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BIOCHEMICAL BULLETIN 



ISSUED QUARTERLY BY THE 

COLUMBIA UNIVERSITY BIOCHEMICAL ASSOCIATION 



PRESS OF 

THE NEW ERA PRINTINQ COMPANY 

LANCASTEK, PA. 



BiocHEMiCAL Bulletin 

Edited, for the Columbia University Biochemical Association, by the 
EDITORIAL COMMITTEE: 

iJuly, igiz-June, 1913) 

ALFRED P. LOTHROP, Chairman, 

PAUL E. HOWE, Secretary, WILLIAM J. GIES, Treasurer, 

WALTER H. EDDY, MAX KAHN, EMILY C. SEAMAN, 

Oct., 1912-July, 191 3 

NELLIS B. FOSTER, ARTHUR KNUDSON, CLAYTON S. SMITH, 

July-September, 19 12 

F. G. GOODRIDGE, EDGAR G. MILLER, Jr., ETHEL WICKWIRE, 

TULA L. HARKEY, H. O. MOSENTHAL, LOUIS E. WISE, 

Oct., 1912-July, 1913 

JOSEPH S. HEPBURN, JACOB ROSENBLOOM, 

JuIy-September, 1912 
ALL OK THE StAFF OF THE BlOCHEMICAL DEPARTMENT OF COLUMBIA UnIVERSITY 



VOLUME II : Nos. 5-8 
1912-1913 



WITH EIGHT PORTRAITS, EIGHT PLATES AND TWO ADDITIONAL 

ILLUSTRATIONS 

LIBRARY 
NnvV YORK 

NEW YORK 

Columbia University Biochemical Association 

1913 

Entered as second-class matter in the Post Ofiice at Lancaster, Pa. 



MEMBERS OF THE COLUMBIA UNIVERSITY 
BIOCHEMICAL ASSOCIATION 

Honorary Members 

PROF, R. H. CHITTENDEN, First Director of ihe Columbia University De- 
partment of Biological (Physiological) Chemistry; Director of the Shef- 
field Scientific School of Yale University 

PROF. HUGO KRONECKER, Director of the Physiological Institute, Uni- 
versity of Bern, Switserland 

PROF. SAMUEL W. LAMBERT, Dean of the Columbia University School of 
Mediane 

DR. JACQUES LOEB, Member of the Rockef eller Institute for Medical Re- 
search; Head of the Department of Experimental Biology 

PROF. ALEXANDER SMITH, Head of the Department of Chemistry, Co- 
lumbia University 

Corresponding Members 

PROF. LEON ASHER, University of Bern, Switserland 

PROF. FILIPPO BOTTAZZI, University of Naples, Italy 

PROF, ROBERT B. GIBSON, University of the Philippines, P. I. 

PROF. VLADIMIR S. GULEVIC, University of Moscow, Russia 

PROF. W. D. HALLIBURTON, King's College, London 

PROF. S. G. HEDIN, University of Upsala, Sweden 

PROF. FREDERICO LANDOLPH, University of La Plata, Argentina 

PROF. A. B. MACALLUM, University of Toronto, Canada 

PROF. D. McCAY, Medical College, Calcutta, India 

PROF. C. A. PEKELHARING, University of Utrecht, Holland 

PROF. S. P. L. SÖRENSEN, Carlsberg Laboratory, Copenhagen, Denmark 

Members Resident in New York City 

Brooklyn Botanic Garden. — C. Stuart Gager. 

College of the City of New York. — Wm. B. Boyd, Louis J. Curtman, 
Benj. G. Feinberg, A. J. Gold färb. 

Columbia University: Departments. — Anatomy: Alfred J. Brown, H. von 
W. Schulte; Bacteriology: James G. Dwyer; Biological Chemistry: Walter H. 
Eddy, Nellis B. Foster, William J. Gies, F. G. Goodridge, Tula L. Harkey, 
Joseph S. Hepburn, Benjamin Horowitz, Paul E. Howe, Max Kahn, Arthur 
Knudson, Alfred P. Lothrop, Edgar G. Miller, Jr., H. O. Mosenthal, Emily C. 
Seaman, Chris. Seifert, Ethel Wickwire, Louis E. Wise; Botany: E. R. Alten- 
burg, C. A. Darling, Fred D. Fromme; Cancer Research: W. H. Woglom; Chem- 
istry: A. M. Buswell, R. P. Calvert, Gustave Egloff, H. L. Fisher, P. W. Punnett, 
A. W. Thomas; Clinical Pathology: Edward Cussler, Peter Irving; Diseases of 
Children: Herbert B. Wilcox; Gynecology: Wilbur Ward; Mediane: T. Stuart 
Hart, I. Ogden Woodruff; Pathology: B. S. Oppenheimer, Alwin M. Pappen- 
heimer; Pharmacology: Charles C. Lieb; Physiology: Russell Burton-Opitz, 
Donald Gordon, Leander H. Shearer, Wm. K. Terriberry; Surgery: Hugh 
Auchincloss, William Darrach, Rolfe Kingsley, Adrian V. S. Lambert, F. T. 
Van Buren, Jr. ; Therapeutics: Maximilian Schulman; University Physician: 

iv 



Members resident in New York (con.) 

Wm. H. McCastline; Vanderbilt Clinic: F. Morris Class, Julius W. Weinstein; 
Zoology: H. B. Goodrich, John D. Haseman, H. J. Muller, Charles Packard. 

Colleges. — Barnard: Helene M. Boas, Ella H. Clark, Ruth S. Finch, Louise 
H. Gregory; College of Pharmacy: Charles W. Ballard; Teachers College: 
Mary G. McCormick, Mrs. A. P. McGowan, Sadie B. Vanderbilt. 

Students. — Graduate: Cora J. Beckwith, Sidney Born, O. C. Bowes, Helen B. 
Davis, Mary C. de Garmo, Frank R. Eider, Louis J. Hirschleifer, Mildred A. Hoge, 
Shojiro Kubushiro, Victor E. Levine, Darwin O. Lyon, W. A. Perlzweig, Edward 
Plaut, Geo. S. Rosenthal, Edward C. Stone, Fred L. Thompson, Jennie A. Walker, 
Charles Weisman, C. A. Wells, Isabel Wheeler. — Teachers College: Anna M. 
Connelly, Ula M. Dow, Ada M. Field, Helen McClure, Alice H. McKinney, 
Elizabeth Rothermel, Mary B. Stark, Helen B. Thompson. — Medical: Louis 
Berman, Ernst Boas, David C. Bull, Will H. Chapman, Robert T. Corry, Calvin 
B. Coulter, Joseph Felsen, Joseph Goldstone, Julius Gottesman, Leon M. Herbert, 
Martin Holzman, Walter F. Hume, Julius Hyman, M. V. Miller, Nathan 
Rosenthal, A. V. Salomon, Harry J. Seiflf, Jacob Shulansky, H. J. Spencer, 
Henry A. Sussman, Wm. W. Tracey, Grover Tracy. 

CoRNELL Univeesity Medical COLLEGE. — Stanley R. Benedict, Ernest D. 
Clark, Robert A. Cooke, Jessie A. Moore, Charles R. Stockard, Geo. W. 
Vandegrift. 

EcLECTic Medical College. — David Alperin. 

Harriman Research Laboratory. — Marston L. Hamlin. 

Hospitals. — Babies': Morris Stark; Bellevue: Edward C. Brenner, Edward 
M. Colie, Jr., Ralph W. Lobenstine; Beth Israel: Charles J. Brim and Alfred A. 
Schwartz; City: Henry H. Janeway, Louis Pine; Flower: Henry L. Weil; 
Flushing: Eimer W. Baker; General Memorial: Clinton B. Knapp; German: H. 
G. Baumgard, Alfred M. Hellman, Melvin G. Herzfeld, Frederick B. Humphries, 
Charles H. Sanford, Fred S. Weingarten; Jewish: Abraham Ravich; Lebanon: 
Samuel Gitlow, M. J. Gottlieb, William Weinberger; Lutheran: Daniel R. Lucas; 
Mt. Sinai: George Baehr, Samuel Bookman, Leo Buerger, Burrill B. Crohn, 
Simon S. Friedman, David J. Kaliski, John L. Kantor, Leo Kessel, Reuben Otten- 
berg, Harry Wessler; TV. Y.: James C. Greenway, Ralph G. Stillman; N. Y. 
Nursery and Child's: Oscar M. Schloss; Presbyterian: Herbert S. Carter, Russell 
L. Cecil, Arthur W. Swann; Roosevelt: J. Buren Sidbury; St. Luke's: Norman 
E. Ditman, Edward C. Kendali, W. S. Schley, Chas. H. Smith. 

Long Island Medical College. — Matthew Steel. 

MoNTEFioRE HoME. — Isidor GrecHwald. 

Museum of Natural History. — Louis Hussakof, Israel J. Kligler. 

N. Y. Aquarium. — Raymond C. Osburn. 

N. Y. Association for Improving the Condition of the Poor. — Donald B. 
Armstrong. 

N. Y. Botanical Garden. — Fred J. Seaver. 

N. Y. City Department of Education. — Boys" High School: Frank T. 
Hughes; Brooklyn Training School: C. A. Mathewson ; Commercial High School: 
' W. J. Donvan, B. C. Gruenberg, Edgar F. Van Buskirk; DeWitt Clinton High 
School: Frank M. Wheat; Eastern District High School: Gertrude S. Burling- 
ham; Girls' High School: Marguerite T. Lee; High School of Commerce: 
Harvey B. Clough, Fred W. Hartwell; Jamaica High School: Ella A. Holmes, 
Charles H. Vosburgh; Manual Training High School: Anna Everson; Morris 



Members resident in New York (con.) 
High School: Charles A. Wirth; Newtown High School: Nellie P. Hewins; 
Wadleigh High School: Helen Gavin, Elsie A. Kupfer, Helen G. Russell, Helen 
S. Watt. 

N. Y. City Department of Health. — Charles F. Bolduan, Alfred F. Hess. 

N. Y. City Normal College. — Beatrix H. Gross. 

N. Y. Eye and Ear Infirmary. — Harold M. Hays. 

N. Y. Milk Committee. — Philip Van Ingen. 

N. Y. Polyclinic Medical School. — Jesse G. M. Bullowa, Mabel C. Little. 

Post Graduate Medical School. — Louis E. Bisch, Arthur F. Chace. 

Pratt Institute. — Grace MacLeod. 

Rockefeller Institute. — Alfred E. Cohn, George W. Draper, Frederic M. 
Hanes, Michael Heidelberger, Gustave M. Meyer. 

Russell Sage Institute of Pathology. — Eugene F. DuBois. 

TuRCK Institute. — Anton R. Rose. 

Vettin School. — Laura I. Mattoon. 

E. V. Delphey, 400 West 57th Street, Manhattan; Leopold L. Falke, 5316 
Thirteenth Avenue, Brooklyn; Mabel P. Fitzgerald, 416 East 6sth Street, Man- 
hattan; Abraham Gross, c/o Arbuckle Sugar Co., Brooklyn; Alfred H. Kropff, 
619 Kent Avenue, Brooklyn. 

Non-Resident Members 

Agnes Scott College (Decatur, Ga.). — Mary C. de Garmo. 

Allegheny General Hospital (Pittsburgh). — James P. McKelvy. 

Carnegie Institution (Cold Spring Harbor, L. I.). — Ross A. Gortner. 

Cornell University (Ithaca). — Jean Broadhurst. 

Drake University Medical School (Des Meines, la.). — E. R. Posner. 

Forest School (Biltmore, N. C). — Homer D. House. 

Iowa University Hospital (Iowa City). — Louis Baumann. 

Isolation Hospital (San Francisco, Cal.). — L. D. Mead. 

Jefferson Medical College (Phila.). — P. B. Hawk, Edward A. Spitzka. 

Johns Hopkins University (Baltimore). — John Howland, W. M. Kraus, 
Burton E. Livingston, Edwards A. Park. 

Lehigh University (Bethlehem, Pa.). — William H. Welker. 

Leland Stanford University (Palo Alto, Cal.). — Hans Zinsser. 

MacDonald College (Quebec). — Kathryn Fisher. 

Mass. Agricultural College (Amherst). — H. D. Goodale. 

New Mexico Agricultural College (State College). — R. F. Hare. 

N. J. Agricultural Experiment Station (New Brunswick). — Carl A. 
Schwarze, Guy West Wilson. 

N. Dakota Agricultural College (Agricultural College). — H. L. White. 

Ohio Agricultural Experiment Station (Wooster). — A. D. Selby. 

Princeton University (N. J.). — E. Newton Harvey. 

Psychopathic Hospital (Boston). — Herman M. Adler. 

Rensselaer Polytechnic Institute (Troy, N. Y.). — Fred W. Schwartz. 

Rochester A and M Institute. — Elizabeth G. Van Hörne. 

Secondary Schools. — Brockport State Normal School (N. Y.) : Ida C. Wads- 
worth; Hermon High School (N. Y.) : Sidney Liebovitz; Indiana State Normal 
School (Terre Haute): Roscoe R. Hyde; Ingleside School (New Milford, 
Conn.) : Mary L. Chase; Knox School (Tarrytown, N. Y.) : Clara G. Miller; 
New Bedford Industrial School (Mass.): Constance C. Hart; North Texas 

vi 



Non-resident members (con.) 

State Normal School (Benton) : Blanche E. Shaffer; Passate High School 
(N. J.) : Hazel Donham, Helene M. Pope; Rochester High School (N. Y.) : 
David F. Renshaw; State Normal School (Truro, N. S.) : Blanche E. Harris. 

Texas A and M College (College Station). — M. K. Thornton. 

Trinity College (Hartford, Conn.). — Max Morse, R. M. Yergason. 

TuLANE University (New Orleans, La.). — Allan C. Eustis. 

U. S. Department of Agriculture (Wash.). — Carl L. Aisberg, W. N. Berg, 
H. E. Buchbinder, William Salant, Clayton S. Smith. 

U. S. Food and Drug Inspection Laboratory (Phila.). — Harold E. Woodward. 

U. S. FooD- Research Laboratory (Phila.). — Joseph S. Hepburn. 

University of Alabama Medical School (Birmingham). — Richard A. Bliss. 

University of California (Berkeley). — William T. Home, 

University of Chicago. — Mathilde Koch. 

University of Georgia Medical School (Atlanta). — ^William D. Cutter. 

University of Illinois (Urbana). — George D, Beal, Isabel Bevier, A. D. 
Emmett. 

University of Indiana (Bloomington). — Clarence E. May. 

University of Kentucky (Louisville). — Mary E. Sweeny. 

University of Manitoba (Winnipeg, Can.). — A. T. Cameron. 

University of Michigan (Ann Arbor). — A. Franklin Shull. 

University of Montana (Missoula). — J. E. Kirkwood. 

University of Pennsylvania (Phila.). — A. N. Richards. 

University of Porto Rico (Las Pietras). — L. A. Robinson. 

University of Tennessee (Memphis). — Edwin D. Watkins. 

University of Texas (Austin). — Mary E. Gearing, Anna E. Richardson. 

University of Toronto (Canada). — Olive G. Patterson. i 

University of Utah (Salt Lake City). — H. A. Mattill. 

University of Wisconsin (Madison). — ^W. H. Peterson. 

Vassar College ( Poughkeepsie, N. Y.). — Winifred J. Robinson. 

Washington State College (Pullman). — Josephine T. Berry, Louise 
McDanell. 

Wesleyan University (Middletown, Conn.). — David D. Whitney. 

West Pennsylvania Hospital (Pittsburgh). — J. Bronfen Brenner, Jacob 
Rosenbloom. 

Williams College (Williamstown, Mass.). — John S. Adriance, 

Yale University (New Haven, Conn.). — Lorande Loss Woodruff. 

Albert H. Allen, Saranac Lake, N. Y. ; Emma A. Buehler, Newark, N. J. ; 
George A. Geiger, West Orange, N. J. ; Edward G. Griffin, Albany, N. Y. ; F. C. 
Hinkel, Utica, N. Y.; Cavalier H. Joüet, Roselle, N. J.; A. E. Olpp, West 
Hoboken, N. J. ; Adeline H. Rowland, Pittsburgh, Pa. ; William A. Taltavall, 
Redlands, Cal. ; David C. Twichell, Saranac Lake, N. Y. 



Vll 



EDITORS OF THE BIOCHEMICAL BULLETIN 

The editorial committee 
with the coUaboration of the members and the 

SPECIAL CONTRIBUTORS: 

DR. JOHN AUER, Rockef eller Institute for Medical Research 

PROF. WILDER D. BANCROFT, Cornell University, Ithaca 

DR. WALTER L. GROLL, Elizabeth Steel Magee Hospital, Pittshurgh, Pa. 

DR. CHARLES A. DOREMUS, 55 W. 53d St., New York City 

DR. ARTHUR W. DOX, Iowa State College Agric. Experiment Station, Arnes 

PROF. JOSEPH ERLANGER, Washington Univ. Medical School, St. Louis 

DR, LEWIS W. FETZER, U. S. Dep't of Agriculture, Washington, D. C. 

PROF. MARTIN H. FISGHER, University of Cincinnati 

DR. MARY LOUISE FOSTER, Smith College, Northampton, Mass. 

PROF. J. E. GREAVES, Utah Agricultural College, Logan 

DR. V. J. HARDING, McGill University, Montreal, Canada 

DR. R. H. M. HARDISTY, McGill University, Montreal, Canada 

DR. J. A. HARRIS, Carnegie Sta. for Exp. Evolution, Cold Spring Harbor, L. I. 

DR. K. A. HASSELBALCH, Einsen Institute, Copenhagen, Denmark 

PROF. G. O. HIGLEY, Ohio Wesleyan University, Delaware 

DR. VERNON K. KRIEBLE, McGill University, Montreal, Canada 

PROF. FRANCIS E. LLOYD, McGill University, Montreal, Canada 

PROF. JOHN A. MANDEL, A''. Y. Univ. and Bellevue Hospital Med. College 

PROF. ALBERT P. MATHEWS, University of Chicago 

PROF. SHINNOSUKE MATSUNAGA, University of Tokyo, Japan 

PROF. LAFAYETTE B. MENDEL, Yale University 

PROF. VICTOR C. MYERS, N. Y. Post-Graduate Med. School and Hospital 

DR. THOMAS B. OSBORNE, Conn. Agric. Experiment Station, New Haven 

DR. AMOS W. PETERS, The Training School, Vineland, N. J. 

PROF. R. F. RUTTAN, McGill University, Montreal, Canada 

DR. E. E. SMITH, 50 East 4ist St., New York City 

DR. A. E. SPAAR, City Hospital, Trincomalee, Ceylon 

PROF. UMETARÖ SUZUKI, University of Tokyo, Japan 

MISS ANNA W. WILLIAMS, University of Illinois, Urbana, III. 

PROF. E. WINTERSTEIN, Polytechnic Institute, Zürich, Switserland 

DR. JULES WOLFF, Pasteur Institute, Paris 



vm 



OFFICERS 

OF THE 

COLUMBIA UNIVERSITY BIOCHEMICAL 

ASSOCIATION 

1912-1913 

HONORARY OFFICERS 

Post President (1910-1912) : 
Prof. Alfred N. Richards, University of Pennsylvania, Phila. 

President: 
Prof. Philip B. Hawk, Jefferson Medical College, Phila. 

Vice Presidents: 
Dr. Herman M. Adler, Psychopathie Hospital, Boston, Mass. 
Prof. Allan C. Eustis, Tulane University, New Orleans, La. 
Miss Olive G. Patterson, Toronto University, Toronto, Can. 
Prof. Winifred J. Robinson, Vassar College, Poughkeepsie, N. Y. 
Prof. Lorande Loss Woodruff, Yale University, New Haven, 
Conn. 

ACTIVE OFFICERS 

President, Dr. Walter H. Eddy. 

Vice President, Prof. Stanley R. Benedict. 

Secretary, Dr. Alfred P. Lothrop. 

Treasurer, Prof. William J. Gies. 

Executive Commiftee — Prof. Stanley R. Benedict, Dr. Wal- 
ter H. Eddy, Dr. Nellis B. Foster, Prof. William J. Gies, Dr. 
Frederic G. Goodridge, Prof. Paul E. Howe and Dr. Alfred 
P. Lothrop. 

Editorial Committee: See the title page. 

ix 



SUMMARY OF CONTENTS: VOL. II, Nos. 5-8. 
No. 5. September, 19 12. 

PAGE 

Ernst ScHxn-ZE. Biography and Bibliography (with portrait). 

Ernst Winterstein. i 

A ReSUME OF THE LiTERATURE ON InOSITE-PhoSPHORIC AciD ("PhYTIN"), 

WITH Special Reference to the Relation of that Substance to 

Plants. Anton Richard Rose 21 

A New Type of Artificial Cell Suitable for Permeability and Other 

BioCHEMicAL Studies. E. Ncwtou Hürvey 50 

On a New Function of the Catalyzer Called Peroxidase and on the 

Biochemical Transformation of Orcin into Orcein. Jules Wolff . ... 53 
Studies of Diffusion through Rubber Membranes : 

1. Preliminary Observations on the Diffusibility of Lipins and Lipin- 

soluble Substances. William J. des SS 

2. Dififusibility of Lipins from Ether through Rubber Membranes into 

Ether. Jacob Rosenbloom 64 

3. Diffusibility of Protein through Rubber Membranes, with a Note on 

the Disintegration of Collodion Membranes by Common Ethyl 
Ether and Other Solvents. William H. Welker 70 

4. The Comparative DiffusibiHty of Various Pigments in Different Sol- 

vents. George D. Beal and George A. Geiger 78 

The Colloidal Nitrogen in the Urine from a Dog with a Tumor of the 

Breast. Max Kahn and Jacob Rosenbloom 87 

General Aspects of Fasting. Paul E. Hozue 90 

The Physico-chemical Basis for the Contraction of Striated Muscle: 

2. Surface Tension. William N. Berg loi 

A Study of Some Protein Compounds. Walter H. Eddy iii 

Effects of Intraperitoneal Injections of Epinephrin on the Partition 

OF Nitrogen in Urine from a Dog. 

Jacob Rosenbloom and William Weinberger. 123 

The Biochemical Society, England. W. D. Halliburton 128 

Proceedings of the Section (II) ON DiETETic Hygiene and Hygienic 

Physiology OF the 15TH International Congress on Hygiene and 

Demography, with Abstracts of Some of the Papers. 

Lafayette B. Mendel, Secretary. 129 
Program of the Proceedings of the Section on Biochemistry Including 

Pharm acology (viii, d), of the 8th International Congress of 

Applied Chemistry. John A. Mandel, Secretary 150 

Proceedings of the Sixth Scientific Meeting of the Columbia Univer- 

siTY Biochemical Association. Alfred P. Lothrop, Secretary 156 

Biochemical News, Notes and Comment: 

General 188 

X 



PAGE 

Columbia University Biochemical Association 200 

Columbia Biochemical Department 201 

Editori ALS : 

Ernst Schulze 205 

Important though Unknown Factors in Nutrition 205 

The Coming of Age of the Babcock Test 207 

Organotherapy 208 

Biochemical Society, England 20g 

"Baustein" or " Construction Unit"? 209 

" Splitting Products " or Cleavage Products ? 209 

A Rare Compliment 210 

X-rays 210 



No. 6. January, 19 13. 

PAGE 

Carl L. Alsberg. Biography and Bibliography (with portrait). H.M.A.. 211 
A Differential Chemical Study of Glucoses from a Case of Pancreatic 

Diabetes. Frederic Landolph 217 

The Detection of Aceto-acetic Acid by Sodium Nitroprussid and Ammonia. 

V. J. Harding and R. F. Ruttan. 223 
Ortho-tolidin as an Indicator for Occult Blood. 

R. F. Ruttan and R. H. M. Hardisty. 225 

Synthetical Properties of Emulsin. Vernon K. Krieble 227 

On the Occurrence of Nicotinic Acid in Rice Bran. 

U. Suzuki and S. Matsunaga. 228 
A Study of the Influence of Cancer Extracts on the Growth of Lupin 

Seedlings. Jacob Rosenhloom 229 

The Biochemistry of the Female Genitalia: 

3. A Quantitative Study of Certain Enzymes of the Ovary, Uterus, and 

Bladder, of Pregnant and Non-pregnant Sheep. 

Thuisco A. Erpf-Lefkovics and Jacob Rosenbloom. 233 

4. On the Absence of Certain Enzymes from the Human Chorion. 

Jacob Rosenbloom. 236 
A Department of Biochemical Research at Vineland, New Jersey. 

Arnos W. Peters. 238 

Biochemistry in New York Twenty Years Ago. E. E. Smith 243 

Immunity in Some of its Biochemical Aspects. Charles Frederick Bolduan. 247 
A Plan for the Organization of the American Biological Society. 

Albert P. Mathews. 261 
Organization of the Federation of American Societies for Experimental 

BlOLOGY, COMPRISING THE AmERICAN PhYSIOLOGICAL SoCIETY, AmERICAN 

Society of Biological Chemists, and American Society for Pharma- 

COLOGY and Experimental Therapeutics. John Auer 269 

Annual Meetings of the Organizations Comprising the Federation of 
American Societies for Experimental Biology : 
I. The American Physiological Society. 

Joseph Erlanger, Acting Secretary. 271 



PAGB 

2. The American Society of Biological Chemists. 

Alfred N. Richards, Secretary. 275 

3. The American Society for Pharmacology and Experimental Thera- 

peutics. John Aner, Secretary 27g 

Meeting of the American Society of Animal Nutrition (American 

Society of Animal Production) . Lewis W. Fetzer 282 

Proceedings of the Eighth Scientific Meeting of the Columbia Univer- 

siTY BiocHEMicAL ASSOCIATION. Alfred P. Lothrop, Secretary ;.. 284 

Folio Microbiologica. C. A. Pekelharing 297 

BiocHEMiCAL Bibliography AND Index. William J. Gies 298 

BiocHEMicAL News, Notes and Comment: 

General 307 

Columbia University Biochemical Association 321 

Columbia Biochemical Department 324 

Editorials : 

New Plan of Quarterly Issue of the Bulletin 329 

Carl L. Aisberg 329 

Stock Poisoning Due to Spoiled Silage. Help ! 330 

Demand for Biological Chemists in the Hospitals 330 

Federation of American Societies for Experimental Biology 331 

Electrons 332 

No. 7. April, 19 13. 

PAGE 

Heinrich Ritthausen (Portrait) 334 

Appreciation. Thomas B. Osborne 335 

Bibliography. Lewis W. Fetzer 339 

Dinner to Professor Chittenden: Testimonial by his Pupils. '94S 349 

Society for Experimental Biology and Medicine: Tenth Anniversary 
Meeting and Dinner. Nineteen O. Three 358 

Methods for the Electrometric Determination of the Concentration of 
Hydrogen Ions in Biological Fluids. K. A. Hasselbalch 367 

A Method for the Determination of Tryptophan Derived from Pro- 
teins. Jesse A. Sanders and Clarence E. May 373 

Physical Chemistry of Muscle Plasma. Filippo Bottazzi 379 

Fasting Studies : IL A Note on the Composition of Muscle from Fast- 
iNG DoGs. H. C. Biddle and Paul E. Howe 386 

SoME Notes on the Form of the Curve of Carbon-dioxide Excretion Re- 
suLTiNG from Muscular Work Following Forced Breathing. 

G. O. Higley. 390 

The Influence of Barometric Pressure on Carbon-dioxide Excretion in 
Man. G. O. Higley 393 

The Relation of Acapniato Shock, andaConsideration of the Mechan- 
iCAL Effects of Artificial Hyper-respiration upon the Circulation. 

Henry H. Janeway and Ephraim M. Ewing. 403 

Cleavage of Pyromucuric Acid by Mold Enzymes. 

Arthur W. Dax and Ray E. Neidig. 407 

Analysis of the Ash of the Castor Bean. Marston Lovell Hamlin 410 

Notes on the Chemical Nature of the " Tannin Masses " in the Fruit 
of the Persimmon. Ernest D. Clark 412 

xü 



PAGE 

HisTON AND iTS Prep ARATION. Walter H. Eddy 419 

DiD VON Wittich Antedate Ostwald in the Definition of Enzyme Action ? 

William N. Berg. 441 

The Biochemical Society, England 446 

Scientific Proceedings of the Columbia University Biochemical Asso- 
ciation. Alfred P. Lothrop, Secretary 452 

Biochemical Bibliography and Index. William J. Gies 470 

Biochemical News, Notes and Comment: 

General 476 

Columbia University Biochemical Association 484 

Editorials : 

Biochemical Society, England 487 

The Bleached Flour Decision 487 

Occupational Diseases in Chemical Trades 488 

The Mathews Plan for an American Biological Society 490 

Antigens 508 



No. 8. July, 19 13. 

PAGE 

An Investigation to Determine the Accuracy of a Modified Meigs 
Method for the Quantitative Determination of Fat in Milk, with 
A Description of an Improved Form of Apparatus. Walter Lewis Croll. 509 

The Occurrence of Arsenic in Soils. /. E. Greaves 519 

Further Notes on the Relationship between the Weicht of the Sugar 
Beet and the Composition of its Juice. 

/. Arthur Harris and Ross Atken Gortner. 524 

Note on the Relationship between Barometric Pressure and Carbon- 
dioxide ExcRETioN IN Man. /. Arthur Harris 530 

The Bleached Flour Decision. Ross Aiken Gortner 532 

Emil Chr. Hansen Fund. 5". P. L. Sörensen 535 

Biological Chemistry in the Philippines. Robert Banks Gibson 536 

Doctorates in Biological Chemistry. Conferred by American Univer- 
sities, 1912-13. P. H. D 538 

Scientific Proceedings of the Columbia University Biochemical Asso- 
ciation. Alfred P. Lothrop, Secretary 541 

Biochemical Bibliography and Index. William J. Gies 559 

Biochemical News, Notes and Comment: 

General 567 

Columbia University Biochemical Association 574 

Columbia Biochemical Department 578 

Editorials : 

Peroxides and Nitrites in Plants 582 

Mathews Plan for the Organization of an American Biological Society. 582 
Crystals 588 

Index: Volume II. (Includes names of authors, and impersonal and per- 
sonal subj ects) 589 

Title Page for Volume II, with Summary of Contents, List of Illus- 

TRATIONS, ETC i-Xvi 

. xiii 



Alphabetic list of authors named in the foregoing summary 

of Contents 

(See author index — page 589 — for additional names of authors of 
abstracts, quotations, comment, etc.) 



Adler, HM, 211 
AuER, J, 269, 279 
Beal, GD, 78 
Berg, WN, ioi, 441 
BiDDLE, HC, 386 
BOLDUAN, CF, 247 

BoTTAzzi, F, 379 
Clark, ED, 412 
Croll, WL, 509 
Dox, AW, 407 
Eddy, WH, III, 419 
Erlanger, J, 271 
Erpf-Lefkovics, TA, 233 
EwiNG, EM, 403 
Fetzer, LW, 282, 339 
Geiger, GA, 78 
GiBSON, RB, 536 
Gies, WJ, 5s, 298, 349, 

358, 470, 559 
GoRTNER, RA, 524, 532 



Greaves, JE, 519 
Halliburton, WD, 128 
Hamlin, ml, 410 
Harding, VJ, 223 
Hardisty, RHM, 225 
Harris, JA, 524, 530 
Harvey, EN, 50 
Hasselbalch, KA, 367 
HiGLEY, GO, 390, 393 
Howe, PE, 90, 386 
Janeway, HH, 403 
Kahn, M, 87 
Kribble, VK, 227 
Landolph, F, 217 
LoTHROP, AP, 156, 284, 

452, 541 
Mandel, JA, 150 
Mathews, AP, 261 
Matsunaga, S, 228 
May, CE, 373 



Mendel, LB, 129 
Neidig, RE, 407 
OsBORNE, TB, 335 
Pekelharing, CA, 297 
Peters, AW, 238 
P. H. D., 538 
Richards, AN, 275 
Rose, AR, 21 

ROSENBLOOM, J, 64, 87, 

123, 229, 233, 236 
Ruttan, RF, 223, 225 
Sanders, JA, 373 
Smith, EE, 243 
Sörensen, SPL, 535 
Suzuki, W, 228 
Weinberger, W, 123 
Welker, WH, 70 
Winterstein, E, i 

WOLFF, J, 53 



XIV 



LIST OF ILLUSTRATIONS 

Eight portraits, eight plates (inserts), and two 
additional illustrations 

No. 5- SEPTEMBER, 1912 

PAGE 

Portrait. Ernst Schulze i 

Plate I. Structure of muscle fibrils (Berg) 107 

Portrait. Paul E. Howe 201 

No. 6. JANUARY, 1913 

Portrait. Carl L. Aisberg 211 

Plate 2. Receptors of three kinds (Bolduan) 254 

No 7. APRIL, 1913 

Portrait. Heinrich Ritthausen 335 

Portrait. Russell H. Chittenden 349 

Engrossed Greetings to Prof. Chittenden by his colleagues of the Governing 

Board of th^^effield Scientific School 351 

Faces of the gol(^medal presented to Prof. Chittenden by the National Insti- 
tute of Social Sciences 353 

Portrait (group). Testimonial dinner to Prof. Chittenden by his pupils, 

Mar. I, 1913 355 

Portrait. Samuel J. Meltzer 359 

Portrait (group). Dinner of the Society for Experimental Biology and 

Medicine, tenth anniversary, Feb. 19, 1913 363 

Plate 3. Apparatus for the electrometic determination of the concentration 

of hydrogen ions (Hasselbalch) 371 

Plate 4. Curve of carbon-dioxide excretion resulting from muscular work 

after forced breathing. (Higley) 390 

Plate 5. Influence of barometric pressure on the excretion of carbon-dioxide 

(Higley) 396 

Plate 6. Micro-Kjeldahl apparatus (Morse) 458 

No. 8. JULY, 1913 

Plate 7. Apparatus for use with the Meigs method for the determination of 

fat in milk (Croll) 517 

Plate 8. Relationship between the weight of the sugar beet and the compo- 
sition of its juice (Harris and Gortner) 526 



XV 



Vol. II September, 1912 No. 5 

Biochemical Bulletin 

Edited, for the Columbia University Biochemical Association, by the 
EDITORIAL COMMITTEE: 
ALFRED P. LOTHROP, Chöirman, 
PAUL E. HOWE, Secretary, WILLIAM J. GIES, Treasurer, 

WALTER H. EDDY, EDGAR G. MILLER, JR., 

NELLIS B FOSTER, HERMAN O. MOSENTHAL, 

FREDERIC G. GOODRIDGE, JACOB ROSENBLOOM, 

TULA L. HARKEY, EMILY C. SEAMAN, 

JOSEPH S. HEPBURN, CLAYTON S. SMITH, 

ARTHUR KNUDSON, ETHEL WICKWIRE, 

all of the Staff of the Biochemical Department of Columbia University. 

CONTENTS 

PAGB 

Ernst Schulze. Biography and Bibliography (with portrait) Ernst Winterstein i 

A ReSUME of the LiTERATURE OX InOSITE-PhOSI'HORIC AcID (" PhYTIN "), WITH 

Special Reference to the Relation of that Substaxce to Plants. 

Anton Richard Rose 21 
A New Type of Artificial Cell Suitable für Permeability and other Bio- 
chemical Studies. E. Newton Harvey . 50 

On a New Function of the Catalyzer Called Peroxidase and on the Bio- 
chemical Transformation of Orcin into Orcein. ßiles Wolff. 53 

Studies of Diffusion Through Rubber Membranes : 

i. Preliminary observations on the diffusibility of lipins and lipin-soluble sub- 

stances. Williajn J. Gies 55 

2. Diffusibility of lipins from ether through rubber membranes into ether. 

Jacob Rosenbloom 64 

3. Diffusibility of protein through rubber membranes, with a note onthe dis- 

integration of collodion membranes by common ethyl ether and other 
solvents. William H. Welker 7° 

4. The comparative diffusibility of various pigments in different solvents. 

George D. Beal and George A. Geiger 78 
The Colloidal Nitrogen in the Urine from a Dog with a Tumor of the 

Breast. Max Kahn and Jacob Roseitbloom 87 

General Aspects of Fasting. Paul E. Hoive 90 

The "Physico-Chemical Basis for the Contraction of Striated Muscle: 

2. Surface tension (with plate i). William N. Berg lOi 

A Study of some Protein Compounds. Walter H. Eddy iii 

Effects of Intraperitoneal Injections of Epinephrin on the Partition of 

Nitrogen IN Urine from A Dog. Jacob Rosenbloom and William Weinberger 123 
The Biochemical Society, England. W. D. Halliburton 128 

PrOCEEDINGS of the SeCTION (II) ON DiETETIC HYGIENE AND HYGIENIC PhYSI- 

ology of the 15TH International Congress on Hygiene and Demog- 

raphy, with Abstracts of some of the Papers. 

Lajayette B. Mendel, Secretary 129 
Program of the Proceedings of the Section on Biochemistry Including 

Pharmacology (viii, d), of the Sth International Congress of Applied 

Chemistry. John A. Mandel, Secretary 150 

Proceedings of the Sixth Scientific Meeting of the Columbia University 

Biochemical Association. Alfred P. Lothrop, Secretary 156 

Biochemical News, Notes and Comment 188 

Editorials 205 



NEW YORK 

Columbia University Biochemical Association. 

Entered as second-class matter in the Post Office at Lancaster, Pa. 



Honorary Members of the Columbia University Biochemical 

Association 

PROF. R. H. CHITTENDEN, First Director of the Columbia University De- 
partment of Biological (Physiological) Chemistry; Director of the Shef- 
field Scientific School of Yale University 

PROF. SAMUEL \V. LAMBERT, Dean of the Columbia University School of 
Medicine 

PROF. ALEXANDER SMITH, Head of the Department of Chemistry, Co- 
lumbia University 

Editors of the Biochemical Bulletin 

EDITORIAL COMMITTEE 

(See names on the outside of this cover) 

CONTRIBUTING EDITORS OF THE BIOCHEMICAL BULLETIN 

PROF. LEON ASHER, University of Bern, Switserland 

PROF. FILIPPO BOTAZZI, University of Naples, Italy 

PROF. W. D. HALLIBURTON, King's College, London 

PROF. S. G. HEDIN, University of Upsala, Sweden 

PROF. FREDERICO LANDOLPH, University of La Plata, Argentina 

PROF. C. A. PEKELHARING, University of Utrecht, Holland 

DR. S. P. L. SÖRENSEN, Carlsberg Laboratory, Copenhagen, Denmark 

SPECIAL CONTRIBUTORS TO THE CONTENTS OF VOLUMES 

I AND II 

PROF. WILDER D. BANCROFT, Cornell University, Ithaca 

DR. CHARLES A. DOREMUS, 55 W. 52d St., New York City 

PROF. MARTIN H. FISCHER, University of Cincinnati 

DR. MARY LOUISE FOSTER, Smith College, Northampton, Mass. 

PROF. FRANCIS E. LLOYD, McGill University, Montreal, Canada 

PROF. JOHN A. MANDEL, A^. Y. Univ. and Bellevue Hospital Med. College 

PROF. ALBERT P. MATHEWS, University of Chicago 

PROF. LAFAYETTE B. MENDEL, Yale University 

PROF. VICTOR C. MYERS, N. Y. Post-Graduate Med. School and Hospital 

DR. E. E. SMITH, 50 Last 4ist St., New York City 

DR. A. E. SPAAR, City Hospital, Trincomalee, Ceylon 

MISS ANNA W. WILLIAMS, University of Illinois, Urbana, III. 

PROF. E. WINTERSTEIN, Polytechnic Institute, Zürich, Switserland 

DR. JULES WOLFF, 26 Rue Dutot, Paris 

ASSOCIATE EDITORS 

DAVID ALPERIN, Eclectic Medical College 

EDGAR ALTENBURG, Department of Botany, Columbia University 

HUGH AUCHINCLOSS, Department of Surgery, Columbia University 

GEORGE BAEHR, Mount Sinai Hospital 

ELMER W. BAKER, Flushing Hospital 

CHARLES W. BALLARD, College of Pharmacy, Columbia University 

HANS G. BAUMGARD, German Hospital Dispensary 

CORA J. BECKWITH, Department of Zoology, Columbia University 



Associate editors (continued) 
STANLEY R. BENEDICT, Cornell University Medical College 
LOUIS E. BISCH, Manhattan State Hospital 
HELENE M. BOAS, Barnard College, Columbia University 
CHARLES F. BOLDUAN, Health Department of New York City 
SAMUEL BOOKMAN, Mount Sinai Hospital 
SIDNEY BORN, Department of Chemistry, Columbia University 
WILLIAM BALLANTINE BOYD, College of the City of New York 
EDWARD C. BRENNER, Bellevue Hospital 

JACOB J. BRONFENBRENNER, Rockefeiler Institute for Medical Research 
LEO BUERGER, Mt. Sinai Hospital 

JESSE G. M. BULLOWA, New York Polyclinic Medical School 
GERTRUDE S. BURLINGHAM, Rastern District High School, Brooklyn 
RUSSELL BURTON-OPITZ, Department of Physiology, Columbia University 
HERBERT S. CARTER, Presbyterian Hospital 
RUSSELL L. CECIL, Presbyterian Hospital 

ARTHUR F. CHACE, New York Post-Graditate Medical School 
ERNEST D. CLARK, Cornell University Medical College 
ALFRED E. COHN, Rocke feller Institute for Medical Research 
EDWARD M. COLIE, Jr., Bellevue Hospital 
BURRILL B. CROHN, Mt. Sinai Hospital 
LOUIS J. CURTMAN, College of the City of New York 
EDWARD CUSSLER, Department of Clinical Pathology, Columbia University 
CHESTER A. DARLING, Department of Botany, Columbia University 
WILLIAM DARRACH, Department of Surgery, Columbia University 
NORMAN E. DITMAN, St. Luke's Hospital 
GEORGE DRAPER, Hospital of the Rockef eller Institute 
BENJAMIN G. FEINBERG, College of the City of New York 
HARRY L. FISHER, Department of Chemistry, Columbia University 
SIMON S. FRIEDMAN, Mt. Sinai Hospital 
C. STUART GAGER, Brooklyn Botanic Garden 
HELEN GAVIN, Wadleigh High School 
SAMUEL GITLOW, Lebanon Hospital Dispensary 
A. J. GOLDFARB, College of the City of New York 
DONALD GORDON, Department of Physiology, Columbia University 
MARK I. GOTTLIEB, Fordham University 
ISIDOR GREENWALD, Montefiore Home Laboratory 
LOUISE HOYT GREGORY, Barnard College, Columbia University 
ABRAHAM GROSS, Arbuckle Sugar Co., Brooklyn 
BEATRIX H. GROSS, N. Y. City Normal College 
BENJAMIN C. GRUENBERG, Brooklyn Commercial High School 
MARSTON L. HAMLIN, Harriman Research Laboratory, Roosevelt Hospital 
FREDERIC M. HANES, Rockef eller Institute for Medical Research 
JOHN D. HASEMAN, Department of Zoology, Columbia University 
HAROLD M. HAYS, New York Eye and Ear Infirmary 
MICHAEL HEIDELBERGER, Rockefeller Institute for Medical Research 
ALFRED M. HELLMAN, German Hospital 
MELVIN G. HERZFELD, German Hospital 
ALFRED F. HESS, Health Department of New York City 
NELLIE P. HEWINS, Newtown High School, L. I. 
ELLA A. HOLMES, Jamaica High School, L. I. 



Associate editors (continued) 

FRANK T. HUGHES, Boys High School, Brooklyn 

FREDERICK B. HUMPHRIES, Gertnan Hospital 

LOUIS HUSSAKOF, American Museum of Natural History 

PETER IRVING, Department of Clinical Pathology, Columbia University 

HENRY H. JANEWAY, City Hospital, New York 

CAVALIER H. JOÜET, Roselle, N. J. 

DAVID J. KALISKI, Mt. Sinai Hospital 

JOHN L. KANTOR, Mt. Sinai Hospital 

EDWARD C. KENDALL, St. Lukc's Hospital 

LEO KESSEL, Mt. Sinai Hospital 

ROLFE KINGSLEY, Department of Surgery, Columbia University 

ISRAEL J. KLIGLER, American Museum of Natural History 

CLINTON B. KNAPP, General Memorial Hospital 

ALFRED H. KRÖPFE, Hoffman and Kropff Chemical Co., Brooklyn 

ELSIE A. KUPFER, Wadleigh High School 

ADRIAN VAN S. LAMBERT, Department of Surgery, Columbia University 

MARGUERITE T. LEE, Girls High School, Brooklyn 

CHARLES C. LIEB, Department of Pharmacology, Columbia University 

MABEL C. LITTLE, New York Polyclinic Hospital 

RALPH W. LOBENSTINE, Bellevue Hospital 

DANIEL R. LUCAS, St. Joseph's Hospital 

CHESTER A. MATHEWSON, Brooklyn Training School for Teachers 

LAURA I. MATTOON, Vcttin School, i6o W. 74th Street 

WILLIAM H. McCASTLINE, University Physician, Columbia University 

MARY G. McCORMICK, Teachers College, Columbia University 

MRS. ELLEN BEERS McGOWAN, Teachers College, Columbia University 

GUSTAVE M. MEYER, Rockefeiler Institute for Medical Research 

JESSIE A. MOORE, Loomis Laboratory, Cornell University Medical College 

HERMANN J. MULLER, Cornell University Medical Collegs 

B. S. OPPENHEIMER, Department of Pathology, Columbia University 

RAYMOND C. OSBURN, New York Aquarium 

REUBEN OTTENBERG. Mt. Sinai Hospital 

CHARLES PACKARD, Department of Zoology, Columbia University 

ALWIN M. PAPPENHEIMER, Department of Pathology, Columbia University 

F. W. PUNNETT, Department of Chemistry, Columbia University 

ABRAHAM RAVICH, Jewish Hospital, Brooklyn 

ANTON R. ROSE, Department of Chemistry, Columbia University 

CHARLES H. SANFORD, German Hospital 

WINFIELD S. SCHLEY, St. Luke's Hospital 

OSCAR M. SCHLOSS, New York Nursery and Chlld's Hospital 

MAX SCHULMAN, Department of Applied Therapeutics, Columbia University 

H. VON W. SCHULTE, Department of Anatomy, Columbia University 

FRED J. SEAVER, Netv York Botanical Garden 

LEANDER H. SHEARER, Department of Physiology, Columbia University 

JAMES B. SIDBURY, Roosevelt Hospital 

MORRIS STARK, Babies Hospital 

MATTHEW STEEL, Long Island Medical College 

RALPH G. STILLMAN, Nezv York Hospital 

CHARLES R. STOCKARD, CorncU Univcrsilv Medical College 

ARTHUR W. SWANN, Presbyterian Hospital 



Associate editors (continued) 
WM. K. TERRIBERRY, Department of Physiology, Columbia University 
F. T. VAN BEUREN, Jr., Department of Surgery, Columbia University 
GEORGE W. VANDEGRIFT, Cornell University Medical College 
SADIE B. VANDERBILT, Teachers College, Columbia University 
CHARLES H. VOSBURGH, Jamaica High School 
WILBUR WARD, Department of Gynecology, Columbia University 
HELEN S. WATT, Wadleigh High School 
WILLIAM WEINBERGER, Lebanon Hospital 
FRED S. WEINGARTEN, German Hospital 

JULIUS W. WEINSTEIN, Vanderbilt Clinic, Columbia University 
HARRY WESSLER, Mt. Sinai Hospital 

H. B. WILCOX, Department of Diseases of Children, Columbia University 
LOUIS E. WISE, Standard Varnish Works, Staten Island, N. Y. 
WILLIAM H. WOGLOM, Dep't. of Cancer Research, Columbia University 
I, OGDEN WOODRUFF, Department of Mediane, Columbia University 
(Locol members of the Columbia University Biochemical Association) 

ASSISTANT EDITORS 

HERMAN M. ADLER, Psychopathie Hospital, Bos.ton, Mass. 

JOHN S. ADRIANCE, Williams College, Williamstozvn, Mass. 

CARL L. ALSBERG, Bureau of Plant Industry, U. S. Dep't. of Agriculture 

D. B. ARMSTRONG, Massachusetts Institute of Technology, Boston 

LOUIS BAUMANN, University Hospital, Iowa City, Iowa 

GEORGE D. BEAL, University of Illinois, Urbana, III. 

WILLIAM N. BERG, Bureau of Animal Industry, U. S. Dep't of Agriculture 

JOSEPHINE T. BERRY, State College, Pullman, Washington 

ISABEL BEVIER, University of Illinois, Urbana, III. 

A. RICHARD BLISS, Birmingham Medical College, Birmingham, Ala. 

JEAN BROADHURST, Cornell University, Ithaca, N. Y. 

WILLIAM D. CUTTER, Medical College of Georgia, Augusta, Ga. 

A. D. EMMETT, University of Illinois, Urbana, III. 

ALLAN C. EUSTIS, Tulane University, Nezv Orleans, La. 

KATHARINE A. FISHER, MacDonald College, Quebec, Canada 

MARY E. GEARING, University of Texas, Austin, Texas 

GEORGE A. GEIGER. Marcus Hook, Fa. 

H. D. GOODALE, Carnegie Sta'n. for Exp. Evolut'n, Cold Spring Harhor, L. I. 

R. A. GORTNER, Carnegie Sta'n for Exp. Evolu'tn, Cold Spring Harbor, L. I. 

R. F. HARE, New Mex. Coli, of Agric. and Mech. Arts, Agric. College, N. M. 

E. NEWTON HARVEY, Princeton University, Princeton, N. J. 

BLANCHE R. HARRIS, State Normal School, Truro, Nova Scotia 

CONSTANCE C. HART, New Bedford Industrial School, New Bedford, Mass. 

P. B. HAWK, JeffersoH Medical College, Philadelphia 

WILLIAM T. HÖRNE, University of California, Berkeley, Cal. 

HOMER D. HOUSE, Forest School, Bilfmore, N. C. 

J. E. KIRKWOOD, University of Montana, Missoula, Mont. 

MATHILDE KOCH, University of Chicago, Chicago, III. 

W. M. KRAUS, Johns Hopkins Medical School, Baltimore, Md. 

SIDNEY LIEBOVITZ, Hermon High School, Hermon, N. Y. 

BURTON E. LIVINGSTON, Johns Hopkins University, Baltimore, Md. 

J. P. McKELVY, Allegheny General Hospital, Pittsburgh, Pa. 

H. A. MATTILL, University of Utah, Salt Lake City, Utah 

CLARENCE E. MAY, Indiana University, Bloomington, Ind. 



Assistant editors (continued) 

L. D. MEAD, Isolation Hospital, San Francisco, Cal. 

CLARA G. MILLER, Knox School, Tarrytczvn, N. Y. 

MAX W. MORSE, Trinity College, Hartford, Conn. 

EDWARDS A. PARK, Johns Hopkins Medical School 

OLIVE G. PATTERSON, Toronto University, Toronto, Canada 

W. H. PETERSON, University of Wisconsin, Madison, Wis. 

E. R. POSNER, Drake University Medical School, Des Maines, la. 

DAVID F. RENSHAW, West High School, Rochester, N. Y. 

ALFRED N. RICHARDS, University of Pennsylvania^ Philadelphia 

ANNA E. RICHARDSON, University of Texas, Austin 

WINIFRED J. ROBINSON, Vassar College, Poughkeepsie, N. Y. 

WILLIAM SALANT, Bureau of Chemistry, U. S. Department of Agriculture 

CARL A. SCHWARZE, N. J. Agricultural Experiment Station, New 

Briuiswick 
FREDERICK W. SCHWARTZ. Rensselaer Polytechnic Institute, Troy, N. Y. 
A. D. SELBY, Ohio Agricultural Experiment Station, Wooster, Ohio 
BLANCHE E. SHAFFER, North Texas State Normal School, Benton, Texas 
A. FRANKLIN SHULL, University of Michigan, Ann Arbor, Mich. 
EDWARD A. SPITZKA, Jcfferson Medical College, Philadelphia 
EDWARD C. STONE, Trinity College, Hartford, Conn. 
MARY E. SWEENY, University of Kentucky, Lexington, Ky. 
WILLIAM A. TALTAVALL, Redlands, Cal. 

IDA C. WADSWORTH, Brockport State Normal School, Brockport, N. Y. 
WILLIAM H. WELKER, Red Hill, Pa. 

DAVID D. WHITNEY, Wesleyan University, Middletown, Conn. 
LORANDE LOSS WOODRUFF, Yale University, New Haven, Conn. 
HAROLD E. WOODWARD, U. S. Food and Drug Inspection Lahoratory, 

Philadelphia 
HANS ZINSSER, Leland Stanford University, Palo Alto, Cal. 
{Non-resident members of the Columbia University Biochentical Association) 




^r^^'i^ ^^>^^^Sz^ 



BiocHEMiCAL Bulletin 



Volume II SEPTEMBER, 191 2 No. 5 



IN MEMORIAM 

ERNST SCHULZE 
Born July 31, 1840 Died June 15, 19 12 

The death o£ Ernst Schulze is an irreparable loss to biology. 
Wherever the biochemistry of plants is appreciated, Schulze's death 
causes profound sorrow. Schulze was one of the founders of our 
present exact biochemical investigation. His researches in phyto- 
chemistry are classical and they have been charged with funda- 
mental ideas that continue to influence research in this great field. 

Dr. Ernst Schulze, professor of agricultural chemistry in the 
Eidgenössischen Technischen Hochschule at Zürich, was born July 
31, 1840, in the hamlet of Bovenden, near Göttingen. In 1858 
Schulze studied chemistry under Wöhler in Göttingen and also spent 
a Semester with Bunsen in Heidelberg. In 1861 he was assistant 
to Lehman, and subsequently to Geuther, at the Chemical Institute 
in Jena. His scientific activity began at the Agricultural Experi- 
ment Station in Weende, under the direction of Henneberg. In 
1871 Schulze was appointed director of the newly founded Agri- 
cultural Experiment Station in Darmstadt. Even while he was at 
Weende, his ability had attracted the attention of the Eidgenös- 
sischen institution. In June, 1872, he was called to Zürich, where 
his activities continued fruit fully for forty years. 

Schulze's first important research was published with the collab- 
oration of his friend Märcker, in 1870, in the Journal für Land- 
zmrtschaft. In this paper it was shown that the principles of pro- 



2 Ernst Schuhe [Sept. 

tein metabolism, as they were stated by Voit on the basis of experi- 
ments on carnivorous animals, applied to the ruminants as well. 
In Zürich, Schulze brought his researches in animal physiology to 
an end by a thorough investigation of wool-fat. He succeeded in 
preparing typical cholesterol in a pure State and in isolating an 
isomer, isocholesterol. 

Since 1872 Schulze had concerned himself exclusively with phy- 
tochemical research ; and forty years of activity in this field f ortified 
the conclusion that plants and animals contain the same classes of 
substances and that the chemical composition of animals is in many 
ways identical with that of plants. 

Schulze developed new methods for the quantitative determina- 
tion of nitrogenous substances and showed how to separate them in 
pure forms from the complex mixtures in plant Juices and extracts. 
With his collaborators Schulze made classical discoveries of the fol- 
lowing nitrogenous Compounds and, by masterly methods, estab- 
lished their Constitution : 

Glutamin, an amide of glutamic acid ; 

Arginin, guanido-ot-aminovalerianic acid ; 

Phenylalanin, yS-phenyl-ct-aminopropionic acid ; 

Vernin (identical with the guanosin subsequently obtained by 
Levene from nucleic acid) ; 

Stachydrin, the dimethylbetain of a-prolin ; 

Lupinin, an alkaloid from lupins. 

Schulze found the following nitrogenous substances in different 
plant materials and studied their role in plant metabolism: amino- 
valerianic acid, leucin, isoleucin, prolin, glutamin, asparagin, Phenyl- 
alanin, tyrosin, arginin, histidin, lysin, vicin, convicin, xanthin, hy- 
poxanthin, guanidin, vernin, allantoin, cholin, betain, trigonellin, 
and stachydrin. Schulze was working with the betains during his 
last illness but, unfortunately, he was unable to complete this re- 
search. Considerable interest was aroused by his discoveiy of the 
presence of allantoin in plants. 

Schulze was the first to make a successful investigation of phyto- 
lecithins ( Phosphatids) and their cleavage products. He found 
that the lecithin in many seeds can be extracted only with hoiling 
alcohol. For this reason he believed that lecithin exists in such 



1912] ■ Ernst Winterstein 3 

seeds in some sort of combination with proteins. In this connection 
he investigated the plant cholesterols, or phytosterols. 

Schulze then began his thorough studies of the carbohydrates 
and nitrogen-free reserve materials in plants. A paper entitled: 
"Untersuchungen über die stickstofffreien Reservestoffe der Samen 
von Lupinus Intens und über die Umwandlung derselben während 
des Keimungsprozesses," was given a prize by the Königlichen 
Gesellschaft der Wissenschaften in Göttingen. 

In this connection Schulze studied the constituents of cell mem- 
branes in plants. He showed that the walls of various plant-cells 
contain carbohydrates which resemble cellulose to a certain extent 
but differ from it by dissolving easily in warm dilute Solutions of 
acids and alkalis. These cell-wall constituents proved to be xylans, 
arabans, galactans, and mannans. They play the part of food 
reserves in seeds. Schulze called them " hemi-celluloses." He 
showed, further, that ordinary cellulose on hydrolysis yields other 
glucoses besides dextrose. Stachys tubers were found to contain 
stachyose, a tetrasaccharid. All these researches yielded data and 
experience that proved useful to Schulze in his discussions and de- 
velopment of analytical methods for phytochemical research. 

The role of asparagin and glutamin in the protein metabolism 
and synthesis in plants greatly interested Schulze to the end of his 
life. Although he was not able fully to explain the process of 
protein synthesis, he made fundamental contributions to the subject. 
He clarified our knowledge of protein metabolism in seedlings. 
What chemist or biologist has not heard of the investigations which 
were begun in 1876, and whose results were usually published in Prus- 
sian agricultural year books and also in the Zeitschrift für physio- 
logische Chemie? Even in his second paper on the subject, in 1878, 
Schulze showed the importance of the characteristic composition of 
etiolated seedlings and their high asparagin content. He concluded 
from his observations that the protein decomposition products do 
not persist, in seedlings, in the proportions in which they were 
originally produced from protein, hut, that after such protein cleav- 
age, these nitro genotis suhstances seem to he changed for the most 
part into asparagin. 

Schulze prepared a great many plant proteins and studied their 



4 Ernst Schulze [Sept 

decomposition products. After his pupils, working outside our 
Institute, had taken an active part in the study of protein metabolism 
in seedlings, and after it had been shown in our laboratory that 
protein decomposition in seedlings is an enzymic process, Schulze 
came to the following general conclusions regarding the protein 
transformations in seedlings: Asparagin is formed in seedHngs at 
the expense of proteins and arises from the same material in etio- 
lated young green plants, and in young leaves and shoots; arginin 
also results in seedlings from direct decomposition of protein. 
Perhaps the individual amids arise in the leaf-buds in the propor- 
tions of their production from protein by hydrolysis with acid^ 
and other agcnts outside the organism, but probably with the differ- 
ence that, in the leaf-buds, neither aspartic acid nor glutamic acid 
is produced, the amids of these amino acids, viz., asparagin and 
glutamin, resulting instead. Amino acids, however, do not occur 
in plants in such proportions, since they are consumed in the plant 
metabolism, some more rapidly, it seems, than others. The accu- 
mulation of asparagin in seedlings is caused by the formation of 
this amide from other products (amino acids) of the trans formation 
of protein. One of the best arguments for this conception is the 
Observation, made by Schulze in his experiments and repeatedly em- 
phasized in his papers, that in many cases asparagin is produced 
abundantly even after the processes of protein decompositions in 
the plant have ceased. 

An admirable outcome of Schulze's investigations is his great 
compilation on the composition of cultivated plants, where he re- 
views briefly the methods of research and gives abundant data on 
the chemical constituents of these plants. 

The later years of Schulze's life were spent in close retirement 
because of a serious and long standing eye-disease that prevented 
him from appearing in public. He lived, at the end, only for his 
science and for his family. His colleagues often wondered how, 
with his weak eyes, he was able to do any experimental work what- 
ever. It was pathetic to see with what extreme care and patience. 
he had to tax himself in order to proceed with his work. 

When Schulze celebrated his seventieth birthday, two years ago, 
we all hoped that the twilight of his life might be long and happy, 



I9I2] Ernst IV int erst ein 5 

but in vain, for pitiless death took him from iis. His pupils mourn, 
a beloved friend and guide; and science, a distinguished inves- 
tigator.^ 

Ernst Winterstein. 

Agriculturchemischen Laboratorium 

der Eidgenössischen Technischen 

Hochschule, Zürich. 

PUBLIICATIONEN VON PROF. DR. E. SCHULZE 

I. In den " Landwirtschaftlichen Versuchsstationen " 

Ueber die Elementarzusammensetzung der tierischen Fette, insbeson- 
dere der Fette vom Schaf, vom Rind und vom Schwein. E. 
Schulze und A. Reinecke. 9: 97-119 (1867). 

Ueber die sensiblen Stickstoff. Einnahmen und Ausgaben des voll- 
jährigen Schafes. E. Schulze und M. MÄRCKER. 11:201(1869). 

Ueber die Zusammensetzung und die Verdaulichkeit des im Wiesenheu 
enthaltenen Fettes. E. Schulze. 15: 81-90 (1872). 

Beiträge zur Kenntnis des Nährwerts und der Zusammensetzung der 
Rüben. E. Schulze. 15: 170-181 (1872). 

Zur Frage über die Verdauung des Heufetts. E. Schulze. 16 : 

329-335 (1873)- 
Notiz über den Aspargingehalt von Lupinen Keimlingen. E. Schulze 

und W. Umlauft. 18: 1-3 (1875). 
Ueber die stickstoffhaltigen Bestandteile der Futter-Rüben. E. 

Schulze und A. Urich. 18: 296-324 (1875). 
Notiz betreffend das Vorkommen des Betains in den Futter-Rüben. 

E. Schulze und A. Urich. 18: 409 (1875). 
Ueber Schwefelsäurebildung in den Keimpflanzen, E. Schulze. 19 : 

172-176 (1876). 
Einige Bemerkungen über die Sachsse-Kormannsche Methode zur Be- 
stimmung des in Amid-Form vorhandenen Stickstoffs. E. 

Schulze. 20: 1 17-123 (1877). 
Ueber die stickstoffhaltigen Bestandteile der Futterrüben. E. Schulze 

und A. Urich. 20: 194-245 (1877). 
Ueber den Gehalt der Kartoffelknollen an Eiweissstoffen und an 

Amiden. E. Schulze und J. Barbieri. 21: 63-92 (1878). 

* The foregoing biographical communication was translated from Prof. E. 
Winterstein's manuscript, in German, by Dr. Ernest D. Clark. Prof. Winter- 
stein's manuscript of the appended bibliography is reproduced verbatim. [Ed.] 



6 Ernst Schulze [Sept 

Ueber ein neues Glukosid (Bestandteil von Lupinus luteus). E. 
Schulze und J. Barbieri. 24: i-ii (1880). 

Ueber das Vorkommen von Leucin und Tyrosin in den Kartoffelknol- 
len. E. Schulze und J. Barbieri. 24: 167-169 (1880). 

Ueber die Bestimmung der Eiweissstoffe und der nicht eiweissartigen 
Stickstoffverbindungen in den Pflanzen. E. Schulze. 24: 358- 

365 (1880); 25: lyz--^?^ (1880). 

Zur Bestimmung der Eiweissstoffe und der nicht eiweissartigen Stick- 
stoffverbindungen in den Pflanzen. E. Schulze und J. Barbieri. 
26:213-283 (1881). 

Neue Beiträge zur Kenntnis der stickstoffhaltigen Bestandteile der 
Kartoffelknollen. E. Schulze und E. Eugster. 27 : 357-373 
(1882). 

Zur quantitativen Bestimmung der Eiweissstoffe und der nicht eiweiss- 
artigen Stickstoffverbindungen in den Pflanzen. E. Schulze. 27 : 
449-465 (1882). 

Ueber das Vorkommen von Hypoxanthin im Kartoffelsaft. E. 
Schulze. 28: 111-115 (1883). 

Ueber das Glutamin. E. Schulze und E. Bosshard. 29: 295-307 
(1883). 

Zur quantitativen Bestimmung des Asparagins, des Glutamins und des 
Ammoniaks in den Pflanzen. E. Schulze und E. Bosshard. 29 : 
399-412 (1883). 

Zur Kenntnis der Methoden, welche zur Bestimmung der Amide in 
Pflanzenextrakten verwendbar sind. E. Schulze. 30 : 459-467 
(1884). 

Ueber einige Bestandteile des Emmentaler Käses. B. Rose und E. 
Schulze. 31: 115-137 (1885). 

Ueber das Vorkommen von Glutamin in den Zuckerrüben und über das 
optische Verhalten desselben. E. Schulze und E. Bosshard. 
32: 129-136 (1887). 

Untersuchungen über die stickstoffhaltigen Bestandteile einiger Rauh- 
futterstoffe. E. Schulze, E. Steiger und E. Bosshard. 33: 
8^123 (1887). 

Ueber die Methoden, welche zur quantitativen Bestimmung der stick- 
stoffhaltigen Pflanzenbestandteile verwendbar sind. E. Schulze. 

33: 124-145 (1887). 
Ueber das Vorkommen von Rohrzucker in unreifen Kartoffelknollen. 

E. Schulze und Th. Seliwanow. 34: 403 (1887). 
Ueber den Nachweis von Rohrzucker in vegetabilischen Substanzen. 

E. Schulze. 34: 408-413 (1887). 



I9I2] Ernst Winterstein 7 

Ein Beitrag zur Erklärung der Veränderungen, welche die stickstoff- 
haltigen Bestandteile eingesäuerter Grünfutterstoffe erleiden. E. 
Schulze. 35: 195-208 (1888). 

Ueber die Zersetzung von Proteinstoffen in verdunkelten grünen 
Pflanzen. E. Schulze und E. Kisser. 36: 1-8 (1889). 

Ueber das Vorkommen eines unlöslichen, Schleimsäure gebenden 
Kohlenhydrats in Rotklee und Luzerne- Pflanzen. E. Schulze 
und E. Steiger. 36: 9-13 (1889). 

Untersuchungen über die stickstofffreien Reservestoffe der Samen von 
Lupinus luteus und über die Umwandlungen derselben während 
des Keimungsprozesses. E. Schulze und E. Steiger. 36 : 391- 
476 (1889). 

Untersuchungen über die chemische Zusammensetzung einiger Legumi- 
nosen-Samen. E. Schulze, E. Steiger und W. Maxwell. 39 : 
269 (1891). 

Ueber einige Bestandteile der Wurzelknollen von Stachys tuberifera. 
A. VON Planta und E. Schulze. 40: 277-298 (1892). 

Bestimmung des Stachyose-Gehalts der Wurzelknollen von Stachys 
tuberifera. A. von Planta und E. Schulze. 41 : 123-129 ( 1892) . 

Zur Kenntnis der in den Leguminosensamen enthaltenen Kohlenhy- 
drate. E. Schulze. 41:207-229 (1892). 

Ueber den Lecithingehalt einiger vegetabilischer Substanzen. E. 
Schulze und S. Frankfurt. 43: 307-318 (1894). 

Untersuchungen über die zur Klasse der stickstoffhaltigen organischen 
Basen gehörenden Bestandteile einiger landwirtschaftlich benutz- 
ter Samen, Oelkuchen und Wurzelknollen, sowie einiger Keim- 
pflanzen. E. Schulze in Verbindung mit S. Frankfurt und E. 
Winterstein. 46: 23-77 (1896). 

Zur Kenntnis der stickstoffhaltigen Bestandteile junger grüner Pflanzen 
von Vicia sativa. E. Schulze. 46: 383-397 (1896), 

Ueber das Vorkommen von Arginin in den Wurzeln und Knollen ein- 
iger Pflanzen. E. Schulze. 46: 451-458 (1896). 

Ueber die Verbreitung des Glutamins in den Pflanzen. E. Schulze. 

48: 33-55 (1897). 

Ueber den Lecithingehalt einiger Pflanzensamen und einiger Oelkuchen. 
E. Schulze. 4g: 203-214 (1898). 

Die Notwendigkeit der Umgestaltung der jetzigen Futter- und Nahr- 
ungsmittel-Analyse. E.Schulze. 49:419-441 (1898). 

Ueber die Verbreitung des Glutamins in den Pflanzen. {Zweite 
Mitteilung.) E.Schulze. 49:442-446(1898). 



8 Ernst Schulze [Sept. 

Ueber die Bestandteile der Samen von Pinus cemhra (Zierbeikiefer 

oder Arve). E. Schulze und N.Rongger. 51:189-204(1899). 
Ueber die Rückbildung der Eivveissstoffe aus deren Zerfallsprodukten 

in der Pflanze. E. Schulze. 55: 33-44 (1901). 
Ueber die Zusammensetzung einiger Koniferen-Samen. E. Schulze. 

55:267-307 (1901). 
Können Leucin und Tyrosin den Pflanzen als Nährstoffe dienen? E. 

Schulze. 56:97-106(1902). 
Ein Nachtrag zu der Abhandlung über die Frage ob Leucin und Tyrosin 

den Pflanzen als Nährstoffe dienen können. E. Schulze. 56: 

293-296 (1902). 
Zur Kenntnis der kristallisierten Stachyose. E. Schulze. 56 : 419- 

423 (1902). 
Ueber das Vorkommen von Hexonbasen in den Knollen der Kartoffel 

{Solanum tuberosum) und der Dahlie {Dahlia variabilis). E. 

Schulze. 59: 331-343 (1904)- 
Ueber Methoden, die zur Darstellung organischer Basen aus Pflanzen- 
säften und Pflanzenextrakten verwendbar sind. E. Schulze. 

59: 344-354 (1904). 

Zur Kenntnis des Glutamins. E. Schulze. 65: 237-246 (1906). 

Ueber die Bestandteile der Samen von Pinus cemhra. E. Schulze. 
67: 57-104 (1907). 

Zur Kenntnis des Glutamins. (Zzveite Mitteilung.) E. Schulze und 
Gh. Godet. 67: 313-319 (1907). 

Ueber die chemische Zusammensetzung der Samen unserer Kultur- 
pflanzen. E.Schulze. 73:35-170(1910). 

Zur Kenntnis des Glutamins. (Dritte Mitteilung.) E. Schulze und 
G. Trier. 77: 1-12 (1912). 

2. In den landwirtschaftlichen Jahrbüchern 

Untersuchungen über einige chemische Vorgänge bei der Keimung der 
gelben Lupine. E. Schulze, W. Umlauft und A. Urich. 5 : 
821-862 (1876). 

Die stickstoffhaltigen Bestandteile der vegetabilischen Futtermittel und 
ihre quantitative Bestimmung. E.Schulze. 6:157-175(1877). 

Ueber die Prozesse, durch welche in der Natur freier Stickstoff in 
Stickstoffverbindungen übergeführt wird, E. Schulze. 6 : 695- 
707 (1877). 

Ueber die Zersetzung und Neubildung von Eiweissstoffen in Lupinen- 
keimlingen. E.Schulze. 7:411-444(1878). 



jgi2] Ernst Winterstein 9 

Ueber den Eiweissumsatz im Pflanzenorganismus. E. Schulze. 9: 
689-748; 12: 909-920; 14: 713-729; 21: 105-130 (1880-1892). 

Untersuchungen über den Emmentaler Käse und über einige andere 
schweizerische Käsesorten. E. Benecke und E. Schulze. 16: 
317-400 (1887). 

Ueber die Bildungsweise des Asparagins und über die Beziehungen der 
stickstofffreien Stoffe zum Eiweissumsatz im Pflanzenorganismus. 
E. Schulze. 17: 683-711 (1888). 

Ueber die stickstofffreien Bestandteile der vegetabilischen Futtermit- 
tel. E.Schulze, 21:79-103(1892). 

Zur Kenntnis der in den pflanzlichen Zellmembranen enthaltenen 
Kohlenhydrate. E. Schulze. 23: 1-26 (1894). 

Ueber die Bildungsweise des Asparagins in den Pflanzen. E. Schulze. 
30: 287-297 (1901). 

Ueber den Abbau und den Aufbau der organischen Stickstoff Verbind- 
ungen in den Pflanzen. E. Schulze. 35: 621-666 (1906). 

3. Im Journal für Landwirtschaft 

Welchen Einfluss haben die Zubereitung des Futters und die Futter- 
mischung auf den Nährwert des Futters? Mit welchen Futter- 
stoffen sind bei den gegenwärtigen Marktpreisen Futterrationen 
mit angemessenem Gehalt an Nährstoffen am billigsten herzu- 
stellen. E. Schulze, 17: 33-48 (1869). 

Untersuchungen über die sensiblen Stickstoff-Einnahmen und -Aus- 
gaben des volljährigen Schafs und die Ausnutzung einiger Futter- 
stoffe durch dasselbe. E. Schulze und M. Märcker. 18: 1-39; 
19: 202-222, 285-326, 347-362; 20: 46-76 (1870-1872). 

Fütterungsversuche mit Schafen, E. Schulze und M. Märcker. 

23: 141-174 (1875)- 
Ueber die Zusammensetzung einer pechschweissigen Schafwolle und 

des daraus gewonnenen Wollfetts. E. Schulze und J. Barbieri. 

27: 125-144 (1879). 
Ueber die zur Gruppe der stickstofffreien Extraktstoffe gehörenden 

Pflanzenbestandteile. E. Schulze. 52: 1-30 (1904). 
Ueber die in den landwirtschaftlichen Kulturpflanzen enthaltenen, nicht 

proteinartigen Stickstoffverbindungen. E. Schulze. 52 : 305- 

336 (1904). 
Ueber den Nährwert der in den Futtermitteln enthaltenen nichtprote- 

inartigen Stickstoffverbindungen. E.Schulze. 54:65-81(1906). 



10 Ernst Schicke [Sept. 

4. In dem Landwirtschaftlichen Jahrbuch der Schweiz 

Ueber die Entstehung der Salpetersäuren Salze im Boden. E. Schulze. 

1890 : 109-121 ; 1891 : 82-86. 
Ueber die in den Futtermitteln enthaltenen Fettsubstanzen und über 

die Bedeutung derselben für die tierische Ernährung. E. Schulze. 

1892 : 1-9. 
Ueber den Humus und seine Beziehung zum Leben der Pflanze. E. 

Schulze. 1901 : 1-13. 
Die Nährstoffnormen und die Beurteilung des Nährwertes der Futter- 
bestandteile nach ihrer Verbrennungswärme. E. Schulze. 1902 : 

1-19. 
Ueber die chemische Zusammensetzung des Holzes und über einige aus 

demselben darstellbaren Produkte. E. Schulze. 1904: i-io. 

5. In der Zeitschrift für physiologische Chemie 

Untersuchungen über die Amidosäuren, welche bei der Zersetzung der 
Eiweissstoffe durch Salzsäure und durch Barytwasser entstehen. 
E. Schulze, J. Barbieri und E. Bosshard. 9: 63-126, 253-259 

(1885). 
Zur Kenntnis des Vorkommens von Allantoin, Asparagin, Hypoxanthin 

und Guanin in den Pflanzen. E. Schulze und E. Bosshard. 9: 

420-444 (1885). 
Notiz betreffend die Bildung von Sulfaten in keimenden Erbsen. E. 

Schulze. 9: 616 (1885). 
Ueber einen neuen stickstoffhaltigen Pflanzenbestandteil. E. Schulze 

und E. Bosshard. ig: 80-89 (1886). 
Untersuchung über die Amidosäuren, welche bei der Zersetzung der 

Eiweissstoffe durch Salzsäure und durch Barytwasser entstehen. 

Zzveite Abhandlung. E. Schulze und E. Bosshard. 10:134-145 

(1886). 
Ueber das Vorkommen von Vernin im Blütenstaub von Corylus avellana 

und von Pinus sylvestris. E. Schulze und A, von Planta, ig : 

326-330 (1886). 
Ueber das Arginin. E. Schulze und E. Steiger, ii : 43-65 (1887). 
Zur Kenntnis der beim Eiweisszerfall entstehenden Phenylamidopro- 

pionsäure. E. Schulze und E. Nägeli. ii : 201-206 (1887). 
Ueber das Vorkommen von Cholin in Keimpflanzen. E. Schulze. 

11:365-372 (1887). 
Ueber einige stickstoffhaltige Bestandteile der Keimlinge von Soja 

hispida. E.Schulze. 12:405-415 (1888). 



1912] Ernst Winterstein ii 

Ueber den Lecithingehalt der Pflanzensamen. E, Schulze und E. 

Steiger. 13:365-384(1889). 
Zur Chemie der Pflanzenzellmembran. E. Schulze, E. Steiger und 

W. Maxwell. 14: 227-273 (1890). 
Bilden sich Cholesterine in Keimpflanzen, welche bei Lichtabschluss 

sich entwickeln? E. Schulze. 14: 491-521 (1890). 
Ueber die Farbenreaktion des Isocholesterins mit Essigsäureanhydrid 

und Schwefelsäure. E. Schulze. 14: 522-523 (1890). 
Ueber die basischen Stickstoffverbindungen aus den Samen von Vicia 

sativa und Pisum sativum. E. Schulze. 15: 140-160 (1891). 
Ueber das Lecithin der Pflanzensamen. E. Schulze und A. Likier- 

NiK. 15: 405-414 (1891). 
Zur Chemie der pflanzlichen Zellmembranen. (Zweite Abhandlung.) 

E. Schulze. 16: 387-438 (1892). 
Ueber einige stickstoffhaltige Bestandteile der Keimlinge von Vicia 

sativa. E. Schulze. 17: 193-216 (1893). 
Ueber die Konstitution des Leucins. E. Schulze und A. Likiernik. 

17: 513-535 (1893). 

Zur Chemie der pflanzlichen Zellmembranen. (Dritte Abhandlung.) 
E. Schulze. 19: 38-69 (1894). 

Ueber die Bestimmung des Lecithingehaltes der Pflanzensamen. E. 
Schulze. 20:225-232(1895). 

Ueber das wechselnde Auftreten einiger krystallinischen Stickstoffver- 
bindungen in den Keimpflanzen und über den Nachweis derselben. 
E. Schulze. 20: 306-326 (1895). 

Ueber das Vorkommen von Glutamin in grünen Pflanzenteilen. E. 
Schulze. 20:327-334(1895). 

Ueber die Verbreitung des Rohrzuckers in den Pflanzen, über seine 
physiologische Rolle und über lösliche Kohlenhydrate, die ihn be- 
gleiten. E. Schulze und S. Frankfurt. 20: 511-555; 21: 108 

(1895). 

Ueber die Zellwandbestandteile der Cotyledonen von Lupinus luteus 
und Lupinus angustifolius und über ihr Verhalten während des 
Keimungs Vorganges. E. Schulze. 21: 392-411 (1895). 

Ueber das Vorkommen von Nitraten in Keimpflanzen. E. Schulze. 
22: 82-89 (1896). 

Ueber einen phosphorhaltigen Bestandteil der Pflanzensamen. E. 
Schulze und E. Winterstein. 22 : 90-94 (1896). 

Ueber das wechselnde Auftreten einiger krystallisierbaren Stickstoff- 
verbindungen in den Keimpflanzen. (Zzveite Abhandlung.) E. 
Schulze. 22:411-434(1896). ^ 



12 Ernst Schuhe [Sept. 

Ueber die beim Umsatz der Proteinstoffe in den Keimpflanzen einiger 
Coniferenarten entstehenden Stickstoffverbindungen. E. Schulze. 
22:435-448 (1896). 

Ueber den Umsatz der Eiweissstoft"e in der lebenden Pflanze. E. 
Schulze. 24: 18-114 (1898). 

Ueber die Spaltungsprodukte der aus den Coniferensamen darstell- 
baren Proteinstoffe. E. Schulze. 24 : 276-284 ; 25 : 360-362 
(1898). 

Ueber die Bildung von Ornithin bei der Spaltung des Arginins und 
über die Konstitution dieser beiden Basen. E. Schulze und E. 
Winterstein. 26: 1-14 (1898). 

Ueber den Eiweissumsatz und die Bildungsweise des Asparagins und 
des Glutamins in den Pflanzen. E. Schulze. 26 : 41 1-426 ( 1899) . 

Ueber die Verbreitung des Rohrzuckers in den Pflanzen, über seine 
physiologische Rolle und über lösliche Kohlenhydrate, die ihn be- 
gleiten. (Zweite Abhandlung.) E.Schulze. 27:267-291(1899). 

Nachweis von Histidin und Lysin unter den Spaltungsprodukten der 
aus Coniferensamen dargestellten Proteinsubstanzen, E. Schulze 
und E. Winterstein. 28: 459-464 (1899). 

Ueber das Vorkommen von Histidin und Lysin in Keimpflanzen. E. 
Schulze. 28: 465-470 (1899). 

Einige Bemerkungen über das Arginin. E. Schulze. 29 : 329-333 
(1900). 

Ueber den Umsatz der Eiweissstoffe in der lebenden Pflanze. (Zweite 
Abhandlung.) E. Schulze. 30: 241-312 (1900). 

Ueber die Ausbeute an Hexonbasen, die aus einigen pflanzlichen Ei- 
weissstoffen zu erhalten sind. E. Schulze und E. Winterstein. 

33: 547-573 (1901)- 
Beiträge zur Kenntnis des Arginins und Ornithins. E. Schutlze und 

E, Winterstein. 34: 128-147 (1901). 
Ueber die Trennung des Phenylalanins von anderen Aminosäuren. E. 

Schulze und E. Winterstein. 35: 210-220 (1902). 
Beiträge zur Kenntnis einiger aus Pflanzen dargestellten Aminosäuren. 

E. Schulze und E. Winterstein. 35: 299-314 (1902). 
Beiträge zur Kenntnis der Hemicellulosen. E. Schulze und N. 

Castoro. 37: 40-53 (1902). 
Beiträge zur Kenntnis der Zusammensetzung und des Stoffwechsels 

der Keimpflanzen. E. Schulze und N. Castoro. 38: 200-258 

(1903)- 
Beiträge zur Kenntnis der Hemicellulosen. E. Schulze und N. 

Castoro. 39: 318-328 (1903). 



igi2] Ernst Wintersfein 13 

Zur Kenntnis der aus Pflanzen darstellbaren Lecithine. (Erste Mit- 
teilung.) E. Schulze und E. Winterstein. 40:101-119(1903). 

Ein Nachtrag zur Abhandlung über einen phosphorhaltigen Bestand- 
teil der Pflanzensamen. E. Schulze und E. Winterstein. 40 : 
120-122 (1903). 

Beiträge zur Kenntnis der in ungekeimten Pflanzensamen enthaltenen 
Stickstoflfverbindungen. E. Schulze und N. Castoro. 41 : 

455-473 (1904). 

Einige Notizen über das Lupeol. E. Schulze. 41: 474-476 (1904). 

Findet man in Pflanzensamen und in Keimpflanzen anorganische Phos- 
phate? E. Schulze und N. Castoro. 41: 477-484 (1904), 

Beiträge zur Kenntnis der Zusammensetzung und des Stoflfwechsels der 
Keimpflanzen. (Zzveite Mitteilung.) E. Schulze und N. Cas- 
toro. 43: 170-198 (1904). 

Ueber das Vorkommen von Ricinin in jungen Keimpflanzen. E. 
Schulze und E. Winterstein. 43: 211-221 (1904). 

Ueber das Verhalten des Cholesterins gegen das Licht. E. Schulze 
und E. Winterstein. 43: 316-319 (1904). 

Ueber die aus den Keimpflanzen von Vicia sativa und Lupinus albus 
darstellbaren Monoaminosäuren. E. Schulze und E. Winter- 
stein. 45: 38-60 (1905). 

Ueber das spezifische Drehungsvermögen einiger aus Pflanzen darge- 
stellten Tyrosinpräparate. E. Schulze und E. Winterstein. 

45:79-83(1905)- 

Neue Beiträge zur Kenntnis der Zusammensetzung und des Stoff- 
wechsels der Keimpflanzen. E. Schulze. 47: 507-569 (1906). 

Ueber den Tyrosingehalt der Keimpflanzen von Lupinus albus. E. 
Schulze und N. Castoro. 48: 387-395 (1906). 

Bildet sich Homogentisinsäure beim Abbau des Tyrosins in den Keim- 
pflanzen? E. Schulze und N. Castoro. 48: 396-411 (1906). 

Ueber das Verhalten des Cholesterins gegen das Licht. (Zweite Mit- 
teilimg.) E. Schulze und E. Winterstein. 48:546-548(1906). 

Ist die bei Luftzutritt eintretende Dunkelfärbung des Rübensaftes 
durch einen Tyrosin- und Homogentisinsäuregehalt dieses Saftes 
bedingt? E. Schulze. 50: 508-524 (1907). 

Ueber den Phosphorgehalt einiger aus Pflanzen dargestellter Lecithin- 
präparate. E. Schulze. 52: 54-61 (1907). 

Zum Nachweis des Rohrzuckers in Pflanzensamen. E. Schulze. 
52:404-411 (1907). 

Ueber die zur Darstellung von Lecithin und anderen Phosphatiden 



14 Ernst Schulze [Sept. 

aus Pflanzensamen verwendbaren Methoden. E. Schulze. 55 : 

338-351 (1908). 

Einige Bemerkungen zu den Arbeiten über den Nährwert der in den 
Pflanzen enthaltenen Amide. E.Schulze. 57:67-73(1908). 

Ueber den Calcium- und Magnesiumgehalt einiger Pflanzensamen. E. 
Schulze und Ch. Godet. 58: 156-161 (1908). 

Ueber das Stachydrin. E. Schulze und G. Trier. 59:233-235(1909). 

Ueber die zur Darstellung von Cholin, Betain und Trigonellin aus 
Pflanzen verwendbaren Methoden und über die quantitative Be- 
stimmung dieser Basen. E.Schulze. 60:155-179(1909). 

Untersuchungen über die in den Pflanzensamen enthaltenen Kohlen- 
hydrate. E. Schulze und Ch. Godet. 61: 279-350 (1909). 

Ueber das Vorkommen von Betain in den Knollen des Topinamburs 
(Helianthus tuberosus). E. Schulze. 65: 293-294 (1910). 

Studien über die Proteinbildung in reifenden Pflanzensamen. E. 
Schulze und E. Winterstein. 65: 431-476 (1910). 

Ein Beitrag zur Kenntnis des Vernins. E. Schulze. 66: 128-136 
(1910). 

Ueber die in den Pflanzen vorkommenden Betaine. E. Schulze und 
G. Trier. 67: 46-58 (1910). 

Ueber das Stachydrin und über einige neben ihm in den Stachysknollen 
und in den Orangenblättern enthaltene Basen. E. Schulze und 
G. Trier. 67: 59-96 (1910). 

Ueber das Vorkommen von Hemicellulosen in den Samenhülsen von 
Pisum sativum und von Phaseolus vulgaris. E. Schulze und U. 
Pfenninger. 68: 93-108 (1910). 

Erwiderung auf R. Engelands Bemerkungen zu den Abhandlungen 
über die pflanzlichen Betaine und das Stachydrin. E. Schulze 
und G. Trier. 6g: 326-328 (1910). 

Ein Beitrag zur Kenntnis der in den Pflanzensamen enthaltenen Kohl- 
enhydrate. E. Schulze und U. Pfenninger. 69: 366-382(1910). 

Ueber die Identität des Vernins und des Guanosins, nebst einigen Be- 
merkungen über Vicin und Convicin. E. Schulze und G. Trier. 

70: 143-151 (1910)- 
Studien über die Proteinbildung in reifenden Pflanzensamen. {Zweite 

Mitteilung.) E. Schulze. 71: 31-48 (1911). 
Untersuchung über die in den Pflanzen vorkommenden Betaine. "E. 

Schulze und U. Pfenninger. 71: 174-185 (1911). 
Zur Frage der Identität des aus Melasse dargestellten Guaninpentosids 

mit dem Vernin. E. Schulze und G. Trier, 76 : 145-147 (1912). 



I9I2] Ernst Winterstein 15 

Untersuchungen über die in den Pflanzen vorkommenden Betaine. 

(Zzueite Mitteilung.) E. Schulze und G. Trier. 76: 258-290 

(1912). 
Dasselbe. (Dritte Mitteilung.) E. Schulze und G. Trier. 79:235- 

242 (1912). 

6. Im Journal für praktische Chemie 

Ueber die Zusammensetzung der rohen Schafwolle. M. Märcker und 

E. Schulze. 108: 193-207 (1870). 
Ueber die Zusammensetzung des Wollfetts. E. Schulze. 7 (n. f.) : 

1-16 (1873). 
Dasselbe. E. Schulze und A. Urich. 9: 321-339 (1874). 
Ueber die Eivveisszersetzung in Kürbiskeimlingen. E. Schulze und 

J. Barbieri. 20: 385-418 (1880). 
Ueber das Vorkommen von Allantoin und Asparagin in jungen Baum- 
blättern. E. Schulze und J. Barbieri, 25: 145-158 (1882). 
Zur Kenntnis der Cholesterine. E. Schulze und J. Barbieri. 25: 

159-180 (1882). 
Ein Nachtrag zu der Abhandlung: "Zur Kenntnis der Cholesterine." 

E. Schulze. 25: 458-462 (1882). 
Ueber Phenylamidopropionsäure, Amidovaleriansäure und einige andere 

stickstoffhaltige Bestandteile der Keimlinge von Lupinns luteus. 

E. Schulze und J. Barbieri. 27: 337-362 (1883). 
Zur quantitativen Bestimmung des Asparagins und des Glutamins. E. 

Schulze. 31:234-246 (1885). 
Zur Kenntnis der stickstoffhaltigen Bestandteile der Kürbiskeimlinge. 

E. Schulze. 32: 433-460 (1886). 

7. In den Berichten der Deutschen Chemischen Gesellschaft 

Über Maltose. E. Schulze. 7: 1047 (1874). 

Über die Zusammensetzung des Wollfetts. E.Schulze. 8:570(1875). 

Selenoidodiglykolsäure. A. Urich und E. Schulze. 8: yyT, (i875)- 

Die stickstoffhaltigen Bestandteile der Rüben. E. Schulze und A. 
Urich. 9:80 (1876). 

Keimung der Lupinensamen. E. Schulze und W. Umlauft. 9: 
1314 (1876). 

Über die stickstoffhaltigen Bestandteile der Runkelrüben (Glutamin). 
E. Schulze und A. Urich. ig: 88 (1877). 

Über das Vorkommen eines Glutaminsäureamids in den Kürbiskeim- 
lingen. E. Schulze und J. Barbieri. ig: 199 (1877). 

Eiweisszersetzung in Keimpflanzen. E. Schulze, ii: 520 (1878). 



l6 Erfist Schuhe [Sept. 

Bildung von schwefelsauren Salzen bei der Eiweisszersetzung in Keim- 
pflanzen. E. Schulze, ii: 1234 (1878). 

Asparagin und Tyrosin in Kürbiskeimlingen. E. Schulze und J. Bar- 
BiERi. 12 : 710 (1879). 

Leucin aus Kürbiskeimlingen. E. Schulze und J. Barbieri. 12 : 1233 

(1879). 
Über ein Glucosid aus Lupinus luteus. E. Schulze und J. Barbieri. 

12: 2200 (1879). 
Über das spezifische Drehungsvermögen des Isocholesterins. E. 

Schulze. 13: 249 (1880). 
Über ein neues Glucosid. E. Schulze und J. Barbierl 13:681(1880), 
Amidosäuren in Lupinenkeimlingen. E. Schulze und J. Barbieri. 

13: 1924 (1880). 
Über die Eiweisszersetzung in Kürbiskeimlingen. E. Schulze und J. 

Barbieri. 13: 2386 (1880). 
Über das Vorkommen von Allantoin im Pflanzenorganismus. E. 

Schulze und J. Barbieri. 14: 1602 (1881). 
Über das Vorkommen von Phenylamidopropionsäure unter den Zersetz- 
ungsprodukten der Eiweissstoffe. E. Schulze und J. Barbieri. 

14: 1785 (1881). 
Zur Kenntnis des Cholesterins. E. Schulze und J. Barbieri. 15: 

953 (1882). 
Über das Vorkommen von Allantoin und Asparagin in jungen Baum- 
blättern. E. Schulze und J. Barbieri. 15:955(1882). 
Beiträge zur Kenntnis der stickstoffhaltigen Bestandteile der Kartoffeln. 

E. Schulze und E. Eugster. 15: 1090 (1882). 
Über das optische Verhalten einiger Aminosäuren. E. Schulze und 

E. Bosshard. 17: 1610 (1884). 
Über die Bildung von Phenylamidopropionsäure beim Erhitzen von 

Eiweissstoffen mit Salzsäure und Zinnchlorür. E. Schulze und 

J. Barbieri. 17: 171 1 (1884). 
Über das optische Verhalten einiger Aminosäuren. E. Schulze und 

E. Bosshard. 18: 388 (1885). 
Über das Vorkommen von Glutamin in den Zuckerrüben und über das 

optische Verhalten desselben. E. Schulze und E. Bosshard. 18 : 

390 (1885). 
Über einen neuen stickstoffhaltigen Bestandteil der Keimlinge von 

Lupinus luteus. E.Schulze. 19:1177(1886). 
Über Paragalactan. E. Schulze. 20: 290 (1887). 
Bilden sich Nitrate im Organismus lebender Pflanzen? E. Schulze. 

20: 1500 (1887). 



I9I2] Ernst Winterstein 17 

Über das Vorkommen von Cholin in Keimpflanzen. E. Schulze. 21 : 
21 (1888). 

Über das Vorkommen von Rohrzucker in unreifen Kartoffeln. E. 
Schulze und Th. Seliwanow. 21 : 299 (1888). 

Über den Nachweis von Rohrzucker in vegetabilischen Substanzen. E. 
Schulze und Th. Seliwanow. 21 : 299 (1888). 

Ein Beitrag zur Veränderung, welche die stickstoffhaltigen Bestand- 
teile eingesäuerter Grünfutterstoffe erleiden. E. Schulze. 21 : 
668 (1888). 

Über das Vorkommen eines unlöslichen Schleimsäure gebenden Kohlen- 
hydrats in Rotklee und Luzerne. E. Schulze und E. Steiger. 
22:345 (1889). 

Über die Zersetzung der Proteinsubstanzen in verdunkelten grünen 
Pflanzen. E. Schulze und E. Kisser. 22:350(1889). 

Über einige stickstoffhaltige Bestandteile der Keimlinge von Soja his- 
pida. E. Schulze. 22: 599 (1889). 

Zur Kenntnis der chemischen Zusammensetzung der Pflanzenzellmem- 
branen. E. Schulze. 22: 1192 (1889). 

Betain und Cholin in den Samen von Vicia sativa. E. Schulze, 22 : 
1827 (1889). 

Untersuchungen über die stickstofffreien Reservestoffe der Samen von 
Lupinus luteus und über die Umwandlung derselben während des 
Keimprozesses. E. Schulze und E. Steiger. 23: 405 (1890). 

Über ein Krystallisieren des Kohlenhydrats. A. v. Planta und E. 
Schulze. 23: 1692 (1890). 

Zur Kenntnis der chemischen Zusammensetzung der pflanzlichen Zell- 
membran. E.Schulze. 23:2579(1890). 

Darstellung von Lecithin aus Pflanzensamen. E. Schulze und A. 
LiKiERNiK. 24:71 (1891). 

Über den Lecithingehalt der Pflanzensamen. E. Schulze und E. 
Steiger. 24: 327 (1891). 

Zur Chemie der Pflanzenzellmembran. E. Schulze, E. Steiger und 
W. Maxwell. 24: 530 (1891). 

Über die Konstitution des Leucins. E. Schulze und A. Likiernik. 
24: 669 (1891). 

Bilden sich Cholesterine in Keimpflanzen, welche bei Lichtabschluss 
sich entwickeln? E. Schulze. 24: 670 (1891). 

Über die Farbenreaktion des Isocholesterins mit Essigsäureanhydrid 
und Schwefelsäure. E. Schulze. 24: 671 (1891). 

Über die Bildung von stickstoffhaltigen Basen beim Eiweisszerfall im 
Pflanzenorganismus. E. Schulze. 24: 1098 (1891). 



i8 Ernst Schuhe [Sept. 

Zur Kenntnis der chemischen Zusammensetzung der pflanzlichen Zell- 
membran. E. Schulze. 24: 2277 (1891). 
Über die Bildung von Harnstoff bei der Spaltung des Arginins. E. 

Schulze und A. Likiernik. 24: 2701 (1891). 
Zur Kenntnis des Stachydrins, A. v. Planta und E. Schulze. 24: 

2705 (1891). 
Über basische Stickstoffverbindungen in den Samen von Vicia sativa 

und Pisum sativum. E. Schulze. 25: 84 (1892). 
Über das Lecithin der Pflanzensamen. E. Schulze und A. Likiernik, 

25:85 (1892). 
Zur Chemie der pflanzlichen Zellmembranen. E. Schulze. 25 : 434 

(1892). 
Über das Vorkommen von Guanidin im Pflanzenorganismus. E. 

Schulze. 25:658(1892). 
Über einen stickstoffhaltigen Bestandteil der Keimlinge von Vicia sativa, 

E. Schulze. 25: 869 (1892). 
Zum Nachweis des Guanidins. E. Schulze. 25: 2213 (1892). 
Über das Vorkommen von Betain und Cholin in Malzkeimen und in 

den Keimen des Weizenkorns. E. Schulze und S. Frankfurt. 

26: 2151 (1893). 
Über die Verbreitung des Rohrzuckers in Pflanzen. E. Schulze und 

S. Frankfurt. 27: 62 (1894). 
Über das Vorkommen von Raffinose im Keime des Weizenkorns. E. 

Schulze und S. Frankfurt. 27: 64 (1894). 
Über krystallisiertes Lävulin. E. Schulze und S. Frankfurt. 27: 

65 (1894). 
Über das Vorkommen von Trigonellin in den Samen von Pisum sati- 
vum und Cannabis sativa. E. Schulze und S. Frankfurt. 27 : 

769 (1894). 
Über ;8-Lävulin. E. Schulze und S. Frankfurt. 27: 3525 (1894). 
Vorkommen von Arginin in Knollen und Wurzeln einiger Pflanzen. E, 

Schulze. 29: 352 (1896). 
Verbreitung des Glutamins in den Pflanzen. E. Schulze. 29: 1882 

(1896). 
Stickstoffhaltige Bestandteile der Keimpflanzen von Ricinus communis. 

E. Schulze. 30: 2197 (1897). 
Über die Spaltungsprodukte des Arginins. E. Schulze und E. Win- 
terstein. 30:2879(1897). 
Bestandteile des Wollfetts. E. Schulze. 31: 1200 (1898). 
Konstitution des Arginins. E. Schulze und E. Winterstein. 32 : 

3191 (1899). 



I9I2] Ernst Winterstein 19 

Über das spezifische Drehungsvermögen des Glutamins. E. Schulze. 

39: 2932 (1906). 
Die Konstitution des Stachydrins, E. Schulze und G. Trier. 42 : 

4654 (1909). 

8. In verschiedenen Zeitschriften 

Ueber die Elementarzusammensetzung der tierischen Fette, insbeson- 
dere der Fette vom Schaf, Rind und Schwein. E. Schulze und 
A. Reinecke : Annalen der Chemie und Pharmacie, 142 ; 191-218 
(1867). 

Untersuchungen über die Respiration des volljährigen Schafes bei 
Erhaltungsfutter. W. Henneberg, E. Schulze, M. Märcker 
und L. Busse : Zentralblatt für die medizinischen Wissenscliaften, 
1870; 353-356; 369-370. 

Ueber Stickstoffausscheidung im Harn der Wiederkäuer. E. Schulze 
und M. Märcker: Zeitschrift für Biologie, 7; 49-62 (1871). 

Ueber die Bestimmung des aus Amiden abspaltbaren Ammoniaks in 
Pflanzenextrakten. E. Schulze. Zeitschrift für analytische 
Chemie: 21 : 1-26 (1882). 

Ueber das Stachydrin. E. Schulze: Archiv der Pharmacie, 231 ; 305 

(1893). ^ 
Zur quantitativen Bestimmung der Kohlenhydrate. E. Schulze: 

Chemikerzeitung, 1894; 527. 

Ueber die Analyse der Pflanzensamen. E. Schulze: Ibid., 1894; 
No. 43- 

Ueber die Cellulose. E. Schulze: Ibid., 1895; No. 65. 

Inwieweit stimmen der Pflanzenkörper und der Tierkörper an ihrer 
chemischen Zusammensetzung überein und inwiefern gleicht der 
pflanzliche Stoffwechsel dem tierischen? E. Schulze: Viertel- 
jahr sschrift der Naturforschenden Gesellschaft in Zürich, 1894; 

243- 

(A) Verbreitung des Glutamins in den Pflanzen. (B) Die in den 
Keimpflanzen der Coniferen enthaltenen Stickstoffverbindungen. 
E. Schulze. Verhandl. d. Schweiz. Naturf. Gesellsch., Zürich 
(1896), 126-127. 

Ueber die Zellwandbestandteile der Cotyledonen von Lupinus Intens 
und Lupinus angustifolius und über das Verhalten während der 
Keimungsvorgänge. E. Schulze : Berichte der Deutschen Bo- 
tanischen Gesellschaft, 14; 66-71 (1896). ^ 

Ueber den Eiweisszerfall und Eiweissbildung in der Pflanze. E. 
Schulze : Ibid., 18 ; 36-42 ( 1900) . 



20 Ernst Schulze [Sept. 

Ueber Tyrosinbildiing in den keimenden Samen von Lupinus albus und 
über den Abbau primärer Eiweisszersetzungsprodukte in den 
Keimpflanzen. E. Schulze: Ibid., 21; 64-67 (1903). 

Ueber die Argininbildung in den Keimpflanzen von Lupinus luteus. 
E. Schulze: Ibid., 22 ; 381-384 (1904). 

9. Dissertationen 

Ueber die Eiweisssubstanz der Kürbissamen und über die Zersetzungs- 
produkte, welche während des Keimprozesses aus derselben ent- 
stehen. J. Barbieri. 1878. 

Zur Kenntnis des Glutamins. Ueber Ammoniakbestimmung in Pflanz- 
ensäften und Pflanzenextracten. E. Bosshard. 1880. 

Ueber die chemische Zusammensetzung der Samen von Lupinus luteus 
und über ein in denselben enthaltenes dextrinartiges Kohlenhy- 
drat. E. Steiger. 1886. 

Ueber das pflanzliche Lecithin und über einige andere Bestandteile der 
Leguminosenchalen. A. Likiernik. 1891. 

Zur Kenntnis des pflanzlichen Amyloids und über einige andere Be- 
standteile der pflanzlichen Zellmembranen. E. Winterstein. 
1892. 

Ueber die Zusammensetzung der Samen und etiolierten Keimpflanzen 
von Cannabis sativa und Helianthus annuus. S. Frankfurt. 
1893. 

Ueber die Zusammensetzung der Samen und der etiolierten Keim- 
pflanzen von Lupinus angustifolius. Miron Merlis. 1897. 

Ueber die Bestandteile der Samen von Picea excelsa und über die 
Spaltungsprodukte der aus diesen Samen darstellbaren Protein- 
stoffe. N. Rongger. 1898. 

Versuche zur quantitativen Bestimmung der bei der Zersetzung der 
Eiweisskörper durch Säuren entstehenden Basen. O. Meyer. 
1900. 

Versuche zur Bestimmung des Gehaltes einiger Pflanzen und Pflanzen- 
teile an Zellwandbestandteilen, Hemicellulosen und Cellulosen. A. 
Kleiber. 1900. 

Beiträge zur Kenntnis der Cholesterine und der Methoden, die zu ihrer 
Abscheidung aus den Fetten und zu ihrer quantitativen Bestim- 
mung verwendbar sind. E. Ritter. 1902. 

Beiträge zur Kenntnis der in den Pflanzensamen enthaltenen Kohlen- 
hydrate. Ch. Godet. 1909. 

Ein Beitrag zur Kenntnis der pflanzlichen Betaine und ihre Bedeutung. 
Das Stachydrin, seine Konstitution und seine Synthese. G. Trier. 
1910. 



A RESUME OF THE LITERATURE ON INOSITE- 

PHOSPHORIC ACID, WITH SPECIAL REFER- 

ENCE TO THE RELATION OF THAT 

SUBSTANCE TO PLANTS 

ANTON RICHARD ROSE 

(Laboratory of Biological Chemistry of Columbia University, at the College of 
Physicians and Surgeons, New York) 

Contents. — Discovery: by the microscopist, 21; by the chemist, 22. Occur- 
rence, 2^; preparation, 25; properties, 26; Constitution, 31; terminology and Clas- 
sification, 35 ; analytical methods, 37 ; role in plants, 39. Bibliography, 46. 

DISCOVERY OF INOSITE-PHOSPHORIC ACID SALTS 
BY THE MICROSCOPIST 

In 1854 Hartig/ engaged in a microscopic study of the seeds o£ 
various plants, noted in all his sections small particles similar to the 
starch grains of the potato, which at that time were absorbing the 
interest of plant physiologists.^ These grains were obviously not 
starch, as they did not give the characteristic blue color with iodine 
in potassium iodid Solution. They are more commonly present in 
seeds than starch, the latter being frequently replaced by fat. 
Hartig considered them an essential reserve product designed to 
play an important part in the germination of the seed and the 
growth of the plant. He first called them " Klebermehl " but 
within a year renamed them " aleurone grains " f rom the Greek 
aXevpov (wheat fiour), a term still in use. Not only did he consider 
them significant for the plants in which they are found but also for 
the animals which eat them. The method which he employed for 
separating them from the other parts of the seed is the one usually 
followed by the investigators who have since worked with these 

^ The papers ref erred to in the text and not accompanied by f ootnote ref er- 
ences are those which pertain specially to the literature of inosite-phosphoric 
acid and are given at the end of this review in the alphabetical order of the 
names of the authors. 

^The starch grains were minutely studied by Nägeli and his coworkers. 
Their work (collected in Die Stärkekörner, 1858) probably afforded the Stimulus 
for the efforts which led to the discovery of inosite-phosphoric acid so early. 

21 



22 Literature on Inosite-Phosphoric Acid [Sept. 

grains, namely, extraction of the macerated seeds with ether and 
removal of the aleuron grains from the cellular debris by Sedimenta- 
tion in this medium. They are insoluble in both alcohol and ether, 
somewhat soluble in water, and quite soluble in dilute acids. 

In the years following Hartig's discovery several other botanists 
turned their attention to the aleuron grains and came to conflicting 
conclusions as to their nature. Von Holle considered them protein 
carriers and referred to them as " Proteinkörner." Both Sachs and 
Gris looked upon the particles as fat concentrates. By far the most 
important study in this field was the comprehensive work of Pfeffer 
in 1872. He differentiated the grains described by Hartig into 
three groups : (i) crystals of calcium Oxalate, (2) a protein sub- 
stance, and (3) a Compound giving no reactions for protein, fat, or 
inorganic salts. This last type was found in all of the one hundred 
different seeds which he examined. These particles he described as 
having rounded surfaces, assuming spheroidal shapes and fre- 
quently twinning, so as to present a convoluted appearance. Enough 
of the grains were obtained for a chemical examination, which was 
made for him by his colleague Brandon. The solubilities were the 
same as those reported by Hartig. Nitrogen could not be detected. 
Positive tests were obtained for calcium, magnesium, and phos- 
phorus. Organic matter was noted and the Suggestion made that 
the substance was a phosphate combined with a carbohydrate. 
These phosphorus-bearing spheroidal bodies occurring with or in the 
aleuron grains Pfeffer named "Globoid." 

DISCOVERY OF INOSITE-PHOSPHORIC ACID BY THE CHEMIST 

Palladin, while engaged on a study of the proteins of Sinapis 
niger in 1893, observed an unusual phenomenon. After extraction 
of the fat-free finely ground seeds with ten per cent. sodium chlorid 
Solution and heating the extract, he obtained a voluminous precipi- 
tate which partly dissolved on standing. A few trials showed that 
he had a substance soluble in cold but insoluble in hot water. By 
filtering off the permanent coagulum, reheating the filtrate and 
filtering while hot, he obtained a fairly pure product rieh in phos- 
phorus, containing calcium and magnesium, but no nitrogen. It 
proved non-reducing when tested with Fehling Solution, both before 



I9I2] Anton Richard Rose 23 

and after hydrolysis with acids. It was soluble in water and aclds, 
and precipitated by the alkali earths and the heavy metals. 

Subsequently Schulze and Winterstein published a paper con- 
firming the observations of Palladin, and also noting that the phos- 
phorus was not precipitated by ammonium molybdate. These 
authors expressed the opinion that the Compound thus discovered by 
chemical procedure is identical with Pfeffer's " globoid." The fol- 
lovving year (1897), a more detailed paper was published by the 
junior author, in which the identity and properties of the substance 
were more fully revealed, and " inosite-phosphoric acid " was sug- 
gested as the proper name for the Compound, inasmuch as it yielded 
inosite and phosphoric acid on hydrolysis. 

The most complete study of this substance has been made by 
Posternak; his findings are embodied in eight papers and several 
applications for patents. He discarded the name suggested by 
Winterstein and proposed a structural formula which does not 
include the inosite ring. He gave to the substance the name 
"phytin" (from the Greek <f>vT7]v) and under this trade name it is 
now placed on the market by a chemical firm in Basel. 

OCCURRENCE OF INOSITE-PHOSPHORIC ACID 

As already noted, phytin was first thought to be a storage product 
in seeds ; and this early Impression has been confirmed by subsequent 
investigation, no case having been reported of a seed in which it is 
completely lacking. The accompanying table (i) lists the plants 
specially mentioned in the literature in connection with the study of 
inosite-phosphoric acid. 

The relative data in the table are not in close accord, but no true 
comparison can as yet be drawn between the species, for most of the 
data were obtained at periods when adequate and uniform analytical 
methods were unavailable. The figures quoted in the table give an 
approximate idea of the quantitative signijficance of this important 
Compound, in relation to other forms of phosphorus available for 
seedling growth and the phosphorus requirement of man and beast. 
Posternak makes the Statement that seeds rieh in fat carry the 
largest amount of phosphorus, which is in harmony with the micros- 
copist's observations that the aleuron grains are particularly numer- 



24 



Literatur e on Inosite-Phosphoric Acid 



[Sept. 



TABLE I 



Recorded analytic data on the occurrence of inosite-phosphoric acid 



Plant. Total 

P 
Per Cent. 

Spruce fir 0.66 

Pea (Pisum sativum).. 0.37 

Pea, yellow 

Bean, white {Phaseolus 

vulgaris) 0.51 

Bean, brown 

Hemp (Cannabis sativa) 1.46 

0.76 
Rice (Ory::a sativa)... 0.95 

Rice flour 

Rice bran 2.22 

Rice germ 6.20 

Wheat ( Triticum sativa) 0.45 
Wheat bran i.ii 

i.il 

Graham flour 

Sesame (Sesamum in- 

dicum) 0.77 

Corn (Zea mays) 0.29 

0.35 
Oats (Avena sativa) . . 0.46 

Barley (Hordium sati- 

vutn) 0.50 

0.47 

0.54 
Barley bran 

Sunflower {Helianthus 

annuus) 0.83 

Rye (Seeale cereale) . . 

0.43 



Ratio of the 

P in phyto- 

phosphate 

to Total P 

Per Cent. 

91-5 
70.8 
19.0 

81.6 
58.0 
91.4 

15-0 

45-9 
69.0 

74.1 
89.2 
29.9 
84.0 

52.0 

29.0 

16.3 

54.0 

48.9 

48.0 
87.4 



Plant. 



38.0 

36.S 
44.0 
60.4 

86.3 
90.3 
28.9 



Total 

P 

Per Cent. 



Ratio of the 

P in phyto- 

phosphate 

to Total P 

Per Cent. 

25.0 



Rye flour 

Rape (Brassica napus 

olifera) 0.98 80.0 

I-I9 44-5 

0.54 38.0 

Rape cake 49.5 

Soy bean (Soja hispida) 0.57 58.0 

Lentil (Lens esculenta) 0.30 82.6 

0.30 9.3 
Cocoanut (Cocus nuci- 

fera) 88.4 

695 



Cottonseed (Gossipium 
herhaceum) 93.6 

Pine : Pinus cembra . . . 0.47 14.39 
Pinus excelsa ... 0.63 21.6 

Castor bean (Ricinus 

communis) 0.26 41.6 

Millet (Panicum millia- 
ceum) 0.77 44-97 

Vetch (Vicia faba mi- 
nor) 0.47 4.4 

Red Clover (Hay) 0.24 70.0 

Radish : Root (Rapha- 

nus vulgaris) 0.02 15.0 

Turnip : Root (Bras- 
sica esculenta?) 0.02 15.0 

Dahlia : Tuber (Dahlia 
variabilis) ..." Spheroids of phjrtin " 

Potato : Tuber (Alliuni 
cepa ) " Spheroids of phytin " 



The analytic results in the above table are those for seeds of the plants, 
except in the last five cases. They are compiled from a number of sources. 
Among the plants studied for their phyto-phosphate content, in which the rela- 
tive amount of this substance is not given, are the following: Beta vulgaris, 
Brassica campestris, Cucurbita pepo, Ervum lens, Lupinus albus, L. angustifolius, 
L. luteus, Pinus laricio, P. maritima, Sinapis nigra, Solanum tuberosum, and the 
tubers of AI Hunt cepa and Dahlia variabilis. In only two materials reported, 
namely, rutabaga root and alfalfa hay, could no phyto-phosphate be found. In 
several instances the total phosphorus was not reported. Where there is a 
close agreement between two or more results, only one figure is given above. 



1912] Anton Richard Rose 25 

ous in oily seeds. He also remarks that the smaller seeds such as 
cereals are the richest in " phytin." This Compound is not entirely 
confined to seeds, its presence having also been noted in the potato 
near the eye and as characteristic spheroids in the tubers of Allium 
and Dahlia. Roots functioning as storage organs, such as those of 
the Brassicae, contain small amounts. None was found by Totting- 
ham and Hart in the mature stems and leaves of the common 
fodder plants, but it occurs in clover leaves and in millet during the 
late flovvering period, and also in tender shoots. 

PREPARATION OF INOSITE-PHOSPHORIC ACID AND ITS SALTS 

To prepare phytin or its closely related Compounds from seeds, 
they should be finely ground and, if fat is present in large amounts, 
it should be removed by extraction with ether and alcohol. Most 
of the preparations reported in the literature have been obtained 
from cereals by leaching with 0.2 per cent. hydrochloric acid Solu- 
tion. Acetic acid has also been used in i per cent. Solution, and 
in a few cases acid Solutions of greater concentration have been 
employed. To remove the soluble proteins from the extract, 
Levene used picric acid; other investigators have coagulated them 
by heating and filtration after cooling; but when acidulated water 
is used the proteins do not seem to interfere appreciably with the 
preparation of pure phytin. The reserve proteins of the seeds are 
of the globulin type and are soluble only in the presence of salt in 
the extracting agent. Precipitation of inosite-phosphoric acid from 
its Solutions can be accomplished by several methods, such as the use 
of the acetates of the heavy metals, barium chlorid in ammoniacal 
Solution or magnesia mixture. In these cases the precipitated Com- 
pound is obviously in a form different from that in which it occurs in 
the original material. To obtain the salt more nearly in the form in 
which it is found in the seed, it may be precipitated by heating the 
Solution to almost boiling and filtering while hot; or better, by 
adding four volumes of ninety-five per cent. alcohol. In obtaining 
pure preparations of inosite-phosphoric acid or its salts, a number 
of reprecipitations are necessary. These have been made alternately 
with copper, lead, and barium. Salts of the first two are decom- 
posed by suspending in distilled water and bubbling hydrogen sul- 



26 Literatlire on Inosite-Phosphoric Acid [Sept. 

phide through the liquid ; the third is removed by adding dilute sul- 
phuric acid. In all cases, the lead or copper salt is the last pre- 
cipitated in this manner of purification; and when the product is 
carefully washed, and the metal removed by hydrogen sulphide, the 
filtrate from the lead or copper sulphide is evaporated at a low tem- 
perature, leaving the inosite-phosphoric acid.^ 

The various salts which have been studied were made from this 
acid. In obtaining the acid for the preparation of pure Compounds, 
the greatest difficulty lies in removing the last traces of magnesium. 
Rising overcame this difficulty by taking up the syrupy acid with 
absolute alcohol, and adding ether until droplets of the acid formed. 
He then filtered off the acid magnesium inosite-phosphate and again 
evaporated. The commonest impurity in phytin is inorganic ortho- 
phosphate which, however, is easily removed. Starkenstein uses the 
calcium salts of the mixed acids and washes with glacial acetic acid, 
which dissolves the inorganic part but not the organic phosphorus 
Compound. Forbes precipitates with magnesia mixture, removes 
the excess of this reagent by washing with ammonia water, washing 
again with alcohol, and extracting with 95 per cent. alcohol con- 
taining 0.2 per cent. mineral acid, which also dissolves all the in- 
organic phosphorus and none of the " phytin." Attempts to prepare 
these salts synthetically will be referred to in a later section. 

PROPERTIES OF INOSITE-PHOSPHORIC ACID AND ITS SALTS 

The substance widely known as "phytin," and described in the 
middle of the last Century by the microscopists as " spheroid bodies," 
frequently assumes the globular shape when forced out of Solution, 
In most cases, the precipitate comes down as a flocculent amorphous 
mass. Inosite-phosphoric acid has not as yet been obtained in crys- 
talline form. At room temperature, it is a syrup of light straw 
color, which becomes very viscid on cooling to — 20° C, and darkens 
on heating to 100° C. Vorbrodt found that this coloration could 
not be prevented by replacing the air with an inert gas during the 
heating, and from this concludes that the change is not due to 
oxidation. If the heat is allowed to reach 125° C, an insoluble dark 
char is produced (cf. Posternak). Inosite-phosphoric acid may 

'For details of the method of preparing the acid see Hart and Patten 
(page48). 



1912] Anton Richard Rose ^y 

form neutral salts, acid salts, double salts, or acid double salts. The 
acid, neutralized with alkali and evaporated to dryness, gives a 
brownish horny mass; but if an alkali earth is also present, double 
salts are formed, which crystallize in fine needles with eight mole- 
cules of water. The magnesium salts crystallize in small and uni- 
form spherules, while the copper salts form large and irregulär 
spherules. Twin forms are frequently produced in the copper pre- 
cipitates, resembling the globoids of which drawings appear in 
Pfeffer's paper. These spheroid masses may be Clusters of needles 
of approximately equal lengths, as is suggested by the regularly 
pitted surfaces sometimes seen, and the term spherocrystal can 
accordingly be applied to them. The copper Compounds are green ; 
the others, as far as reported, are white. Occasionally a faint 
pinkish cast has been noticed in pure preparations. 

The acid is miscible in all proportions with water. It is soluble 
in alcohol but not in the other common lipoid solvents. Ether added 
to an alcoholic Solution precipitates the acid in droplets. According 
to Posternak, the acid-alkali and acid-magnesium salts are soluble 
in alcohol and water. The double salts are soluble in water, forming 
opalescent Solutions from which they are precipitated by chlorid and 
acetate of potassium, redissolving if these are added in excess. The 
decrease in solubility of the salts of inosite-phosphoric acid is in the 
following Order : alkali, alkali earth and heavy metal. The magne- 
sium Compounds are more soluble than the calcium salts and the 
latter more soluble than those of barium or Strontium. The same 
Order of solubility also holds for acid salts, double salts and normal 
salts. These phytophosphates are more soluble in cold than in hot 
water, and heating frequently precipitates them, even in the presence 
of dilute acetic acid. This precipitation is largely influenced by 
other Compounds in Solution, halogens and sulphates inhibiting, and 
phosphates facilitating the reaction. Posternak noted that the 
precipitates thus formed by heating were not always completely dis- 
solved on cooling; also that the phytophosphates not readily soluble 
in cold water were changed to more soluble forms by dissolving in 
dilute acid and precipitating with alcohol. Dilute mineral acids are 
solvents for all of these Compounds. Acetic acid does not dissolve 
the salts of inosite-phosphoric acid with the heavy metals, barium. 



28 Literature on Inosite-PJwsphoric Acid [Sept. 

calcium and Strontium ; but the magnesium and alkali salts, and the 
double salts, are very soluble in this reagent. Posternak says that 
the alkali salts of this acid are solvents for the Compounds with 
alkali earths, and that on standing, crystals of double salts form in 
these Solutions, tending to arrange themselves in rosettes — a further 
Suggestion as to the mode of formation of the characteristic sphero- 
crystals mentioned above. Inosite-phosphoric acid Solutions do not 
polarize light, and pass through semi-permeable membranes com- 
paratively slowly. 

All the salts of inosite-phosphoric acid, except the alkali salts and 
the acid magnesium salts, are precipitated f rom aqueous Solution by 
four volumes of alcohol. The acid and its salts in alcoholic Solu- 
tion are precipitated by ether. The addition of neutral Solutions of 
silver, lead, copper, cadmium, iron, uranium, Strontium, barium and 
calcium precipitates the acid f rom its Solutions ; so also do magnesia 
mixture and albumin. According to Posternak, precipitation with 
copper is prevented by the presence of fat. The copper salts are 
soluble in ammonium hydroxid Solution. Ammonium molybdate in 
nitric acid does not cause precipitation in dilute Solutions of inosite 
phosphates, but in concentrated Solutions white needles are formed 
on long Standing which are insoluble in nitric acid and soluble in 
water. Preparations f rom seeds retain persistently small amounts of 
magnesium, several reprecipitations being necessary to get a salt 
containing a single metal. It is equally difficult to get a preparation 
free from the hydrogen ion, and it may be said in general that the 
property of forming acid salts and double salts is very characteristic 
of inosite-phosphoric acid. The most important contribution to our 
knowledge of the nature and properties of its salts has been made 
recently by Anderson. From acid purified by means of barium pre- 
cipitation and the method described by Hart and Patten, the follow- 
ing Compounds have been prepared : tri-barium, penta-barium, penta- 
barium di-ammonium, penta-magnesium, penta-magnesium di-am- 
monium, tetra-cupric di-calcium, tetra-calcium, penta-calcium, hexa- 
cupric, octa-silver, and hepta-silver salts. Most of them are white 
amorphous powders, but the tri-barium and tetra-calcium salts can 
be reprecipitated in irregulär crystalline form. 

Pure preparations have been made and analyzed by several other 
investigators. The results of their work are given in Table 2. 



I9I2] 



Anton Richard Rose 



29 



TABLE 2 



Analytic data pertaining to inosite-phosphoric acid 
(Compüed from results reported in the literature of the subject) 



Name of 
Author 



Anderson. 



Contardi. 



Hart, Patten. 

Hart. 
Tottingham. 

Horner. 

Levene. 



Elements 



Carbon Hydro- 
gen 



Phosphorus 



Barium 



Magne- 
sium 



Calcium 



Other 
Metals 



Ratios' 



C=6 
P=x 



P=6 
M'=x 



(Prepared from the purified commercial product.)^ 



6.42 
4-59 



10.56 
10.76 



1-44 
1-15 



3-21 
3.22 



37-21 
48.87 
46.99 


14-13 
42-9 


13-03 
22.46 
17.66 






14.69 









16.87 
13.46 
14.07 

21.29 

26.37 

26.16 

19.07 

20.62 

21.75 
22.53 
16.88 
11.94 
13-02 
(Synthesized by means of inosite and ortho-phosphoric acid.) 



6.42(K) 

33-54(Cu) 
55-98(Ag) 
52.43(Ag) 


6 
6 
6 
6 
6 
6 
6 
6 
6 
6 
6 
6 
6 



14.24 
14-23 

9.16 

9.60 

( 

12.62 

8.17 



3-45 
3-61 
1.64 
1.68 



24.09 
24.31 

16.34 
16.05 



35-57 
34-48 







4 
4 
4 
4 



(Synthesized by means of inosite and pyro-phosphoric acid.) 



3-24 
1.58 



(acid) 



26.51 

16-55 

( 

26.08 
21.08 



35-90 



(Prepared from rice bran.) 



9.0 



13-8 



10.89 



10.63 
17-30 



3-00 



3-40 
3-63 



28.10 
12.50 
15.60 
21.40 

(Preparations from wheat bran.) 
25-98 



56.2 

23.2 


8.9 


13-5 



22.1 (Cu) 



16.38 



26.08 












5-8 


I-13 



2.6 (K) 



6 
2 



(Similar preparations.) 



(The commercial preparation.)- 
20.32 1 I 1.45 I 11-96 I 



9-84 

5.66 

17.90 

7-70 



1.47 
3-57 



23-00 

13-93 
13-16 

11-95 



(Preparations from hemp.) 



44-50 



40.14 



5-5 
6 

1-5 
2 



6 
10 
12 
12 



12 

10 

8 

IG 
12 



6 
4 



(Synthesized by means of inosite and ortho-phosphoric acid.) 



12 



12 
12 

12 



63 



- 1 7 



8.5 

4 

9-5 



^ The empirical formulas have been calculated from the analytic results, and 
the relations between the carbon and phosphorus, and the phosphorus and base 
(M'), are given in these two columns. The carbon and the phosphorus are, in 
each case, assumed to be six atoms per molecule. 

^ Placed on the market by the Gesellschaft für Chemische Industrie in Basel. 

^ A 0.2 per cent. HCl extract of wheat bran several times reprecipitated from 
weakly acid Solution by alcohol. 

* The first substance; obtained by extracting with sodium chlorid Solution 
and, after removing protein with picric acid, precipitating with copper and 
removing the copper. 



30 



Literature on Inosite-Phosphoric Acid 



[Sept. 



TABLE 2 (coniinued) 



Name of 
Author 



Elements 



Carbon 



Hydro- 
gen 



Phosphorus 



Barium 



Magne- 



Calcium 



Other 
Metals 



Ratiosl 



C=6 
P=x 



P=6 
M'=x 



Posternak. 



Rising. 

Suzuki, 

Yoshimura, 

Takaishi. 

Vorbrodt. 



Winterstein. 

Winterstein, 
Schulze. 



(A large number of preparations were raade in which several kinds of 

seeds were used.) 



9-97 
4-79 
7-25 
7-44 



6.51 





26.08 
25.89 
12.70 
19-42 
19-73 










6 
6 
6 
6 


3-70 

I.OO 

1-34 
1.49 










50-45 










8.41 








i9.02(Na) 









5-37 1 1-03 I 



(Preparations from barley bran.) 
13-08 I I I l52.6s(Ag)| 

(Preparations from rice bran.) 



1.21 



I 23.48 I 



I S-81 I 17-48 I 



(Preparations from corn.) 



15.18 I42.62 I I 

(Preparations from black mustard.) 
18.42 I I 7-8 I I 



9.65 I 2.83 I 15-20 I 



I 



I 5-5 
I - 

I 3.7 



II 

4 
12 

I 7 



1 6 I 10.6 



8 
6.5 

— ?* 



The alkali salts are hygroscopic, but the others do not change in 
weight under ordinary conditions. Posternak assigns to the double 
alkali salts of the alkali earths eight molecules of water. The 
barium salt prepared by Vorbrodt lost 9.33 per cent. of its weight in 
the presence of phosphorus pentoxid, but regained it when exposed 
to a moist atmosphere. At 110° C, 11.5 per cent. was lost. Ander- 
son reports for his tri-barium salt five molecules of water, for his 
tetra-calcium salt twelve, and for his penta-magnesium salt twenty- 
four. 

Inosite-phosphoric acid is easily decomposed by heating with 
strong acids in sealed tubes but does not spontaneously break down 
into its cleavage p'roducts. Mendel and Underhill kept a Solution 
of the acid for many months and found at the end of the time no 
apparent change. Posternak states that heating phytin in alkaline 
Solution to 100° C. causes no decomposition, but Winterstein found 
that if a twenty per cent. Solution of alkali were used (sodium 
hydroxid) and the temperature raised to 230° C, cleavage occurred. 
Contardi states that cleavage does not occur when these salts are 

* Extracted with sodium chlorid Solution, precipitated by heating, and fil- 
tered hot. 



I9I2] 'Anton Richard Rose 31 

heated in water to 200° C. under pressure. According tc Giacosa it 
is more readily hydrolyzed than lecithin. From the fact tbat the 
products of hydrolysis are inosite and phosphate, Winterstein came 
to the conclusion that the Compound is inosite-phosphoric acid. 

CONSTITUTION OF INOSITE-PHOSPHORIC ACID 

Posternak, who has done more work on this substance than any 
other one investigator, did not agree with Winterstein in the con- 
clusion set forth above. From his analyses he first constructed the 
following formula: 

HC(OH)OP .(OH) 



(X) I 

HC(OH)0 



P^ (OH) 



but, as benzoyl chlorid gave no positive test for the hydroxyl group, 
a second formula was proposed (anhydro-oxymethylene-diphos- 
phoric acid) : 

CH,-0-P(' 
/ ^(OH), 

(«) o: 

\ /^ 

\cHj-o— ?<;■ 

%(0H), 

He was of the opinion that inosite is synthesized from the products 
of hydrolysis when the "phytin" is heated under pressure with 
mineral acids. A number of chemists have expressed doubt concern- 
ing the probability of such a formation of inosite, either from this 
organic group or any part of it that might result from the action of 
the acid thereon. In 1907 Suzuki and his co-workers obtained 
inosite from "phytin" by the action of an enzyme, from which they 
concluded that inosite is an integral part of the "phytin" molecule 
and constructed the following formula to represent their view : 



32 



Litcrature on Inosite-Phosphoric Acid 



[Sept. 



HO-^P— O— Q 
Ho/ 



H 
-C— 0-P' 






/^ 



-OH 
\0H 



(3) 




P_0— CH HC— 0— Pf 



HO 
O^ 



=0 



v\0H 



sOH 



hq// 



;P— O— c- 

h 



-C— O— P= 

h 



=0 



\\0H 



HO/ \0H 

M. W., 660; C = 10.91%; P = 28.18%. 

Neuberg came to similar conclusions the following year when he 
obtained inosite and furfurol on mixing " phytin " with phosphoric 
acid and distilling under reduced pressure, and also showed that 
furfurol can be obtained from inosite. He proposed the following 
f ormula : 



H 

-c- 



(4) 



HO. 

H0\ H 

HO>P-0-(; 

HO^. 

HO-7P— O— CH HC— O 

HO/ 



H 



/OH 

O— ^<0H 

/OH 
-P.^OH 



\0H 



t 



-CH 



HO— P\ /P— OH 
HO/ ^0^ \0H 

M. W., 714;C = 10.085% ; P = 26.05%. 

Levene, working with a preparation from hempseed, was led to 
believe that the "phytin" of this grain contained in its molecule 
phosphate, inosite and a carbohydrate of the pentose group. His 
work was criticized by Neuberg, who claimed that there were 
impurities in the preparation. In view of the known intimate asso- 
ciation of the phytin with protein and carbohydrate in the aleuron 
grain, and the possible occurrence of a chemical combination of 
both phyto-phosphate and carbohydrate with protein, it is conceiv- 
able that Levene had a product holding pentose as an integral part 



igi2] 'Anton Riclmrd Rose 33 

and not as an impurity, though in view of all the available evidence 
Neuberg's criticism seems at the present time somewhat justifiable. 
Starkenstein also refused to accept the simple formula proposed 
by Posternak and offers the f ollowing : 

HO-^P— OH.HO— C-C— OH.HO— P^OH 

^\ j i /^ 

HO— P— OH. HO— C C— OH.HO— P^H 

o/ 11 \) 

(5) HO— C— C— OH 



HO OH 
0=P P=( 



/\/\ 
OH O OH 
M. W., 714 ; C = 10.985^ ; P = 26.05 

He argues that the phosphoric acid is in the pyro-form^ from the 
fact that its silver salt is the same color as silver pyro-phosphate, 
and that its behavior when titrated with Standard uranium acetate 
Solution is also like that of pyro-phosphoric acid. That it is not 
combined in the usual form of an ester but held loosely in a complex 
"addition form," he maintains from the fact that an increase of 
inosite and inorganic phosphate resulted from heating some of the 
calcium salt for an hour at ioo° C. These arguments are not alto- 
gether convincing. Anderson has prepared a silver salt of the ortho- 
tetra-phosphoric acid ester with inosite and reports it as being white 
like the pyro-phosphoric acid salt. 

In the quantitative titration of pyrophosphoric acid with a uran- 
ium acetate Solution, standardized by ortho-phosphate and using fer- 
rocyanide as indicator, only one half of the phosphorus value is 
obtained. Starkenstein explains this phenomenon by the assump- 
tion that one half of the more reactive ions of the phosphoric acid 
have been removed in the dehydration. Now if two phosphoric- 
acid groups had formed esters with one polyalcohol, analogous con- 
ditions would have resulted as far as the ions are concerned, and 
bivalent ions would be expected to connect the two phosphoric acid 

* That phosphorus occurs in plants in the pyro form may seem stränge to 
many, but this is not the first time that such an occurrence has been suggested. 
In 1892 Hardin (5". C. Exp. Sta. Bull., N. S., No. 8) reported his finding both 
pyro- and meta- phosphate in cottonseed meal, when he sought, in this feeding 
material, a substance toxic to cattle. 



34 Literature on Inosite-Phosphoric Acid [Sept. 

radicles. This may be illustrated by these two graphic represen- 
tations : 

.HO: OH^^^* \ /OH«-^ 

\ / 0=:P OH 

0=P— OH ^ -f— ua. 

The idea of this type of reaction for ortho and pyro-forms 
is in harmony with the fact that when inosite-phosphoric acid is 
precipitated in acid Solutions by divalent metals, the tri-metal salt is 
the more readily formed. The activity of the hydrogen ions is 
relatively greater in the inosite-phosphoric acid than the sum total 
of the ions of the ortho-forms would be, probably due to the influ- 
ence of increased negative electric charges in the many phosphorus 
atoms held in one molecule, so that, altho six having been eliminated 
in the assumed ester formation, the very reactive ions are eight 
in number. The last column in Table 2 is interesting in this con- 
nection. Finally, the Statement that the inosite-phosphoric acid 
is decomposed by dry heat has been shown to be erroneous by both 
Anderson and the writer. 

That phytin is a salt of inosite-phosphoric acid seems to be con- 
clusively demonstrated by the synthetic work of Contardi, whose 
preparations from rice bran gave analyses identical with a synthetic 
preparation obtained by heating anhydrous inosite with ortho- 
phosphoric acid (sp. g. 1.7). Other workers have attempted to 
substantiate this result, but so far without success. Carre could 
obtain only a mixture of the two chemicals ; Anderson was able to 
produce tetra-ortho and di-pyro-phosphoric acid esters. 

It does not follow that these preparations of inosite-phosphoric 
acid are identical in form with the organic phosphorus Compound 
occurring in plants. The writer calls attention elsewhere to a differ- 
ence of behavior between the phytophosphate in seeds and in prepa- 
rations.^ Certain data, not as yet published, as well as differences 
in the products described in the literature, fall in with those 
suspicions.^ 

What inosite-phosphoric acid is, in terms of a definite chemical 

'Rose: Technical Bulletin 20 of the New York Agricultural Experiment 
Station. 

' Cf . Preparations analyzed by Patten and Hart, Winterstein and Schulze, 
and Levene, in table 2, p. 29. 



I9I2] Anton Richard Rose 35 

structure, is an open question. It is probably an ester of phosphoric 
acid with inosite, in which six phosphoric acid groups are united 
with each inosite molecule. This ratio, Cß : Pg, is indicated for the 
vast majority of the pure preparations analyzed; exceptions are the 
preparations by Levene, Vorbrodt and Rising (Table 2), these 
authors giving the ratio Cq : P5.5. Anderson has pointed out that 
bis and Rising's silver salts are probably identical, Rising's analysis 
being equally well adapted to the formula C6Hi7027P6Ag7. It 
seems probable that the molecular weight when accurately deter- 
mined will be reported as 714 or will differ from this by the molec- 
ular weight of three molecules of water, The molecule seems to 
contain twelve hydrogen atoms readily separated in ionization, six of 
which are exceedingly reactive ; the remaining hydrogen atoms 
gradually diminish in reactivity by twos, the last four being slow 
to enter into an exchange with bases. The most readily formed 
salts are therefore those corresponding to an octavalent acid and the 
other common ones are in six and tenvalent combinations. 

TERMINOLOGY AND CLASSIFICATION 

The investigations of which this paper is a brief review have 
brought to the biological chemist and plant physiologist a type of 
phosphorus Compound from the plant world which is relatively 
new and probably of prime importance. The phosphorus which 
occurs in acid and water extracts was formerly considered inor- 
ganic phosphate, and awakened no especial interest, and the mention 
of organic phosphorus immediately brought to mind nucleoproteins 
and lecithins. In the organic laboratories combinations of phos- 
phorus with various organic radicals have been made and recently- 
prepared phosphoric acid esters'^ resemble " phytin " in some respects, 
and so may be considered of special concern to the biological 
chemist, as possibly bearing on problems in his field. A cleavage 
product of inosinic acid, c?-arabinose phosphoric acid,^ is significant 
as showing that carbohydrate esters are not confined to those pro- 
duced by the synthetic Operations of the laboratory. Even more 

'v. Lebedew: Biochem. Zeit., 1909, 20, 114; Neuberg and his coworkers : 
Ber., 43, 2060; Biochem. Zeit., 23, 515; 26, 115 and 529; 36, 5; Langheld, Ber., 44, 
2076. 

'Levene and Jacobs, Ber., 191 1, 44, 746. 



36 Literatur e on Inosite-Phosphoric Acid [Sept. 

striking are Iwanow's experiments® in which, when yeast was 
allowed to ferment sugar in the presence of sodium phosphate, there 
was noted a disappearance of the inorganic phosphorus, amounting 
to f rom eighty to ninety-three per cent. ; and in the liquors there was 
foiind an organic phosphorus Compound optically active and giving 
reactions for aldehydes and ketones. Biochemical syntheses of this 
class have also been successfully made by other investigators.^*' Of 
special interest are the inosite esters with ortho- and pyrophos- 
phoric acid prepared by Anderson. Mention may also be made of the 
spherocrystals discovered by Hansen" in the parenchyma cells of 
the Euphorbia caput medusae, which he describes as amorphous 
masses of calcium and magnesium phosphate, but which Belzug^^ 
later has shown to be salts of a new organic acid, phosphomalic acid. 
We may reasonably expect that additional phosphorus-bearing sub- 
stances of this kind will be discovered in nature by the phyto- 
chemist for which a rational System of nomenclature will be required. 

Rising, in his paper on inosite-phosphoric acid, refers to soluble 
phosphorus Compounds obtained from grains, which he promises to 
discuss in later contributions. These substances he considers closely 
related to "phytin," and proposes classifying them as a single group 
with the generic name " phyto-phosphoric acid." This term we may 
profitably adopt to indicate the acid radicals of those organic phos- 
phorus Compounds which may be found in the water and dilute acid 
extracts of plant materials. In this group will be included the gly- 
cerophosphates, phosphomalates, such hexose and pentose phos- 
phates as may be discovered in plants, and the phytin-like substances. 

The term " phytin " as used at present seems to designate that 
substance which is extracted from seeds by leaching with dilute 
acids, reacting positively in the tests for calcium, magnesium, and, 
after hydrolysis, for phosphoric acid and inosite. The multiplica- 
tion of trade names for definite chemical Compounds is not desirable. 
There are many students and other workers who must of necessity 

'Iwanow: Zeit, für physiol. Chein., 1107, 50, 281-288. 

"Young and Hardin : Biochem. Zeit., 191 1, 32, 173-188; Proc. Chem. Soc, 
21, 23, 24; Proc. Roy. Soc., London, 77, 80, 81, 82; Euler and Ohlsen: Biochem. 
Zeit., 191 1, 37, 313. 

" Hansen : Arbeit, des bot. Inst., Würzburg, 1888, 92-122. 

"Beizug: Jour. de bot., 1893, 7, 211-229. 



I9I2] Anton Richard Rose 37 

carry in memory more names of organic Compounds than they can 
reasonably be expected to define in terms of chemical formulae, if 
the common names do not in themselves off er suggestions of chem- 
ical structure. Unsystematic naming is contrary to the modern 
spirit of chemical nomenclature. Winterstein's " inosite-phosphoric 
acid" has priority over Posternak's "phytin" and the further ad- 
vantage of being a chemically descriptive term. The preference 
of several authors for this latter designation is evidenced by the 
fact that the name phytin is not adhered to or is given in parenthesis 
after the name " inosite-phosphoric acid." In this particular case 
the probability of confusion is very miich increased by the fact that 
the term " phytine " is already applied to Chlorophyll preparations 
whose chemical composition we cannot hope to know for some time 
and for convenience must perforce carry a non-chemically descrip- 
tive appellation. The word " phytin " seems to have all the Psycho- 
logie requirements of a really good trade name and the substance 
which it designates in the market is widely advertised in the Euro- 
pean medical Journals for its therapeutic properties, which are more 
than likely of questionable character, and the term will undoubtedly 
persist. 

It can be readily conceived that this may not be the only inosite- 
phosphoric acid in plants and we should look for other combinations 
in which the phosphorus may be in the ortho or pyro form — even 
the meta phosphate — and be present as the hexa, tetra, di, or mono 
phosphoric acid. Various incidents have suggested to the writer 
that some of these forms occur in preparations from seeds when 
certain treatment other than those described above is used. 

ANALYTICAL METHODS^ 

The quantitative estimation of phytin phosphorus has been 
effected only by determining the difference between the total soluble 
phosphorus and the inorganic phosphorus. Phytin research in 
animal and plant metabolism is therefore very largely dependent 
upon the accuracy of the determination of inorganic phosphorus. 

" The analytical methods are here treated very briefly, for their development 
is as yet imperfect and the literature conflicting. Many papers have not been 
mentioned and the reader is referred for these to the bibliography on page 
46. A more complete Statement with experimental data will be published later. 



38 Literature on Inosite-Phosphoric Acid [Sept 

The term soluble phosphorus above and elsewhere means of course 
the phosphorus Compounds which dissolve in cold acidulated water ; 
the amount is obtained by evaporating the extract and destroying 
the organic matter with sulphuric and nitric acids according to the 
method of Neumann,^^ after which the phosphorus is determined 
by the usual ammonium molybdate and magnesia mixture method 
as described by Sonnenschein and later modified by Woy. As ex- 
tracting agents both acetic acid and hydrochloric acid have been 
used. 

The first method to approximate an accurate determination of 
inorganic phosphorus in the presence of soluble organic phosphorus 
was that used by Hart and Andrews in 1903. Their extracting 
agent was 0.2 per cent. hydrochloric acid Solution, a solvent which 
has since been used by most investigators. Hart and Andrews 
noted that ammonium molybdate did not precipitate the phytin 
phosphorus, and used this fact to devise a method for separating 
the two kinds of phosphorus combination in Solution. They 
had some apprehension lest the strong acid in the usual molybdate 
Solutions would hydrolyze some of the organic phosphorus Com- 
pounds and thus yield high results for the inorganic portion. They 
determined the minimum amount of nitric acid necessary to give a 
rapid, complete, and crystalline Separation of the yellow precipitate 
(2 c.c. of nitric acid, specific gravity 1.2, in each 250 c.c. of Solu- 
tion) and added to the liquid of this acidity neutral ammonium 
molybdate Solution. 

Vorbrodt, in his excellent monograph on " phytin," developed a 
method which is based on a triple precipitation of the inorganic 
phosphorus, first precipitating in general by means of magnesia 
mixture and dissolving the precipitate in the least amount of 
nitric acid. This is diluted to 50 c.c, heated to 100° C, and treated 
with an equal volume of ammonium molybdate Solution. The 
yellow precipitate is dissolved in ammonia water, 25 c.c. of 5 per 
cent. barium chlorid are added,^^ and the precipitate after being 
washed and dried is weighed; or the phosphorus may be precipi- 
tated with magnesia mixture and weighed as magnesium pyro- 
phosphate. 

"Neumann: Zeit, für physiol. Chetn., 1902, 37, 115. 
"Riegler: Zeit. Anal. Chem., 1902, 41, 675. 



1912] 'Anton Richard Rose 39 

Stutzer in Germany and Forbes in this country, working inde- 
pendently, introduced a new idea in the determination of inorganic 
phosphorus, namely, the use of acid alcohol. Forbes and bis asso- 
ciates make an acidulated water extract and precipitate with mag- 
nesia mixture; the precipitate is then washed successively with 
ammonia water and alcohol, and the inorganic phosphorus separated 
f rom the phytins by digesting in cold 95 per cent. alcohol containing 
0.2 per cent. of nitric acid. This alcoholic Solution is finally filtered, 
the filtrate evaporated, and phosphorus determined in the residue 
in the usual way. 

Starkenstein has studied in some detail the application of titra- 
tion methods to this problem, and his results point to the possibility 
of determining quantitatively these different forms of phosphorus 
in the same Solution. He found that titration of a Solution con- 
taining ortho-phosphate, pyro-phosphate and inosite-phosphate with 
uranyl acetate standardized by ortho-phosphate, using cochineal as 
an indicator, gave in each case true values for total phosphorus; 
that with ferrocyanide as an indicator, the total phosphorus was 
equivalent to all of the phosphorus as ortho-phosphate, one half of 
that as pyro-phosphate and inosite-phosphate, the glycero-phosphate 
not entering into the reaction at all. Anderson notes that pyro- 
phosphoric acid can be converted into the ortho form by heating 
with dilute acids, while the inosite-phosphoric acid is not affected 
by this treatment. With these facts in mind a Volumetrie process 
may readily be devised. 

THE ROLE OF INOSITE-PHOSPHORIC ACID AND ITS 
SALTS IN PLANTS 

The literature on phosphorus metabolism in animals has become 
voluminous, but the botanists have published comparatively little on 
the changes of these Compounds and their probable significance in a 
plant's life history. That cell functioning is impossible in the 
absence of phosphorus is again emphasized in the recent work of 
Frouin,^^ which shows its absolute necessity in the growth of 
micro-organisms. The study of the role of phytin in plant life in- 
volves an investigation of the changes and distribution of all the 

^'Frouin: Compt. Rend. Soc. Biol., 1910, 68, 801-803. 



40 Literature on Inosite-Phosphoric Acid [Sept. 

pliosphorus Compounds and of inosite in the several stages of plant 
development. Since the methods of differentiating between the 
various combinations of phosphorus in plant substances are becom- 
ing highly perfected, we may expect rapid developments in our 
knowledge of their functions in plant processes. The universal 
presence of phytin in propagating and growing parts must be highly 
significant. This constant occurrence led Starkenstein to assume 
that "phytin" plays a specific role in the mechanism of growth of 
both plants and animals. If this be so, its biochemical reactions 
must be closely linked with carbohydrate and protein formation, 
and its occurrence with these substances in the aleuron grain must 
be more than a mere coincidence. 

In this connection it may be well to review briefly the literature 
regarding the aleuron grains. The best summary was found in 
Vines's text book of plant physiology (1886) but this is too brief 
to be satisfactory. From Pfeffer's comprehensive description, it 
appears that these grains form in the vacuoles during the ripening 
and desiccation of the seed; that the forms assumed are globular, 
which are less distorted and attain a larger size in the more fatty 
seeds. They consist morphologically of three parts: the large pro- 
tein particle, Pfeffer's globoid, and a membrane. Crystals of cal- 
cium Oxalate are sometimes present. Weyl^'^ isolated the grains from 
the " Paranuss " employing the method of all the previous inves- 
tigators, and made an extensive study of their proteins. This was 
in the days of the vegetable vitellins (globulins), and the chief protein 
of the aleuron grain having been shown to belong to this group, 
Weyl thought that the membrane was a modified form of the same 
protein, an albuminate. Three or four years later, Vines^^ under- 
took a study of these proteins and from his observations on mate- 
rial from a large variety of seeds, grouped them into five classes: 
vegetable peptone (water soluble), vegetable myosin, crystalloid, 
vit ellin (all three soluble in sodium chlorid Solution), albuminate 
(soluble in sodium carbonate Solution). These are described as 
plastic proteins, in part transported to the cells of the seed from 
other portions of the plant. According to Posternak, these pro- 

^ Weyl : Zeit, für physiol. Chem., 1S77, i, 84-96. 

"Vines: Proc. Roy. Soc. London, 28, 218; 30, 387; 31, 59, 62; see also 
Lundtke : Jahrb. wiss. Bot., 1890, 21, 62. 



I9I2] Anton Richard Rose 41 

teins constitute from fifty to seventy-five per cent. of the aleuron 
grain. It is doubtful whether they are simple proteins; more likely 
they contain both phosphorus and bases in their molecules. Besides 
protein, Posternak found carbohydrates which were not free, but 
combined with some other substances of the grain; also ash, to the 
extent of twenty-five to fifty per cent. The following analytic 
data were recorded : 

Per Cent. Per Cent. 

Phosphorus O.11-3.83 Magnesium 0.28-1.27 

Sulphur 0.64-0.81 Calcium 0.11-0.37 

Silicon 0.01-0.36 Iron 0.03-0.09 

Potassium 2.29-2.71 Manganese Trace 

These results were obtained from aleuron grains of sunflower, 
white lupin, hemp, and red fir. The author calls attention to the 
interesting fact that all the elements essential to plant growth are 
present in these bodies. These results are notable when com- 
pared with Bernardini's analyses of rice embryo: P2O5, 0.95; SO3 
(not given); SiOs, 0.25; K2O, 1.691; MgO, 1.389; CaO, 0.279; 
FesOg, 0.06; Mn, trace; NagO, trace. One would like to know 
whether the Silicon in these two substances is present in organic 
combination. 

The globoid or "phytin" is a calcium-magnesium salt of 
inosite-phosphoric acid. Phyto-phosphate is also combined in the 
protein granules, possibly in the form of the potassium salt, as 
Posternak believed, inasmuch as he could not separate the potassium 
salt from the globulin, although it is soluble in alcohol and globulin 
is not soluble in this liquor. He concluded that it is chemically 
attached to the protein. In germination, the aleuron grains swell 
up, forming a granulär viscid mass; both globoid and crystalloid 
go into Solution, enzymatic action sets in, and both phytin and pro- 
tein are hydrolyzed. 

The presence of an enzyme having the power to decompose phy- 
tin into inosite and an inorganic phosphate was first demonstrated 
by Suzuki and his associates in the bran of rice. It has also been 
found by Vorbrodt in other small grain, including wheat, rye, and 
barley ; likewise in larger seeds, as vetch and lentils. An extract of 
the kemel of indian corn gave no evidence of the presence of a phy- 
tase, but it was shown to develop during the germination of the 
grain. 



42 Literatlire on Inosite-Phosphoric Acid [Sept. 

In the earlier analyses of seeds, the inorganic phosphorus was not 
given a very prominent place and was usually reported as that por- 
tion obtained by subtracting the sum of the protein and lecithin phos- 
phorus from the total phosphorus; but as this dement began to re- 
ceive special attention its direct determination was attempted, and 
consequently the amount of inorganic phosphorus reported was 
lessened. Thus Umikoff^^ estimates the inorganic portion as fully 
half of the total phosphorus, but the more recent workers report 
it in very small percentages. With the germination of the seed, 
the inorganic phosphorus gradually increases at the expense of the 
organic form. This was at first attributed to the breaking up of the 
phosphorus-bearing proteins and lecithins. Tammann,^'' one of the 
earliest investigators to make direct determinations of inorganic 
phosphorus in seeds and their sprouts, found during germination 
an increase of this form which, in terms of P2O5, was from 0.324 
per Cent, to 0.443 P^^ cent. in a period of only twelve days. Ac- 
cording to the work of Prianischnikow^^ and Merlis,^^ lecithin de- 
creased one half during fifteen days' germination of Vicia sativa 
and Lupinus angustifolius. Iwanow found that the inorganic phos- 
phorus increased from a very small amount to 93.7 per cent. of the 
total phosphorus in germinating seeds of Vicia faba. He held 
that lecithin is the most stable of the organic forms and is altered 
very little. Phytin, owing to the presence of phytase, is practically 
all changed by this process. 

Vorbrodt has shown that the phosphorus Compounds, especially 
inosite-phosphoric acid, are particularly abundant in the germ. 
When the seed begins to sprout, this supply is increased by trans- 
portation of phosphorus from other parts of the grain, as is indi- 
cated by Zalesky's^^ Observation that in the sprouts of Lupinus 
angustifolius the total phosphorus increased in twenty-five days 
from 0.302 gram to 0.514 gram; the inorganic phosphorus doubled 
in amount, while the protein and lecithin phosphorus remained prac- 

"Umikoff: Russian Dissertation, 1895. (Cited by Zalesky: Ber. bot. Ges., 
1902, 20, 426-433.) 

^Tammann: Zeit, für physiol. Chem., 1885, 9, 416-418. 

" Prianischnikow, 1895. (Cited by Zalesky. See footnote 19.) 

^'Merlis: Landw. Vers. Stat., 1897, 18. (Cited by Zalesky. See footnote 19.) 

^Zalesky: Ber. bot. Ges., 1902, 20, 426-433. 



I9I2] Anton Richard Rose 43 

tically unchanged. Bernardini found that the phytin decreased also 
in the germination of wheat, but the lecithins increased. 

In the early stages of plant development subsequent to germina- 
tion, inorganic exceeds organic phosphorus. Staniszkis could find 
no trace of inosite-phosphoric acid in millet during this period. The 
phosphorus is now drawn f rom the soil and its increase in the plant 
is proportional to the increase in dry matter. The organic forms of 
phosphorus are synthesized from this supply, according to Stan- 
iszkis, very slowly until the heads are formed. Hart and Totting- 
ham found no phytin phosphorus in the dried forage plants. 
Balicka-Iwanowska, working with barley, found the phosphorus 
Compounds at the end of the fourth week present in the same pro- 
portion as that in the seed ; thereafter there was a constant decrease 
in the inorganic phosphorus. In the seventh week the protein phos- 
phorus had doubled and the inosite-phosphoric acid had increased to 
seven times the amount present in the fourth week. As the barley 
seeds began to form, in the ninth week, the increase of organic 
phosphorus occurred mostly through the synthesis of nucleoproteins. 
The small increase which Staniszkis reports was more in the form of 
lecithins and protein phosphorus Compounds than of inosite-phos- 
phoric acid. At the period of flowering, the lecithins reach their 
maximum, which may be, according to Stoklasa, 71.6 per cent. of 
the total phosphorus. During the formation of millet seed, the syn- 
thesis of phytin goes on energetically at the expense of both inor- 
ganic and protein phosphorus. In the barley, Balika-Iwanowska 
found an increase of inosite-phosphoric acid and also one of nucleo- 
protein which was even greater than that of the phytophosphate. In 
the ripening of the seeds of millet the formation of inosite phos- 
phates and nucleoprotein ran parallel, but in barley the inosite phos- 
phate increased at the expense of some of the protein phosphorus. 
As the panicle grew and matured in both of these plants, there was 
a transportation of both the phosphorus and the protein to this part 
of the plant. 

The mobilisation of phosphorus and changes in its form in 
Vicia faha and other plants were also studied by Iwanow. The 
most interesting part of his contribution is the relation between 
sunlight and changes in the form of phosphorus. The plants which 
remained in a dark room contained more inorganic phosphorus than 



44 Literature on Inosite-Phosphoric Acid [Sept. 

those which had snnlight. Opaque shields on the leaves produced 
the same results in the protected part of the leaf, hence the change 
of inorganic into organic phosphorus may be attributed in part if not 
altogether to photosynthesis. Stoklasa and his pupils^'* consider 
phosphorus an integral part of Chlorophyll, existing in a form which 
does not give the HPO4 ion. Schimper^^ in his rather comprehen- 
sive study of the assimilation of the ash constituents in plants also 
noted the decrease of the inorganic phosphorus through the action 
of light. Posternak, attempting to account for the formation of 
"phytin," which he then thought to be anhydro-oxymethylene-di- 
phosphoric acid, assumed that it was formed simultaneously with 
the reduction of carbon dioxide by a direct combination of the orod- 
ucts of the photo-chemical action and inorganic phosphates, an 
hypothesis suggested by Schimper's experiments. If Posternak's 
assumption is true, even in part, phosphorus may play a very signi- 
ficant röle in carbohydrate anabolism. Several authors have ex- 
pressed doubt about this explanation of "phytin" synthesis, advanc- 
ing the argument that inosite-phosphoric acid is not found in the 
early stages of growth, and when formed later is not uniformly 
produced, as would be expected if it were due entirely to the action 
of the chloroplasts. There is still the possibility that it is so syn- 
thesized and instantly broken down in the formation of other Com- 
pounds. 

Soave could find no inosite in dormant seeds unless they were 
first boiled in strong acid, but after they began to sprout its pres- 
ence could be easily demonstrated until the reserve material of the 
cotyledons was almost exhausted. It was also present in unripe 
seeds, indicating that the inosite-phosphoric acid is formed in the 
seed by the combination of the inorganic phosphorus, abundantly 
present at this stage, with the inosite. The occurrence of inosite in 
the unripe seed and the green parts of the mature plant, and the later 
disappearance of this substance as inosite-phosphoric-acid -forms, 
indicate that phytin is probably produced by the reversible action 
of an intracellular phytase, and that Posternak's explanation is in- 
correct. 

Rising suggests that inosite-phosphoric acid may be an interme- 

** Stoklasa, Brdlika and Ernest : Ber. d. deut. bot. Ges., 1910, 27, i, 
"Schimper: Flora, 1890, 23, 207-261; Bot. Zeifg., 1888, 46, 81. 



igi2] 'Anton Richard Rose 45 

diary product in the formation and destruction of the lecithins, and 
in this way may play an exceedingly important part in the life cf the 
plant. The possible relation between the two Compounds is shown 
in the following graphic representation : 



HjOjPOHC— CHOPO3H, — CHOPOjHj 

HjOgPOHg (JHOPOjH, ^ CHOPOjH, 



HjOjPOHC— CHOPOjHj — CHOPO,H, 



2 




The most striking Suggestion as to the functions of inosite-phos- 
phoric acid is contributed by Starkenstein, who thinks that the 
inosite is in itself inert and incidental and functions only in its com- 
bination with phosphorus. He has demonstrated that, in the body, 
inosite yields lactic acid, an interesting fact in view of its possible 
significance in carbohydrate formation. He assigns to inosite- 
phosphoric acid, as its special function, some part in the process of 
growth, basing his view on his experiments with animals. In har- 
mony with this view is the distribution of phosphorus in the seed, 
the greater part being localized in the germ; according to Bernardini 
over eighty per cent. of the total phosphorus in the rice germ is in 
the form of inosite-phosphoric acid. It is interesting to note in this 
connection the Observation of Iwanow that there is a tendency to 
concentration of phosphorus in the parts of the plant where growth 
is most active, and also that when the phosphorus supply of the Sub- 
strate is insufficient, the phosphorus of the other parts of the plant 
is rapidly transported to the growing shoots. As previously stated 
the phosphorus of the seed is in the form of the calcium-magnesium 
Salt of inosite-phosphoric acid, but, according to Posternak, the 
phosphorus, in transportation, is in the potassium salt of this acid. 

Phyto-phosphoric acids, whether they are inosite esters or other 
Compounds, undoubtedly play very significant roles in all higher 
plants, but as their specific functions have not as yet been ascertained, 
even the chemical structures being as yet uncertain, nearly all State- 
ments on the subject must be pure conjectures. The chief sug- 
gestions from experimental work are that these acids are concerned 
in the process of photo-synthesis or in the changes of the photo-syn- 
thetic products, for example, the formation of carbohydrates and 
fats; that it is an intermediary step in the synthesis of phospho-pro- 
teins and lipoids; and that it acts as a specific Controlling factor in 



4^ Literature on Inusite-Phosphoric Acid [Sept. 

growth. The varioiis functions as thus outlined are probably over- 
estimated, but those who have worked in this field seem to be 
strongly of the opinion that inosite-phosphoric acid is more than 
a reserve material. It is attracting considerable attention and as the 
necessary analytical methods are perfected, we may expect to 
see, in increasing number, valuable contributions that will eluci- 
date in detail the part which this interesting Compound plays in 
nature. 

BIBLIOGRAPHY OF INOSITE-PHOSPHORIC ACID ("PHYTIN") 

Anderson. N. Y. Agr. Exp. Sta., Geneva, N. Y. ; A''. Y. Agr. Exp. Sta. Tech. 
Bull, ig, 21, 1912; /. Biol. Chem., 1912, 11, 471-487; 12, 97-113, and 447-464. 
Important contribution on the salts of inosite-phosphoric acid, also impor- 
tant synthetic work. 

Aso and Yoshida. Imp. Univ., Tokio; /. Coli. Agr., Imp. Univ., Tokio, 1909, i, 
153-168. Compares the value of " phytin " with other forms of phosphorus 
as a fertilizing material. 

Balika-Iwanowska. Agr. Chem. Inst., Krakow; Ras. Akad. Univ., 1906, 2° ser, 
6, B, 24. (From Maly's Jahrsb., 1907, 36, 741.) Gives the influence of the 
HPO4 ion on plant growth; also the changes in form and distribution of 
phosphorus at the various stages of development of barley plants. 

Bernardini. Chemica Agr. Scuola, Portici ; Atti. Acc. Lincei, 1912, 21, 1°, 283-289. 

Bernardini and Morelli. Atti. Acc. Lincei, 1912, 21, 1°, 357-362. 

Boorsma. Batavia ; Von Bemmelen Festschrift, 210-213 ; (see Chem. Centr., 19H 
(I), 296). Preparation and properties of the inosite-phosphoric acid from 
rice bran. 

Carte. Bull. soc. chim., 1901, (4), 9, 195-199. (From Chem. Zentr., 1911 (I), 
1196.) Repeats the work of Contardi and reports negative results. 

Collison. Ohio Agr. Exp. Sta., Wooster, O. ; /. Biol. Chem., 1912, 12, 65-73; 
/. Ind. and Eng. Chem., 1912, 4, 606-608. Modified method of determining 
inorganic phosphorus in the presence of phyto-phosphates. Cf. Forbes. 

Contardi. Lab. di chim. org. della Reale Scuola Agr., Milan; Atti. Acc. Lincei, 
1909, 5 ser., 18 (1° sem.), 64-67; 1910, 5 ser., 19 (1° sem.), 823-827. Prepa- 
ration of inosite-phosphoric acid from rice bran, analyses of the pure prepa- 
ration, synthesis of phytin by heating inosite with phosphoric acid. Syn- 
thetic product gave same analytic data as the preparation from rice bran. 
Important papers. 

Cook. U. S. Dept. Agr., Washington, D. C; Bur. of Chem., U. S. Dept. Agr., 
Bull. 123, 1909. Feeding experiments on rabbits. 

Dox and Golden. la. Agr. Exp. Sta., Ames, la. ; /. Biol. Chem., 191 1, 10, 183- 
186. Demonstration of the presence of phytase in certain common fungi. 

Donath. Wien. klin. Woch., 191 1, 24, 1192-1197. Therapeutic. 

Fingerling and Hecking. Versuchsstation, Hohenhain; Biochem. Zeit., 1912, 
37. 452. Contains a note on Stutzer's method. 

Forbes, Lehmann, Collison and Whittier. Ohio Agr. Exp. Sta., Wooster, 



1912] Anton Richard Rose 47 

Ohio; Ohio Agr. Exp. Sta. Bull., 215, 1910. Gives a good method for the 

determination of inorganic phosphorus. See Collison. 
Fürst. Centr. Kinderheilk., 1904, 409. Therapeutic. 
Giacosa. Giorn. della Real. Accad. di med. di Torina, 1905, 68, 369-374; 1907, 

70, 290-295. (From Maly's Jahrsb., 1906, 35, 124 ; 1908, 37, 473 ; also Biochem. 

Centr., 6, 573.) Pharmacological study on man. 
Gilbert and Posternak. L'Oeuvre Medico-chirurgical, 1903, No, 36. (From 

Maly's Jahrsb., 1904, 34, 729. Therapeutic. 
Gilbert and Lippmann. La Presse Medicale, 1904, Aug. 2y and Sept. 10. Phar- 
macological studies on rabbit and guinea pig. 
Hardin. S. C. Exp. Sta., Fort Hill, S. C. ; 5". C Exper. Sta. Bull., n. s. 8, 1892. 

Pyrophosphoric acid in cottonseed meal; probably associated with phytin. 

Cf. Crawford, /. Pharm, and Exp. Ther., 1910, i, 519. 
Hart, McCoUum and Humphrey. Wis. Agr. Exp. Sta., Madison, Wis. ; Wis. 

Agr. Exp. Sta. Research Bull., 5, 1909; Am. J. Physiol., 1909, 23, 86-102; 24, 

246-277. Feeding experiment with the cow. Cf. Jordan. 
Hart and Andrews. N. Y. Agr. Exp. Sta., Geneva, N. Y. ; A''. F. Agr. Exp. 

Sta. Bull., 238, 1903; Am. Chem. Jour., 1903, 30, 470-486. Gives the first 

approximately reliable method of determining inorganic phosphorus in the 

presence of phytin and other organic forms of phosphorus. 
Hart and Tottingham. Univ. of Wis., Madison, Wis.; /. Biol. Chem., 1909, 6, 

431-444. Determination of the amounts of phytin in certain feeding stuffs. 
Hartig. Braunschweig; Bot. Ztg., 1855, 13, 881-882; 1856, 14, 257-355. Micro- 

scopic study of seeds; discovery of the substance later known as phytin. 

See also "Lehrbuch der Anatomie und Physiologie der Pflanzen," 1891, 48. 
Horner. Pathol. Inst., Univ. of Berlin ; Biochem. Zeit., 1906, 2, 428-434. Phar- 
macological studies on dog and rabbit. 
Iljin. Russ. Wratsch., 1906, No. 13, from Maly's Jahrsb., 1907, 36, 54. Com- 

pares the properties of phytin, lecithin and nucleoprotein. 
Iwanoff. Jahrb. wiss. Bot., 1901, 36, 355-379; /. Exp. Agr. (Russian), 1902, i 

(cited by Zaleski) ; Ber. bot. Gesell., 1902, 20, 366-372; Zeit, physiol. Chem., 

1907, 50, 281-288. Studies on the changes of the forms of phosphorus in 

germinating vetch seeds and in the growing plant. Production of phyto- 

phosphates by yeast-fermentation of sugar in the presence of di-sodium 

phosphate. 
Jegorow. Landw. Inst. Petrowskoje-Rasumowskoje, Moskow; Biochem. Zeit., 

igi2, 42, 432-439. Study on stability of inosite-phosphoric acid and disput- 

ing the existence of phytase. 
Jordan, Hart and Patten. N. Y. Agr. Exp. Sta., Geneva, N. Y. ; N. Y. Agr. 

Exp. Sta. Tech. Bull, i, 1903; Am. J. Physiol., 1906, 16, 268-313. Feeding 

experiments with the milk cow. Cf. Hart. 
Korolev. Moscow. Izv. Moscov. Selsk. Khoz. Inst., 1910, 16, 1-98. (From 

Chemical Abstracts, 191 1, 1962.) Studies on organic phosphorus in the soil. 
LeClerc and Cook. U. S. Dept. Agr., Washington, D. C. ; /. Biol. Chem., 1906, 

2, 203-217. Feeding experiments on rabbits. 
Levene. Rockefeller Inst., N. Y. ; Biochem. Zeit., 1909, 16, 399-405- Studies 

of phytin preparations made from hemp. Obtained a Compound consisting 

of three groups : inosite, pentosan and phosphate. 
Maestro. Lo Spermentale, 1904, 59, 456-458. (From Maly's Jahrsb., 1905, 35, 

91.) Pharmacologic. 



48 Literature on Inosite-Phosphoric Acid [Sept. 

McCoUum and Hart. Univ. of Wis.. Madison, Wis. ; /. Biol. Chem., 1908, 4, 
497-500. Showing the presence of phytase in animal tissues. 

Mendel and Underhill. Yale University; Am. J. PhysioL, 1906, 17, 75-88. In- 
fluence of phytin on bacteria; pharmacological studies on dog and rabbit. 

Nagaoka. Imp. Univ., Tokio; Bull. Coli. Agr., Tokio, 1906, 6, No. 3. Value of 
inosite-phosphoric acid and other phosphorus Compounds in plant waste 
products as fertilizers, as compared with animal wastes, showing that the 
latter are the more efficient. 

Neuberg. Pathol. Inst., Univ. of Berlin; Biochem. Zeit., 1908, 9, 557-560; 1909, 
16, 405-410. Analyses of several preparations of phytin. Concludes that 
the substance is inosite-phosphoric acid and does not contain a carbohy- 
drate group. 

Novi. Pharm. Lab. Bologna; Mem.r. accad. sei. Bologna, igii,^,s^r. 6. (From 
Zentr. Biochem. u. Biophys., 1911,11,871.) Influence of phytin and glycero- 
phosphates on muscle reaction. 

Palladin. Zürich Polytechnicum ; Zeit. Biol., 1894, 31, 191-203. Discovery of 
inosite-phosphoric acid by chemical procedure. 

Patten and Hart. N. Y. Agr. Exp. Sta., Geneva, N. Y. ; A''. Y. Agr. Exp. Sta. 
Bull., 250, 1904; Am. Chem. Jour., 1904, 31, 564-672. An important contri- 
bution on the properties and composition of inosite-phosphoric acid. 

Peters. Allg. med. Centralzeitg., 1908, No. 9. (From Centr. Nervenheilk. u. 
Psychiatrie, 1908, 31, 1081.) Therapeutic. 

Pfeffer. Marburg; Jahrb. wiss. Bot., 1872, 8, 429-574. Comprehensive study of 
aleuron grains, identification of "globoid," and approximation of its chem- 
ical nature. 

Polacci. Royal Bot. Inst., Univ. of Pavia; Malphigia, 1894, 8, 361-379. Study 
of phosphorus in the aleuron grain. 

Posternak. Zürich Polytechnicum; Pasteur Inst.; Basel; Compt. Rend., 1905, 
140, 322-324; 1903, 137, 202-203; 2>37-2>29; 439-441; Bul. soc. chim., 1904, 33» 
116; Rev. Gen. Bot., 1900, 12, 5-24; 65-73; Compt. Rend. Soc. Biol., 1903, 55, 
1190-1192. German Patents, Kl. 12, Nos. 155798, 159749, 160470, 164298; 
Münch. med. Woch., 1907, p. 827. The first and most comprehensive study 
of the physical and chemical properties of phytin, giving methods of prepa- 
ration, results of analyses of pure products, and speculations on the Consti- 
tution and biological function of this product. 

Rising. St. Albonvorstadt, Basel; Svensk Kern. Tidskrift, 191 1, 22, 143-150. 
Study of organic phosphorus Compounds in food materials ; the name phyto- 
phosphoric acid is suggested, methods are given for analysis of the various 
forms of phosphorus, and analyses are reported for a preparation of the 
silver salt of inosite-phosphoric acid. 

Rogosinski. Anz. Akad. Wiss., Krakow, 1910, 260-310. (From Chem. Centr., 
1910 (II), 1549; Chem. Abstr., 1911, 5, 1476.) Feeding experiment with 
the dog. 

Rose. N. Y. Agr. Exp. Sta., Geneva, N. Y. ; N. Y. Agr. Exp. Sta. Tech. Bull., 
20, 1912. Feeding experiment with the milk cow; Dept. Biol. Chem., Colum- 
bia Univ., N. Y., BiocHEMiCAL Bulletin, 1912, i, 428-438. Influence of 
phytin on the growth of lupin seedlings. 

Sechert. These de Paris, 1904. (From Maly's Jahrsb., 1904, 34, 729.) Therapeutic. 

Schulze and Castoro. Zürich Polytechnikum; Zeit, physiol. Chem., 1904, 41, 



I9I2] Anton Richard Rose 49 

477-484. Presents a method of analysis, and data on content of inosJte- 

phosphoric acid in various seeds. 
Schulze and Winterstein. Zürich Polytechnikum; Zeit, physiol. Chem., 1903, 

40, 120-122. Phytin prepared and analyses of it made. 
Scofne. Pharm. Inst., Turin; dorn, della Real. Acc. di med. dt Torino, 190S, 

56, 630. (From Biochem. Zentr., 1905.) Fate of inosite-phosphoric acid in 

the animal organism and paths of elimination. 
Soave. Sias, spernt. agrar. Ital., 1906, 39, 413-427, 434-438. (From Chem. Zentr., 

1906, 1726.) Ann. R. Accad. di Agr. di Torino, 1906, 49, p. i et seq. (From 
Centr. Physiol., 1906, 772.) Shows the relation between inosite and phytin 
in seeds. 

Staniskis. Jagel Univ., Krakow ; Ans. Akad. Wiss., Krakow, 1909, 95-123. (From 
Chem. Zentr., 1909 (II), 114.) Determination of the distribution of the 
forms of phosphorus in millet at various stages of development. 

Starkenstein. Pharm. Inst., Univ. of Prague; Zeit. exp. Path. u. Therapie, 
1908, 5, 378-389; Biochem. Zeit., 1910, 30, 56-98; 191 1, 32, 234-265. A study 
of inosite and its relation to animal and plant life; the relation between 
inosite and inosite-phosphoric acid — chemical, biochemical and biological ; 
the nature and Constitution of phytin; its significance in animal and plant 
growth ; toxicity ; reaction to common indicators ; and estimation by Volu- 
metrie methods. A noteworthy contribution. 

Streffer. Zentr. ges. Therapie, 1908, 25, 135. Therapeutic. 

Stutzer. Biochem. Zeit., 1908, 7, 471-487. Methods of analysis. 

Suzuki, Yoshimura and Takaishi. Imp. Univ., Tokio; Bull. Coli. Agri., Tokio, 

1907, 7, 495-502, 503-512. Preparation of inosite-phosphoric acid from rice 
bran; discovery of phytase; Suggestion of a new formula, containing the 
inosite nucleus. 

Tsuda. Imp. Univ., Tokio; /. Coli. Agri., Tokio, 1909, i, 167-168. Study of the 

forms of phosphorus in vegetable wastes. 
Tyshnjenko. Therap. klin. militarmed. Akad., St. Petersburg; Dissertation 

(Russian), 1909, p. 117. (From Maly's Jahrsh., 1909, 39, 589.) Feeding 

experiments on man. 
Vorbrodt. Univ. Krakow; Bull, de l'Acad. des Sei. de Cracovie, 1910, ser. A, 

414-511. A study of the general reactions of phytin; comparison of methods 

of analysis, and Suggestion of a desirable modification; the phytin content 

of a number of seeds reported; comparative study of phytases; ultimate 

composition of phytin and a proposed empirical formula. An excellent 

paper. 
Weismann. Therap. Monatshefte, 1908, 22, 470. Therapeutic. 
Windisch. Jahrb. Vers. u. Lehrs. f. Brauerei, Berlin, 1907, 10, 56-58. (See 

also Wochschr. Brauw., 1906, 23, 516; Chemical Abstracts, 1907, i, 81.) 

Showing that the inosite-phosphoric acid of barley does not pass into beer 

but disappears in the malting process. 
Winterstein. Zürich Polytechnikum ; Ber., 1897, 30, 2299-2302. The first inten- 

tional preparation of phytin; analyses of the material; cleavage products; 

introduction of the name inosite-phosphoric acid. Zeit, physiol. Chem., 1908, 

58, 118-121. Discusses the Constitution of phytin. 



A NEW TYPE OF ARTIFICIAL CELL SUITABLE FOR 
PERMEABILITY AND OTHER BIOCHEMICAL 

STUDIES 

E. NEWTON HARVEY 
(Physiological Lahoratory, Princeton University) 

Research on the permeability of membranes has been largely 
confined to a study of the properties of what may be termed macro- 
scopic membranes; composed of parchment, collodion, rubber, or 
silk impregnated with various substances. The best example of 
membranes of a type and size comparable to the surface film of cells 
and yet available for permeabiHty studies are the precipitation 
membranes of Traube, investigated by Waiden, Tamann, and 
Meerburg. 

Protein membranes of exceeding fineness are formed at the 
surface of various non-miscible fluids shaken with protein Solutions, 
such as the surface film of oil globules in protein-oil emulsions, or 
the films formed on Chloroform or benzol when shaken with albu- 
men Solutions.^ Such membranes are useless for permeability 
studies so long as they Surround fluids that do not mix with water. 
However, it is an easy matter to replace the fluid within the mem- 
brane by a watery Solution, provided the former fluid is readily 
volatile and slightly soluble in water. Chloroform conforms to 
these conditions. When Chloroform is shaken with egg albumen 
Solutions, the globules, in the course of 10-15 minutes, shrink in 
size and their membranes become crumpled, due to the passage of 
Chloroform from water to air and from globule to water. Lecithin, 
if previously dissolved in the Chloroform, will take up water as the 
Chloroform passes out. In the course of one to two hours, in an 
open vessel, all the Chloroform disappears and we obtain, instead 
of a Chloroform Solution of lecithin, a water Solution of lecithin 
enclosed in a fine protein membrane, the whole of a size comparable 
with cell sizes. The diameter of the droplets may be varied at will 

^ Robertson : Journal of Biological Chemistry, 4, p. i, 1908. 

50 



igi2] E. Newton Harvey 5^ 

according to the degree of shaking. The role of the lecithin is to 
hold the water as the water replaces the Chloroform. The protein 
membrane is impermeable to lecithin. 

These artificial lecithin cells are stable, persisting until destroyed 
by bacteria. In many ways — in shape, in general appearance and in 
consistency — they resemble, to a very remarkable degree, sea-tirchin 
or star-fish eggs. Some of their properties have been described in 
Science (n. s.), V'ol. 36, p. 564, 1912. 

The point I wish to emphasize here, however, is not that we 
can prepare artificial cells closely resembling real cells, but that a 
Solution of lecithin may be obtained within a protein membrane, 
the whole of known composition and of a size comparable with cell 
sizes. Much can be inferred concerning the living cell from a 
knowledge of the properties of such artificial cells where compo- 
sition is definitely known. 

As Chloroform is exchanged for water, some of the lecithin 
separates in the form of granules, most of which agglutinate in a 
dense clump. The cell as a whole, but more particularly these 
granules, take up neutral red from dilute Solution, becoming red 
in color. (Chloroform alone takes up only the yellow base of neu- 
tral red. When lecithin is dissolved in Chloroform it unites with 
the yellow base, forming a red salt.) 

If the permeability for alkalies of such red-stained cells is 
studied, a marked difference from that of living cells is noted. 
Both ammonium hydroxid and sodium hydroxid in w/2000 con- 
centration enter rapidly and at the same rate. It will be remem- 
bered that all living cells are very easily permeable to ammonium 
hydroxid, but very slightly so to sodium hydroxid.^ The surface 
membrane of living cells is evidently of quite different composition 
from the protein film which condenses on Chloroform droplets. 

Living cells behave toward alkalies as though they were sur- 
rounded by a layer of a fat solvent, as postulated by Overton. 
Lipoid-soluble alkalies (ammonium hydroxid) penetrate readily, 
lipoid-insoluble alkalies (sodium hydroxid) do not. The lipoid 
solubility of ammonium hydroxid can be readily demonstrated by 
means of a benzol-lecithin Solution shaken with egg albumen solu- 

^ Harvey, E. N. : Journal of Experiniental Zoology, 10, p. 507, 1911 and 
BiocHEMicAL Bulletin, i, p. 227, 1911. 



52 New Type of Artificial Cell [Sept. 

tion. The same type of protein-film is formed on these globules 
but they differ from chloroform-lecithin globules in that the benzol 
is not replaced by water. If the benzol-lecithin globule is stained in 
neutral red Solution and placed in 7?/iooo ammonium hydroxid 
Solution, the color change from red to yellow takes place almost 
instantly. But it is only after 15-20 minutes that sodium hydroxid 
in relatively high (;i/io) concentrations can enter. Ammonium 
hydroxid is readily soluble in the benzol droplet, while sodium 
hydroxid is not; and in this fact lies the explanation of the differ- 
ence in penetrability. When stained in neutral red Solution, prac- 
tically all living cells behave as though they were protected from 
alkali by a benzol-lecithin surface layer. 

It is a simple matter to introduce various substances into these 
cells by dissolving or suspending the material in the chloroform- 
lecithin Solution before it is shaken with the protein Solution. 
Thus, oil may be dissolved by Chloroform and will separate in the 
cell in several large droplets much like those in a Nereis egg. Or 
cholesterol, starch grains and finely divided protein particles can be 
likewise included ; or substances to be used as indicators in study- 
ing the permeability of the protein film. 

Such cells, regarded as complex Systems of biological sub- 
stances, offer exceptional advantages for interpreting phenomena 
observed in living cells under special conditions; for example, dur- 
ing the passage of an electric current. Movements and disintegra- 
tions take place which I have as yet only partially investigated. In 
the near future I intend to describe these phenomena and shall give 
more complete data upon the permeability of the film which sur- 
rounds the cells. 



ON A NEW FUNCTION OF THE CATALYZER CALLED 

" PEROXIDASE " AND ON THE BIOCHEMICAL 

TRANSFORMATION OF ORCIN TO ORCEIN^ 

JULES WOLFF 

In a recent publication I have described the influence which 
peroxidase exerts on certain phenols in the presence of various salts 
and alkalies.^ When dissolved in a weak sodium carbonate Solu- 
tion freely exposed to the air, orcin, for example, fixes from four 
to five times more oxygen in the presence of peroxidase than in its 
absence. In this note I wish to call attention to the fact that perox- 
idase has other powers than the fixation of atmospheric oxygen. 

In determining the combined influence of ammonia and perox- 
idase upon aqueous Solutions of orcin, I have studied conditions 
which favor the transformation of orcin^ into orcein, the beautiful 
coloring matter which is one of the principal constituents of com- 
mercial orseille. 

My observations are interesting from many points of view. 
They show that (i) if a 2 per cent. Solution of orcin is exposed 
to the air in a thin layer and subjected to the influence of different 
proportions of ammonia, orcein is not formed even after a month 
under such conditions, but, instead, there is produced a substance 
which imparts a brownish-red color to the liquid. (2) If, how- 
ever, a portion of the same Solution is put in a narrow tube, so that 
the reaction takes place in a deeper layer, and the surface of con- 
tact with air is limited (other conditions being equal), one observes 
a very slow but regulär formation of orcein. (3) If (everything 
eise being equal) one repeats the first experiment (i), but adds to 
the ammoniacal Solution of orcin a suitable quantity of peroxidase, 
orcein fails to appear, just as in the first experiment. (4) If, how- 

* Translated from the author's manuscript, in French, by Dr. J. J. Bronfen- 
brenner. [Ed.] 

''Wolff: Comptes rendus, 1912, clv, p. 618. 

'Orcin has been the subject of interesting work by Robiquet, Dumas, Liebig, 
and Laurent and Gerhardt. 

53 



54 A New Function of " Peroxidase" 

ever (all other conditions being eqiial), one repeats the second ex- 
periment (2), but adds to the ammoniacal Solution of orcin a siiitable 
amount of peroxidase, the transformation into orcein takcs place 
very rapidly and is quite advanced in four or five days^ Compar- 
ing the coloration intensities of products 2 and 4, one sees that in 
five days 4 contains more than twice as mtich orcein as 2. This 
gain is due to the action of the peroxidase. By boiling the perox- 
idase Solution for f rom 5 to 6 minutes, before adding it to the ammo- 
niacal Solution of orcin in experiment 4, there is no acceleration in 
the formation of orcein, evidently because the peroxidase, as the 
active agent in the transformation, is thus destroyed. 

In Order to determine the rate of oxidation in the different ex- 
periments, I measured the volumes of absorbed oxygen. In experi- 
ment 2, eight to nine times more oxygen was absorbed than in i, 
in the course of 48 hours. In experiments 3 and 4 there was a 
similar difference but, because of the presence of peroxidase, the 
proportions of absorbed oxygen were larger. 

Without discussing the nature of the combined action of am- 
monia, oxygen, and peroxidase upon orcin, we may conclude that 
in dilute Solutions of orcin, slozv oxidation by ammonia is the pri- 
mary condition for the formation of orcein. If, however, to this 
condition is added the accelerating influence of peroxidase, the 
action is directed toward formation of coloring matter rather than 
toward increased absorption of oxygen. These facts may possibly 
be helpful in the commercial preparation of orseille. 

Paris, France. 

* My first and third experiments could easily be performed in flasks with flat 
bottoms. A small test tube would be satisfactory for experiments 2 and 4. 
The proportions of materials indicated below are well adapted for the purposes 
of the experiments : 

^ , ( 2 c.c. of 2.8 per Cent. Solution of ) tnm x j -.i 

For I and 2 < , , , ^j^ > Diluted with water to 3.5 c.c. 

( orcm and 50 mg. of NH3 ) 

2 c.c. of 2.8 per Cent. Solution of 
For 3 and 4 ■{ orcin, 50 mg. of NH3 and \- Diluted with water to 3.5 c.c. 

I c.c. of active peroxidase Solution 



STUDIES OF DIFFUSION THROUGH RUBBER 

MEMBRANES 

I. Preliminary observations on the diffusibility of lipins and 

lipin-soluble substances 

WILLIAM J. GIES 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

CONTENTS 

I. Introduction c :- 

IL On the diffusibility of biological substances through rubber cg 

III. A demonstration of osmotic pressure exerted by fat 50 

IV. A demonstration of the diffusion of pigments from fat through rubber 

into fat 60 

V. Comparative dialysis experiments, with demonstrations 61 

VI. Experiments on the diffusibility of alkaloids through rubber 62 

I. INTRODUCTION 

When I proposed to my biochemical associates in Cancer re- 
search, in 1909, that we undertake a study of intracellular chem- 
istry/ I realized that new analytic methods and unconventional 
experimental procedures were prerequisites for material progress in 
this as in any other chemical relation. The greatest obstacle in the 
path of progress in intracellular chemistry is the evident lability of 
the essential intracellular constituents. Our best chemical methods 
increase this predicament because each is essentially anti-biological 
in character. Biochemical discoördinations are enforced whenever 
any of our present chemical processes is efifectively applied to proto- 
plasmic material. 

In reflecting on the properties and possible coördinations of 
intracellular lipins, it seemed probable that such lipins might be 
separated from protoplasmic material with the least chemical vio- 

* Gies : Studies in cancer and allied subjects, conducted under the auspices of 
the George Crocker Special Research Fund; Volume III, Department of Bio- 
logical Chemistry, Introduction (in press). 

55 



56 SUidies of Diffusion through Rubber Membrancs [Sept. 

lence, and isolated with the least possible alteration of their qualities, 
if they cotild be removed by dialysis.^ When this idea first came to 
mind, however, execution of its essential feature appeared to be im- 
possible. I believed that the diffusion of a solute depends very 
largely on chemical affinity between the separating membrane and 
the solvents on both sides of the partition. In that view, it seemed 
highly improbable that any of the ordinary membranes, except 
possibly collodion, could be of Service in the dialysis of lipins under 
any circumstances. Collodion appeared to possess favorable quali- 
ties because of its solubility in common lipin solvents and its pos- 
sible affinity for the latter under conditions of dialysis.^ 

Collodion is the only one of the available membranes which, 
while soluble in ether-alcohol Solutions, freely permits the passage 
of salins, extractives, carbohydrates, and proteins from aqueous 
Solutions to water, or to aqueous Solutions, outside, and vice versa, 
At first thought this suggested special availability of collodion for 
the work in mind. On the other hand lipins could not be expected 
to dialyze through collodion in the presence of much water and, as 
preliminary dehydration seemed an inevitable necessity for the 
dialysis of lipins from cellular matter, the permeability of collodion 
membranes to zoater-soluble substances did not appear, after all, to 
imply any practical advantages for the diffusion of lipins. I also 
recalled the fact that, in some experiments in another relation, we 
found that collodion was occasionally rendered defective by ether 
when the latter was used as a preservative of aqueous Solutions 
undergoing dialysis.^ 

Continuing actively to consider these matters from one view- 
point and then another, I thought of rubber as a possible choice of 
membrane. Recalling the well-known fact that rubber swells very 
markedly in ether and even in ether vapor, I assumed that the rub- 
ber expands in ether under such conditions because ether dissolves 
in the rubber or combines with it. This was but the prelude to the 

' After preliminary desiccation by treatment with anhydrous sodium sulfate 
or other suitable process. 

° Collodion is a serviceable membrane for such purposes. See page 70. 

* In some experiments which Professor Welker has conducted at my request, 
we have found that the disintegrative effect of ether on collodion membranes 
may be due to contained alcohol and other impurities. (See page 70.) 



igi2] William J. Gies 57 

deduction that if ether dissolves in or combines with rubber, ether 
would also carry dissolved lipins with it into a rubber membrane; 
and if ether were on the opposite side of such a membrane, to work 
inwardly under such conditions, ether currents would develop; and 
lipins would pass from the Solution o£ higher concentration to that 
of the lower, and there accumulate until an equilibrium was estab- 
lished. 

This conception was so attractive that I proceeded at once to 
State it to Dr. Rosenbloom and, with his Cooperation, immediately 
tested it. The solid residue from an evaporated ether extract of 
egg yolk offered the greatest advantages for a preliminary test. 
We accordingly made an ether Solution of such a yellow residue, 
transferred the deep yellow Solution to a rubber condom, immersed 
and supported the latter in ether in a stoppered bottle, and almost 
immediately observed diffusion currents as well as the rapid egress 
of lipochrome. Fat and cholesterol were easily detected in the 
diffusate. 

Assuming that this prompt positive result might be due to defects 
in the rubber, we made many tests to satisfy ourselves that the ob- 
servations were or were not what they appeared to be. Dr. Rosen- 
bloom gave very earnest attention to this phase of the matter for 
some time and established the fact that we were dealing, except in 
a few cases of obviously imperfect membranes, with true diffusion 
phenomena. 

The original experimental observations were made on March i, 
1910. At that time I was ignorant of similar results of previous 
work with rubber membranes, although I recalled rather vaguely the 
fact that Kahlenberg had made use of such membranes in another 
connection. The references to Kahlenberg's work which are given 
in the Chemisches Zentralblatt [1906 (2), pp. 1391 and 1772], the 
only ones we could find on this subject at that time, satisfied us that 
if we extended these experiments, the observations of a previous 
observer would not be repeated.^ A month or two after the work 

"The references to which I allude gave the substance of a paper in the 
Transactions of the Wisconsin Academy of Sciences, Arts, and Letters, 1905, xv 
(i), pp. 209-272, entitled: " On the nature of the process of osmosis and osmotic 
pressure, with observations concerning dialysis." The results with which our 
own could be directly compared were the f ollowing ones : Copper oleate was 



$8 Stiidies of Diffusion through Rubber Membranes [Sept. 

was inaugiirated we also saw a late reference to the well-known 
fact, regarding the swelling of rubber in lipin solvents, on which our 
work was based.*^ Ten months later we demonstrated these findings 
at a meeting of the American Society of Biological Chemists (see 
page 64). 

The succeeding sections of this paper present reprinted prelim- 
inary reports on various portions of the studies which thus far have 
developed from the observations described above. It is my Inten- 
tion to discuss in detail each seotion of the work, and additional ex- 
periments, at the earliest opportunity, when I hope to dwell more 
particularly on the significance of such results for the Student of the 
functions of cell membranes, and for the investigator of the co- 
ordinations and equilibria in intracellular affairs. 

IL ON THE DIFFUSIBILITY OF BIOLOGICAL SUBSTANCES 

THROUGH RUBBER' 

The writer and his associates have f ound that many ether-soluble 
substances of biological origin, such as fat and cholesterol, pass 
readily from ether Solutions through rubber membranes into ether 
when the mechanical conditions for such diffusions are favorable. 
Lecithans appear to be wholly indiffusible. 

Many substances which are soluble in fatty oils, Chloroform, al- 
cohol, acetone, ethyl acetate, and other solvents of similar powers, 
or in mixtures of such solvents, promptly diffuse through rubber 
under suitable conditions. Collodion is one of the products which 
appears to be indiffusible under such circumstances. When an ordi- 
nary ethereal Solution of collodion (containing 24 per cent. of alco- 
hol) is dialyzed in a rubber Condom against ether in a closed vessel, 
the alcohol rapidly passes to the exterior and the collodion gradually 
gelatinizes. Liquid accumulates in the bag under these conditions. 

Various inorganic substances diffuse through rubber under the 

found to diffuse from benzene through a rubber membrane into benzene; and 
camphor diffused from pyridin, alcohol, and toluol through rubber membranes 
into the same solvents, respectively. A recent study of Kahlenberg's paper in the 
original makes it evident that our results may be explained on the theory of 
diffusion which Kahlenberg has done much to render convincing. 

'Flack and Hill: Journal of Physiology, 1910, xl, p. xxxiii. 

' Gies : Proceedings of the Biological Section of the American Chemical 
Society: Science, 1911, xxxiv, p. 223; Biochemical Bulletin, 1911, i, p. 125. 



igi2] William J. Gies 59 

conditions mentioned above. Ferric sulphocyanate readily passes 
from ether Solution through rubber into ether. 

The writer inaugurated these studies, with Dr. Rosenbloom's 
Cooperation,^ in the hope of devising improvements in the methods. 
for the isolation of Hpins. The work is progressing along several 
lines, especially with reference to methods of isolation and purifi- 
cation, and to osmosis (see page 64). 

III. A DEMONSTRATION OF OSMOTIC PRESSURE EXERTED 

BY FAT' 

In the first of two demonstrations, a cylindrical rubher bag (Con- 
dom), 13^ inches in diameter and 8 inches long, was lowered into an 
oiled miislin bag of aboiit the same dimensions. The rubber bag 
was then filled to overflowing with olive oil. The rubber bag ex- 
panded, as the oil filled it, to the füll length and width of the muslin 
sheath. The sheath prevented further extension of the rubber bag 
and imparted rigidity to the Osmometer that was ultimately con- 
structed. The double bag, füll of oil and with its mouth wide open, 
was then raised so as to inclose about an inch of the lower end of 
a long glass tube which was firmly supported vertically above the 
demonstration table. The glass tube was 5 feet long and its bore 
was 4 mm. in diameter. Ligatures were tightly secured around the 
neck of the double bag against the immersed lower end of the verti- 
cal tube. The bag then hung directly from the end of the tube. 
The bag and its sheath were in a tightly distended condition and a 
stationary column of oil an inch high in the tube was visible above 
the protruding edge of the sheath. The tube and bag were then 
lowered into a salt-mouth liter bottle on the table until the bag 
almost touched the bottom of the bottle. The height of the bottle 
and the length of the bag were nearly equal. The tube was then 
marked with a label on the plane of the oil meniscus just above the 
neck of the bag, and enough ether was poured into the bottle to pro- 
vide Immersion for the bag to the depth of an inch. For a mo- 
ment no change in the volume of oil was apparent, and the lateral 

^Rosenbloom and Gies: Proceedings of the American Society of Biological 
Chemists, 191 1, ii, p. 8; Journal of Biological Chemistry, 191 1, ix, p. xiv. 

•Rosenbloom and Gies: Proceedings of the Society for Experimental 
Biology and Mediane, 191 1, viii, p. 71. 



6o Studies of Diffusion through Rubber Memhranes [Sept. 

pressure of the ether was obviously without mechanical effect. But 
in a miniite or two downward diffusion currents were visible along 
the surface of the bag and oil rose rapidly in the tube. 

After the initial effects of the ether had been shown, the bottle 
was filled with ether containing Sudan III, and a 5-foot vertical 
extension of the same bore was added to the upright glass tube. 
In a moment the upward movement of the liquid was markedly 
accelerated. 

The demonstration was started at about 9 p. m. At 10 p. m. 
the osmotic pressure had carried the column of oily fluid to the top 
of the lo-foot tube, and liquid continued to run rapidly from the 
Upper orifice until the apparatus was dismantled after the adjourn- 
ment of the meeting, at about 11.30 p. m. 

During the progress of the demonstration, Sudan III diffused 
rapidly from the exterior, through the rubber, to the very top of the 
rising column of fluid, before any of the liquid passed out of the 
Upper opening. Oil diffused rapidly through the rubber into the 
ether. 

The second demonstration was essentially the same in principle 
and technic as the first. Instead of a lo-foot upright tube, however, 
the authors substituted an L tube with an inside diameter of 6 mm. 
The vertical extension of the tube was 17 inches, the horizontal ex- 
tension was only 3 inches. The latter extension was drawn out to 
a narrow bore in an inclined plane, to facilitate direct delivery of 
any liquid that might pass through that end of the tube. 

When partial immersion of the bag first occurred there was no 
visible response, but, in a minute or two, oil began to rise in the 
tube. The bag was then completely covered with ether. The up- 
ward movement proceeded rapidly ; and in about an hour nearly 200 
c.c. of liquid passed through the upper orifice into a graduated cylin- 
der which was supported underneath the outlet to catch the overflow. 

IV. A DEMONSTRATION OF THE DIFFUSION OF PIGMENTS 
FROM FAT THROUGH RUBBER INTO FAT»" 

The writer has found that many fat-soluble pigments, such as 

Sudan III and Scarlet R, diffuse readily from solid and liquid fats 

" Gies : Proceedings of the Society for Experimental Biology and Mediane, 
191 1, viii, p. 73. 



1912] 



William J. Gies 



6i 



through rubber into various solid and liquid media, among them 
both solid fat and oil. Thus, when Sudan III is dissolved in melted 
lard, the red liquid poured into a rubber bag, the bag supported in 
melted lard in a bottle, and the apparatus promptly immersed in ice 
water — the fatty matter will congeal before any sign of pigmentary 
diffusion occurs but, in a few hours, a reddish tinge will develop 
outside of the bag, and on each successive day for several weeks 
further extension of the pigmented matter may be witnes'sed, until 
the whole of the external lard is deeply suffused with red. This 
process takes place quite rapidly when the lard and apparatus are 
kept in a thermostat at 40° C. 

The demonstrations were intended to exhibit a few instances of 
such pigmentary diffusions as occur speedily enough at room tem- 



No. 


Contents of the Rubber Bag 


Nature of the 

Liquid in which 

the Bag was 

Suspended 




Oil 


Pigment 




I 
2 

3 

4 
S 


Olive oil 
Cocoanut oil 
Lard oil 
Paraffin oil 
Olive oil 


Scarlet R 
Scarlet R 
Sudan III 
Sudan III 
Sudan III 


Olive oil 
Cocoanut oil 
Lard oil 
.Paraffin oil 
Ether 


Visible diffusion of the pigment oc- 

curred promptly 
Visible diffusion of the pigment oc- 

curred promptly 
Visible diffusion of the pigment oc- 

curred promptly 
Visible diffusion of the pigment oc- 

curred promptly 
Visible diffusion of the pigment oc- 

curred almost immediately 



perature to yield positive results within an hour. The accompany- 
ing summary indicates briefly the precise nature and results of the 
demonstrations (including two control tests — 4 and 5), which were 
made with thin rubber bags in ordinary glass bottles. 

The bags were securely supported in the bottles, and the mix- 
tures were shaken occasionally during the demonstration. The 
bags were found, after the adjournment of the meeting, to be with- 
out defects. 



V. COMPARATIVE DIALYSIS EXPERIMENTS, WITH 
DEMONSTRATIONS" 

When dry bags of rubber, gold-beater's skin, parchment, and 

collodion, each containing olive oil and Sudan III, are separately 

" Goodridge and Gies : Proceedings of the Society for Experimental Biology 
and Mediane, 191 1, viii, p. 74. 



62 Studies of Diffusion through Rubber Membranes [Sept. 

immersed in olive oil, and the remaining conditions of the environ- 
ment are uniform, diffusion of the pigment promptly occurs through 
rubber, but does not take place at all through any of the other three 
membranes. When the bags are lifted from the oil, washed ex- 
ternally with ether, and then immersed in ether,^^ the pigment 
quickly passes through the rubber, but diffuses very slowly if at all 
through the remaining membranes. 

Successive immersions of nioist impermeable membranes (gold- 
beater's skin and parchment) in alcohol and ether, for different 
periods of time, fail to render the treated membranes more perme- 
able to Sudan III than before. 

The authors demonstrated the general facts in this connection 
pertaining to rubber and gold-beater's skin. 

VI. EXPERIMENTS ON THE DIFFUSIBILITY OF ALKALOIDS 

THROUGH RUBBER^" 

Various ether-soluble substances, when dissolved in ether and 
placed in rubber bags immersed in ether, readily pass through the 
rubber membranes thus imposed (I-V). We have found that 
various alkaloids and some related substances readily diffuse 
through rubber under such conditions. 

Our experiments were conducted as follows : A moderate quan- 
tity of the pure ether-soluble substance was mixed with 15 to 25 c.c. 
of ether.^^ This mixture was poured through a funnel into a new 
air-tight rubber condom in such a way as to preclude the possibility 
of overflow upon the external surface. The bag was then immersed 
in about 50 c.c. of ether in a narrow salt-mouth bottle 7 inches high. 
With the bag suspended at füll extension in this position, its mouth 
was about an inch above the opening in the bottle. The protrud- 

"In experiments which the senior author has been conducting with Prof. 
Welker's Cooperation, it has been found that collodion bags are disintegrated by 
ether containing more than about 1.5 per cent. of alcohol. Pure ether does not 
dissolve or in any way disorganize collodion membranes. A collodion bag con- 
taining pure ether may be immersed for a week or more in pure ether without 
undergoing any appreciable deterioration. (See page 70.) 

^ Sidbury and Gies : Proceedings of the Society for Experimental Biology 
and Medicine, 191 1, viii, p. 104. 

" Substances which did not dissolve readily were triturated with ether in a 
mortar. 



igi2] William J. Gies 63 

ing Condom was supported in the neck of the bottle by a tightly 
fitting cork Stopper, which also served to keep the bag closed. After 
a diffusion period of convenient length (sometimes 2 to 5 days)/^ 
the Condom was cautiously removed from the bottle, the ether diffus- 
ate was poured into a porcelain dish, and the ether completely re- 
moved by evaporation on a steam bath. At least one appropriate 
test was then applied to the residue.^^ 

Meanwhile, the ether Solution in the Condom was removed. A 
large volume of water was then poured into the suspended bag, 
which, during its distention by the water, was carefully examined 
for signs of leakage. In a few instances defective membranes tem- 
porarily rendered the outcome doubtful. All results with such bags 
were ignored, of course. Each of the tests, even after reliable pos- 
itive responses, was repeated at least once with a nezv rubber bag. 

The substances named below (the complete list of those already 
tested in this connection) are readily diffusible under the conditions 
of these experiments : — 

A. Apomorphin, atropin, brucin, caffein, Cocain, codein, col- 
chicin, coniin, morphin, narcein, narcotin, nicotin, physostigmin, 
quinin, strychnin, veratrin. 

B. Acetanilid, antipyrin, phenacetin, picric acid, picrotoxin, 
pyramidon, salicylic acid. 

Experiments with other solvents, and with additional substances 
of alkaloidal type, will be added to this series. 

" Some of the alkaloids pass through rubber almost immediately under the 
conditions of these experiments. 

*' In the experiments with nicotin, the " tobacco odor " of the concentrated 
liquids was very pronounced. 



STUDIES OF DIFFUSION THROUGH RUBBER 

MEMBRANES 

2. Diffusibility of lipins from ether through rubber membranes 

into ether 

JACOB ROSENBLOOM 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

I. INTRODUCTION 

Many experiments, in completion of the diffusion work I have 
been doing in collaboration with Dr. Gies/ have been performed to 
determine the diffusibihty or non-diffusibihty of hpins and similar 
substances (page 57). Such data must obviously be obtained in 
detail, if any attempt to devise methods for the isolation and purifi- 
cation of hpins by dialysis through rubber can be successful. 

I present here briefly the essential resuks of the work aheady 
completed in this connection. 

II. DIFFUSION EXPERIMENTS 

Methods. In the experiments described below, ordinary rubber 
Condoms were used as diffusion membranes.^ Various kinds of 
"sheet rubber," known as "pure Mexican plantation rubber," and 
fumed with carbon-disulfid, were found to be good membranes for 
this kind of work, but besides allowing fats, fatty acids, soaps, cho- 
lesterol and hpochrome to diffuse through it, this sheet rubber also 
permits the passage of lecithans under the conditions to be described, 
although the lecithans pass through the sheet rubber very slowly 
compared with other lipins such as fat and cholesterol. Condoms 

^ Rosenbloom and Gies : Proceedings of the American Society of Biological 
Chemists, 191 1, ii, p. 8; Journal of Biological Chemistry, 191 1, ix, p. xiv. 

° Bef ore the Condoms were used for this purpose, they were placed in f resh 
portions of ether daily for several days, to free them from the powder adherent 
to them. This is especially important when one proposes to test the dialysates 
for phosphorus, since the adherent powder has been found to contain phosphorus. 

64 



I9I2] Jacoh Rosenbloom 65 

do not permit the diffuslon of lecithans under the conditions of the 
tests to be described, and they were preferred for this work for that 
reason. The cause of the observed difference in permeability is 
unknown to us, but will soon be made the subject of special inquiry. 
The substances to be tested in the diffusion experiments were dis- 
solved in 100 to 200 c.c. of ether (anhydrous and distilled over 
sodium), the concentrations of the Solutions varying from 0.5 to 
5 per cent, The Solution or Suspension was carefully poured 
through a funnel into a new air-tight rubber condom in such a way 
as to preclude the possibility of overflow upon the external surface. 
The bag was then immersed in from 100 to 200 c.c. of pure ether in 
a wide-mouthed bottle of convenient size, and suspended loosely by 
a thin cord held securely between the stopper and neck of the 
tightly stoppered bottle. The bottle was kept well stoppered 
throughout the whole of each test to prevent egress of ether and 
ingress of dust and other extraneous matter. 

1. Ether extract of egg yolk. Within five minutes after ether 
extract of egg yolk is subjected to the diffusion treatment described 
above, the lipochrome appears in the dialysate, diffusion currents 
being visible about the same time. The following substances can be 
detected in the dialysate after short periods of dialysis: fat, fatty 
acid, cholesterol, and lipochrome. The lecithans do not pass 
through the Condoms, even during prolonged periods of dialysis. 

We tested for lecithans in the dialysate by analyzing the evapora- 
tion residue for phosphorus by the fusion and Neumann methods. 
and by seeking an " acetone precipitate " in the concentrated ether 
Solution after the addition of electrolyte (sodium chlorid). Some- 
times a positive phosphorus test was obtained from a dialysate 
which did not yield an " acetone precipitate." In such cases, it was 
found that this result was due to the presence of glycerophosphoric 
acid in the dialysate. If to a Solution of lecithans, which, after 
dialysis in a condom, does not yield a phosphorus Compound to the 
dialysate, one adds some glycerophosphoric acid, and then dialyzes 
this Solution through the same rubber condom, glycerophosphoric 
acid appears in the diffusate. 

2. Ether extract of brain. The dialysate from ether extracts 
of brain contained fat, fatty acid, and cholesterol. Lecithans failed 
to dialyze. 



66 Studies of Diffusion through Rubber Membranes [Sept. 

3. Ether extract of heart muscle (ox). Fat, fatty acid, lipo- 
chrome, and cholesterol were detected in the dialysate from ether 
extracts of the heart muscle of oxen. Lecithans did not diffuse. 

4. Ether extract of kidney and liver (dog). Fat, fatty acid, 
lipochrome, and cholesterol appeared in the diffusates from ether 
extracts of dog kidneys and livers. Lecithans did not dialyze. 

5. Ether extract of blood (dog). Fat, fatty acid, lipochrome, 
and cholesterol occurred in the dialysates from ether extracts of dog 
blood. No lecithans dialyzed. 

6. Ether extract of carrots. The coloring matter dialyzed 
very rapidly from ether extracts of carrots. A small amount of fat 
was also present in the dialysate. 

7. Ether extract of Xanthoma (skin). The yellow coloring 
matter dialyzed very quickly from an ether extract of xanthomatous 
skin, but it faded in twelve hours. 

8. Ether extract of cerumen. Cholesterol, fat, and fatty acid 
were present in the dialysates from ether extracts of cerumen. 
Neither the coloring matter nor the lecithans diffused. 

g. Ether extract of yeast. The dialysates from ether extracts 
of yeast exhibited a peculiar opalescence, even at the end of six 
weeks' dialysis. A small amount of fat dialyzed, but lecithans did 
not diffuse. 

10. Ether Solutions of individual substances er special prod- 
ucts. The following substances or special products, when subjected 
to diffusion by the method described above, were found to be 
diffusible : 

Acetic acid Ethyl butyrate Palmitic acid 

Acetone Formic acid Potassium palmitate* 

Beta-hydroxy-butyric acid GlyceroP Potassium stearate* 

Butter (fresh and rancid) Lactic acid Propionic acid 

Butyric acid Lead oleate Sodium palmitate* 

Cholesterol-acetate Mutton tallow Sodium stearate* 

' When ether-alcohol Solutions of glj'cerol are dialyzed against ether-alcohol, 
and alcohol Solutions of glycerol are dialyzed against ether, the dialysates contain 
glycerol. 

* Treated with water, then with alcohol to the point of precipitation, then 
with ether until a precipitate was produced. The filtrate was dialyzed against 
water, alcohol, and ether in identical proportions. 



912] Jacob Rosenhloom 6y 

Cholesterol-benzoate Oleic acid Stearic acid 

Cholesterol (from brain, Olive oil Sudan III 

egg yolk, and gall-stones) Olive oil stained with Urochrome' 

Ethyl acetate Sudan III Valerianic acid 

In some special experiments we found^ that cholesterol benzoate, 
cholesterol stearate, cholesterol oleate and cholesterol palmitate, 
when dissolved in ether, readily diffuse through rubber into ether. 
Cholesterol stearate with a molecular weight of 652.61 diffuses, 
whereas the various lecithans, with molecular weights considered 
to be 770 to 785, do not. If we assume that the diffusion of a sub- 
stance depends on the size of its molecules, the above facts 
strengthen Hiestand's conclusion that the molecular weight of ^gg- 
yolk lecithin is 1446, which figure he obtained by a molecular weight 
determination, 

II. Indiffusible substances. The following substances, when 
subjected to diffusion by the method described above, were found to 
be indiffusible.'^ 

Sodium chlorid Lecithans from yeast 

Lecithans from brain Lecithans from wheat embryo 

Lecithans from egg yolk Kephalin from brain 

Lecithans from heart muscle Cuorin from heart muscle 

Lecithans from pig testicle Compound of lecithin with platinic chlorid 

Lecithans from liver and kidney 

Koch^ has lately described the preparation of various Compounds 
with lecithans, but it is uncertain whether these Compounds are 
colloidal adsorptions, mechanical mixtures, or true chemical Com- 
pounds. It seemed of interest to study the behavior of these sub- 
stances in ether Solution, when subjected to dialysis in rubber bags 
suspended in ether. 

The preparations used in these experiments were made accord- 

' Ether-alcohol Solution (equal amounts) dialyzed against ether-alcohol. 

' Boas and Rosenbloom : Proceedings of the Society for Experimental 
Biology and Medicine, 191 1, viii, p. 132. 

''We have found that lecithans prepared by the Zuelzer, Bergeil, or Roaf 
and Edie method, when dialyzed, always yield traces of cholesterol to the 
dialysate; and often fat. 

* Koch and collaborators : Journal of Pharmacology and Experimental 
Therapeiitics, 1910, xii, 239-269. 



68 Studies of Diffusion through Rubber Membranes [Sept. 

ing to the method described by Koch. For the dialysis tests the 
Solutions of the lecithan Compounds were evaporated to dryness at 
38° and the residues triturated with ether. The extracts were fil- 
tered, and the filtrates placed inside of rubber bags and dialyzed 
against ether for thirty-seven days. The dialysates were tested 
weekly to see if the substance combined with the lecithan had 
diffused. 

Compounds of lecithin with glucose, lactic acid, strychnin, digi- 
tonin, salicin, urea, creatin, Creatinin, and caffein were prepared. 
It was found that the glucose and lactic acid dialyzed completely, 
the strychnin, digitonin, and salicin dialyzed partially, while urea, 
creatin, Creatinin, and caffein did not dialyze at all.^ 

It was thought that some of the various substances which did not 
diffuse might do so in the presence of a considerable amount of dif- 
fusible material, but on dialyzing various mixtures of the above- 
named indiffusible substances with varying amounts (up to 15 
grams), of neutral fat, fatty acid, cholesterol, or olive oil, no diffu- 
sion of lecithan occurred. 

When Solutions of lecithans are subjected to dialysis by the 
method described above, they take up a great deal of ether, and the 
volume of liquid in the bag is greatly increased. We have demon- 
strated that lipins exert strong osmotic pressure. ( See page 59. ) 

We have also placed ether Solutions of lecithans with cholesterol 
and fat in closed rubber bags suspended in Soxhlet extractors. 
Soxhlet extraction in the usual way failed to remove lecithan from 
the bag under these conditions. These findings favor the develop- 
ment of a method for the thorough removal of impurities from 
lecithan Solutions. 

It is perhaps superfluous to add that the results already mentioned 
may be obtained by placing the Solution to be tested outside the 
rubber bag and allowing dialysis to take place into pure ether con- 
tained in the bag. 

III. SUMMARY OF GENERAL CONCLUSIONS 

I. Most lipins, chief among them, fat, fatty acid, soaps, cho- 
lesterol, cholesterol-esters, lipochrome, and various other ether-sol- 

• Boas and Rosenbloom : Loc. cit. 



I9I2] Jacob Rosenhloom 69 

üble substances, diffuse from ether Solution through rubber 
membranes into ether, 

2. Sodium Chlorid, lecithans prepared from various sources, 
kephalin, cuorin, and the Compound of platinum with lecithin, do not 
diffuse under such conditions. 

3. One or more of the diffusible substances in these experiments 
may be dialyzed from Solutions containing them, together with one 
or more of the indiffusible ones, without inducing any of the latter 
to pass through the membrane. 



STUDIES OF DIFFUSION THROUGH RUBBER 

MEMBRANES 

3. Diffusibility of protein through rubber membranes, with a 

note on the disintegration o£ coUodion membranes 

by common ethyl ether and other solvents 

WILLIAM H. WELKER 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

l. INTRODUCTION 

In the course of our studies of proteins, under the auspices of the 
George Crocker Special Research Fund, we obtained a protein 
product which is soluble in a mixture of equal parts of absolute 
alcohol and absolute ether, and which responds to the biuret, xantho- 
proteic, Millon and Hopkins-Cole tests. The material was prepared 
by the following method : 25 grams of Witte peptone were dissolved 
in 500 c.c. of 0.2 per cent. hydrochloric acid Solution. The liquid 
was evaporated to a thick syrup on a water bath. This syrup was 
thoroughly extracted with absolute alcohol and the resultant yellow 
liquid filtered. The filtrate was treated with an equal volume of 
absolute ether, which produced a white flocculent precipitate. After 
the Sedimentation of the precipitate, the supernatant liquid was de- 
canted and filtered. When five volumes of absolute ether were 
added to this filtrate, a white flocculent precipitate was produced. 
This product was isolated by filtration, washed with absolute ether, 
and exposed to the air in a thin layer on a watch glass, where it 
solidified as yellowish and somewhat hygroscopic granulär material, 
which could easily be pulverized. The product dissolved promptly 
in absolute alcohol. From concentrated alcoholic Solution the prod- 
uct may be precipitated by the addition of an equal volume of abso- 
lute ether. Whether the product is peptone or a much simpler Poly- 
peptid has not yet been determined. 

Dr. Gies and his collaborators have lately given much attention to 

70 



1912] 



William H. Welker 



71 



the diffusibility of lipins and other substances throtigh rubber mem- 
branes. The solubility of the above mentioned product in alcohol- 
ether Solution led Dr. Gies to propose a study of the comparative 
diffusibility of the protein through membranes of rubber, parchment 
and collodion. Such an investigation was accordingly conducted 
by the diffusion process described on page 55. The data in the 
accompanying tables indicate the conditions and the results of the 
tests in this connection.^ 

IL COMPARATIVE DIFFUSION EXPERIMENTS 

Experiments with rubber membranes. The results of the 
tests in the first four series show collectively [Table i (1-15)] that 
biuret-reacting matter appeared in the diffusates; that the rubber 
itself did not yield such substance; and that the occurrence of biuret- 

TABLE I 

Results of experiments with rubber membranes 

A. Data showing the dißusibility of biuret-reacting material 
First Series. With Rubber Condoms. 











Results of 


Results of the 




Duration of 




Liquid outside 


the biuret 


test for leaks at 


Exp. No. 


the experi- 


Contents of the bag 


of the bag 


test in the 


the end of the 




ment, days 






diffusate 


experiment 


I 


4 


r 30 c.c. ether-alc. sol. 
\ 10 c.c. abs. ether 


Abs. ether 


+ + + 


Small hole in 










the bag 


2 


4 


f 20 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ + 


No leak 












3 


4 


f 10 c.c. ether-alc. sol. 
l 30 c.c. abs. ether 


Abs. ether 


+ 


No leak 













Second Series. With Rubber Condoms. 



4 


5 


/ 30 c.c. ether-alc. sol. 
\ 10 c.c. abs. ether 


Abs. ether 


+ + + 


Small hole 










very high up 












in the bag 


5 


5 


f 20 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ + 


No leak 












6 


5 


f 10 c.c. ether-alc. sol. 
\ 30 c.c. abs. ether 


Abs. ether 


+ 


No leak 













* In the tables, " ether-alc. sol." indicates the protein Solution as it was made 
available by the above mentioned process before final precipitation with five 
volumes of ether. Such precipitation was efifected only when the solid was 
desired for special reasons. See the data pertaining to the eighth series of tests, 
Table i. 



72 Studies of Diffusion through Rubber Membranes [Sept. 



TABLE I (continued) 
Third Series. With Rubber Condoms. 



Exp. No. 


Duration of 
the experi- 
ment, days 


Contents of the bag 


Liquid outside 
of the bag 


Results of 
the biuret 
test in the 
difTusate 


Results of the 

test for leaks at 

the end of the 

experiment 


7 


S 


/ 30 c.c. ether-alc. sol. 
1 lo c.c. abs. ether 


Abs. ether 


+ + + 


No leak 












8 


5 


f 20 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ + 


No leak 


9 


5 


f lo c.c. ether-alc. sol. 
\ 30 c.c. abs. ether 


Abs. ether 


+ 


No leak 


lO« 


5 


Abs. ether ("control") 


Abs. ether 


— 


No leak 


II» 


5 


Abs. ether ("control") 


Abs. ether 


— 





FouRTH Series. With Bags of Sheet Rubber.* 



12 


30 


/ 60 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ + + 


No leak 












13 


30 


f 40 c.c. ether-alc. sol. 
1 40 c.c. abs. ether 


Abs. ether 


+ + 


No leak 












14 


30 


/ 20 c.c. ether-alc. sol. 
L 60 c.c. abs. ether 


Abs. ether 


— 


No leak 












IS 


30 


Abs. ether ("control") 


Abs. ether 


— 


No leak 



B. Data showing that the diffusible biuret-reacting material ii-15) 

was true protein 

Fifth Series. With Rubber Condoms. 





Duration of 
the experi- 
ment, days 


Contents of the bag 


Liquid outside 
of the bag 


Results of the tests for 
protein in the diffusate 


Results of 
the test for 


Exp. 
No. 


Biuret 


Hop- 
kins- 
Cole 


Xan- 

thopro- 

teic 


leaks at the 

end of the 

experiment 


16 
17 


3 
3 


/ 30 c.c. ether-alc. sol. 
\ 10 c.c. abs. ether 
30 c.c. ether-alc. sol. 


Abs. ether 
Abs. ether 


+ 
+ 


+ 
+ 


+ 

+ 


No leak 
No leak 



SixTH Series. With Bags of Sheet Rubber.* 



18 


3 


/ 60 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ 


+ 


+ 


No leak 
















19 


3 


r 60 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


+ 


+ 


+ 


No leak 

















Seventh Series. With Rubber Condoms. 



20 

31 



3 
3 



40 c.c. ether-alc. sol. 
40 c.c. ether-alc. sol. 



Abs. ether 
Abs. ether 



+ 
+ 



+ 
+ 



+ 
+ 



No leak 
No leak 



•This control experiment (10) was carried out in duplicate, with negative 
results in each case. 

*In this experiment (11) a new, clean, empty condom, with the bottom 
removed, was suspended in absolute ether, for " control " purposes. 

* " Pure Mexican plantation rubber." 



I9I2] 



William H. Welker 



73 



TABLE I (continued) 
C. Data showing the eßect of water on the diffusion phenomena (.r-21) 
EiGHTH Series. With Rubber Condoms.' 



Exp. No. 


Duration of 
the experi- 
ment, days 


Contents of the bag 


Liquid outside of 
the bag 


Results of 
the biurei 
test in the 
diffusate 


Results of the 

test for leaks at 

the end of the 

experiment 








3 C.C. abs. alc. sol. 


6 c.c. abs. alc. 






22 


5 


' 


9 c.c. abs. ether 
6 c.c. H2O 
3 c.c. abs. alc. 
3 c.c. abs. alc. sol. 


18 c.c. abs. 
ether 
. 1.2 c.c. H2O 
9 c.c. abs. alc. 


•" 


No leak 


23 


5 




21 c.c. abs. ether 
I c.c. H2O 
6 c.c. abs alc. 


21 c.c. abs. 
ether 
I c.c. HsO 


— 


No leak 


24 


5 


\ 


r 3 c.c. abs. alc. sol. 
l 3 c.c. abs. ether 


Equal volumes 
of abs. ether 


+ 


No leak 








and abs. al- 












cohol 







D. Data showing the effect of fat on the diffusion phenomena {1-24) 
NiNTH Series. With Rubber Condoms. 



25 


10 


Olive oil and Witte Pep- 
tone (solid) 


Olive oil 


_e 




26 


10 


Olive oil, Witte peptone 
(solid) and H20^ 


Olive oil 


^" 





Tenth Series. With Rubber Condoms. 



27 
28 



30 
30 



Olive oil and Witte pep- 
tone (solid) 

Olive oil, Witte peptone 
and HjO^ 



Olive oil 
Olive oil 



Eleventh Series. With Rubber Condoms. 



29 


10 


f Ether-alc. sol. 
iLard 


Abs. ether 


_ 














30 


10 


f Ether-alc. sol. 
"iLard 


Abs. ether 


— 















Twelfth Series, With Rubber Condoms. 



31 


1 


/ Ether-alc. sol. 
LLard 


Abs. ether 


+ 


No leak 












32 


I 


/ Ether-alc. sol. 
iLard 


Abs. ether 


+ 


No leak 













Thirteenth Series. With Rubber Condoms. 



33 


2 


f 30 C.C. ether-alc. sol. 
ILard 


30 c.c. abs. ether 


+ 


No leak 












34 


3 


/ 30 C.C. ether-alc. sol. 
ILard 


30 c.c. abs. ether 


+ 


No leak 













'For this series of tests, we used 0.2 gram of the protein material dissolved 
in 10 c.c. of absolute alcohol. 

• On the water-soluble extract of the oil. 

^ Water sufficient to make a paste of the Witte peptone was used. The paste 
was triturated into the oil. 



74 SUidies of Diffusion throngh Rubber Membranes [Sept. 

reacting material in the diffusate was not due to perforations of the 
bags. The diffusion of biuret-reacting material was always greatest 
in degree through the bags containing the largest proportion of 
protein. 

The results of tests 1-15 show that biuret-reacting material dif- 
fiised through the rubber membranes under the conditions imposed. 
In Order to determine more definitely, however, whether protein dif- 
fused through the rubber, we repeated the essential features of tests 
1-15, but applied additional tests to the diffusates [Table i (16- 
21)]. 

The results of tests 16-21 (Table i) confirm the findings of tests 
1-15, and also show definitely that the dififusible biuret-reacting ma- 
terial was triie protein. 

The data of tests 1-2 1 suggest that osmosis depends upon affin- 
ities between the membrane, and the solvent or solute, or both. We 
made a direct test of this mattter in a preliminary way by adding 
water to the solvent and thus disturbing its affinities with the mem- 
brane without decreasing the solubility of the solute. The findings 
are given in the summary pertaining to the eighth series (Table i ). 

The results of tests 22-24 show that water exerted a disturbing 
osmotic influence and that diffusion of the protein was entirely pre- 
vented by the water. We extended these experiments to determina- 
tions of the influence of associated, readily diffusible lipins, in the 
presence or absence of water. The results are given in tests 25-34. 

In the tenth series the oil in the diffusates was emulsified with 
a little soap Solution and then repeatedly extracted with ether until 
all the fat was removed. The water containing the soap, and the 
aqueous extract of the oil, were evaporated to dryness and the 
biuret test applied to a concentrated Solution of the residue. 

Experiments with parchment-paper bags. The foregoing re- 
sults with rubber membranes naturally increased our desire to make 
comparative observations with bags of parchment and collodion. 
The results of the tests with parchment are given in tests 35-42, 
Table 2. That osmosis depends upon accord between the solvent 
and the membrane is obvious from these results also, for the protein 
substance, which is readily diffusible through parchment from 
aqueous Solution, does not dialyze through such a membrane from 
an alcohol-ether Solution. 



I9I2] 



William H. Welker 



n 



TABLE 2 

Results of experiments with bags of parchment paper 

FOURTEENTH SeRIES. 





Duration of the 






Results of the 


Exp. No. 


experiment, 


Contents of the bag 


Liquid outside of 


biuret lest in 




days 




the bag 


the diffusate 


35 


I 


f 10 c.c. ether-alc. sol. 
\ 30 c.c. abs. ether 


Abs. ether 













36 


2 


/ 3 c.c. ether-alc. sol. 
\ 3 c.c. abs. ether 


Abs. ether 


— 










FiFTEENTH SERIES.« 


37 


10 


/ 30 c.c. ether-alc. sol. 
\ 10 c.c. abs. ether 


Abs. ether 


^ 










38 


10 


f 20 c.c. ether-alc. sol. 
1 20 c.c. abs. ether 


Abs. ether 


— 










39 


IG 


f 10 c.c. ether-alc. sol. 
\ 30 c.c. abs. ether 


Abs. ether 


— 










SiXTEENTH SERIES.» 


40 


10 


f 30 c.c. ether-alc. sol. 
\ 10 c.c. abs. ether 


Abs. ether 


_ 










41 


10 


f 20 c.c. ether-alc. sol. 
\ 20 c.c. abs. ether 


Abs. ether 


-" 


42 


10 


/ 10 c.c. ether-alc. sol. 
\ 30 c.c. abs. ether 


Abs. ether 


— 











III. ON THE UTILITY OF COLLODION.BAGS IN EXPERIMENTS OF 
THE KIND DESCRIBED IN THE FOREGOING SECTIONS 

Experiments like those in the sixteenth series (Table 2) were 
attempted with bags made of collodion, but in each case the bags 
were perforated and in part dissolved, by the Contents, before the 
experiment could be fairly started. 

Several years ago, Dr. Gies observed, in some dialysis experi- 
ments with collodion membranes, that ethyl ether could be kept on 
the aqueous Contents of collodion bags, for preservative purposes in 
such tests, without inducing distintegration of the bags. In repeti- 
tions of the experiments a year or two later, however, it was found 
that ether under such circumstances often caused deterioration of 

*The results in this series, while apparently negative in each case, were 
somewhat doubtful owing to the fact that the paper contained some soluble 
material, which rendered the biuret test more or less uncertain. See the results 
of the tests in the sixteenth series. 

*The parchment paper was washed free from soluble matter before the 
beginning of the tests. 



76 Studies of Diffusion through Rubber Membranes [Sept. 

the membrane, but that occasionally it did not. The reason for such 
variations in the action of the ether could not be conveniently ascer- 
tained at the time. 

The prompt Perforation of the collodion bags in our several 
attempts, as stated above, to determine the diffusibihty through col- 
lodion of the alcohol-ether soluble protein, recalled Dr. Gies' pre- 
vious experiences and led him to suspect that the alcohol, in the Solu- 
tions employed by us, was responsible for the observed destructive 
effects on the collodion membrane in these experiments. He be- 
lieved, also, that the previous variations in the action of ether on 
collodion in dialysis experiments, as already related, were due to 
differences in the degrees of purity of the ether employed. At Dr. 
Gies' request, therefore, I made direct tests of the solvent action of 
ether containing alcohol, and various other substances related in one 
way or another to alcohol and ether. 

Collodion bags were made, in test tubes, from U. S. P. col- 
lodion.^*^ It was found that such bags were not perforated by abso- 
lute ether when it was poured into them 10 minutes after their re- 
moval from the tubes, i. e., after fairly complete evaporation of the 
residual alcohol. The time required for the evaporation of the 
residual alcohol is dependent on the prevailing temperature. At low 
temperatures the alcohol disappears from the collodion membrane 
very slowly. Common ether (Merck's 0.720 sp. gr.), however, 
when poured into such bags, passed through them almost immedi- 
ately, with general Solution of the collodion, even after 2 hours of 
preliminary exposure of the bag outside the mould. In the first 
tests of the effects of alcohol it was found that absolute ether con- 
taining 1.5 per Cent, or more of added absolute alcohol promptly 
penetrated the bags. 

In a series of more careful tests of absolute ether containing 
various percentages of added absolute alcohol, it was found that the 
bags were penetrated promptly by ether containing more than 1,25 
per Cent, of alcohol, but that the mixture containing 1.25 per cent. 
of alcohol acted more slowly. Ether containing less than 1.25 ^r 
cent. of alcohol exhibited no destructive action. 

Qualitative tests showed that acetone, acetaldehyde, ethyl acetate, 
methyl alcohol and glacial acetic acid attack and penetrate collodion 

"An ether Solution containing alcohol. 



igi2] William H. Welker 77 

bags^^ immediately, toluol slowly, whereas formic acid, formalde- 
hyde (40 per cent.), Chloroform, petroleum ether, carbon tetra- 
chlorid, carbon bisulfid and paraffin oil were without distinguishable 
solvent action, even after long periods of contact. Acetone (5 per 
cent.) in absolute ether attacks collodion bags slowly, while a 10 per 
cent. Solution acts rapidly. Acetaldehyde (4 per cent.) in absolute 
ether attacks the bags slowly, but a 5 per cent. Solution acts rapidly. 
Methyl alcohol (3 per cent.) in absolute ether dissolves the bags, but 
a 2 per cent. Solution does not. Glacial acetic acid (2 per cent.) in 
absolute ether attacks the bags slowly, a 3 per cent. Solution acts 
rapidly but a i per cent. Solution appears to be inert. Five per 
cent. Solutions of Chloroform, toluol, petroleum ether, carbon tetra- 
chlorid, carbon bisulfid, benzol, ethyl acetate, and paraffin oil, 
in absolute ether, were without visible effect on collodion bags. 
Five per cent. Solutions of formic acid (sp. gr. 1.2) and formalde- 
hyde (40 per cent.), in absolute ether, immediately attacked and 
penetrated collodion bags. 

Further work along these lines is in progress. 

My cordial thanks are due Dr. Gies for his kind direction and 
assistance in these experiments. 

"Bags practically free from residual alcohol were used. 



STUDIES OF DIFFUSION THROUGH RUBBER 

MEMBRANES 

4. The comparative diffusibility of various pigments in 

different solvents 

GEORGE D. BEAL and GEORGE A. GEIGER 

{Biochemical Laboratory of Columbia University, at the College of 
Physicians and Surgeons, New York) 

I. INTRODUCTION 

Dr. Gies and his associates have shown that many biological 
substances diffuse through rubber under suitable conditions (page 
55). Inorganic as well as organic substances exhibit this capacity 
and various colloids share it with crystalloids. Lipochrome and 
ferric sulfocyanate are among the colored substances which, in the 
early experiments, were found to be diffusible from ether Solution 
through rubber membranes into ether. 

At Dr. Gies' Suggestion we undertook a similar study of the 
diffusibility of common pigments, especially " food colors." Fol- 
lowing his advice we also sought data which might be of service in 
devising methods for the purification of pigments, and for their 
Separation and detection under various circumstances. 

Our diffusion tests were conducted by the following general 
method: A moderate quantity of the pigment was mixed with 15- 
25 c.c. of the solvent. The Solution, or Suspension, was carefully 
poured into a rubber Condom in such a way as to preclude the pos- 
sibility of overflow upon the external surface. The bag was then 
immersed in about 50 c.c. of solvent in a narrow salt-mouth bottle 7 
inches high. With the bag suspended at füll extension in this Posi- 
tion, its mouth was about an inch above the opening in the bottle. 
The protruding condom was supported in the neck of the bottle by 
a tightly fitting cork stopper, which served to keep both the bag and 
the bottle closed. The diffusion periods varied from a few minutes 
\o a week or more, according to the obvious requirements in each 
case for a definite conclusion regarding diffusibility. 

78 



I9I2] 



George D. Beul and George A.. Geiger 



79 



Most of the original tests were made with new Condoms. Many 
tests were repeated with Condoms which had previously been em- 
ployed by us in pigment-diffusion experiments but which, prior to 
being used again, had been thoroughly washed with portions of the 
solvent to which they were soon to be subjected in the new diffusion 
tests. Defects in the rubber could easily be detected. All doubtful 
results were ignored. Numerous repetitions prevented erroneous 
deductions. 

In the accompanying summary we present an outline of the 
various tests and the main results of each. For the sake of con- 
venience we use in the summary the following abbreviations : 

D, diffusion; Di, pigment appears in the diffusate within lo minutes; 
D2, pigment does not diffuse within 10 minutes, but appears in the 
diffusate within 30 minutes; D3, pigment does not diffuse within 30 
minutes, but appears in the diffusate within i hour; D4, pigment does 
not diffuse within i hour, but appears in the diffusate before the lapse 
of 2 hours ; D5, pigment cannot be seen in the diffusate before the third 
hour of diffusion, but appears before the fourth hour; D6, pigment 
cannot be seen in the diffusate before the sixth hour of diffusion, but 
appears before the eighth hour; D7, pigment cannot be seen in the 
diffusate before the tenth hour of diffusion, but appears before the 
twelfth hour; D8, pigment appears in diffusate in about 24 hours; 
O, no visible diffusion at any time within a week. 

IL SUMMARY OF DIFFUSION DATA 



Inside 


Outside solvent 


Result 




Solvent 


Pigment 


Remarks 


I Ether. . . 


Sudan III 


Ether 

Ether + alco- 
hol (25%). 

Ether + alco- 
hol (50%). 

Ether -f alco- 
hol (75%). 

Alcohol 

(100%)... 

Chloroform . . 


Di 
Di 
Di 
Di 
Di 

Di 


D very rapid. 


2 Ether. . . 


Sudan III 


3 Ether . . . 

4 Ether. . . 

5 Ether. . . 

6 Ether . . . 


Sudan III 


D rapid though slower than i. 


Sudan III 


D slower than 2. 


Sudan III 


D slower than 3. 


Sudan III 


Ether was withdrawn, leaving a 
concentrated Solution of 
Sudan III. Currents very 
distinct. CoUection of color 
near top very marked. 

Color zone on top. No diffusion 
currents downward. 







8o Studios of Diffusion through Rubber Membranes [Sept. 
II. SUMMARY OF DIFFUSION DATA {continued) 



Inside 



Solvent 



7 Ether. 



8 Ether . . . 

9 Ether. . . 

10 Ether . . . 

11 Alcohol. . 

12 Chloro- 

form . . . 

13 Alcohol . . 

14 Ethyl 
acetate . . 

15 Acetone.. 

16 Gl. acetic 

acid .... 

17 Ether. . . 

18 Ether. . . 

19 Ethyl 
acetate . . 

30 Acetone.. 
21 Alcohol. . 
27 Ether. . . 

23 Ether. . . 

24 Ether . . . 

25 Ether. 



Pigment 



26 Ether . . 

27 Ether. . 

28 Ether. . 

29 Ether. . 

30 Ether . . 

31 Ether. . 

32 Ether. . 

33 Ether. . 

34 Ether . . 

35 Ether. . 

36 Ethyl 
acetate . 

37 Ethyl 
acetate . 



Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Sudan III 

Picric acid 

Hematoxylin 

Methyl violet 

Methyl violet 

Methyl violet 

Methyl violet 

Magenta 

Naphthol yellow . . . 

Methyl violet and 

Sudan III 



Outside solvent 



Methyl alco- 
hol 



Acetone . 



Petroleum 

ether. . . . 
Gl. acetic acid 
Ether 



Chloroform . 
Alcohol . . . . 



Ethyl acetate 
Acetone .... 



Gl. acetic acid 
Ether 



Ether 

Ethyl acetate 



Acetone . 
Alcohol . 
Ether. . . 
Ether. . . 
Ether... 



Resnlt 



Ether. 



Chlorophyll Ether. 



Annatto 

Alcannin 

Metanil yellow. . 
Martius yellow. . 

Scarlet R 

Malachite green. 
Brazil wood . . . . 

Chrysoldin 

Turmeric 



'Ether. 
Ether. 
Ether. 
Ether. 
Ether. 
Ether. 
Ether. 
Ether. 
Ether. 



Annatto . 



Chlorophyl . 



Ethyl acetate 



Ethyl acetate 



Di 



Di 



Di 
Di 
Di 

Di 
Di 

Di 
Di 

Di 
Di 

D3 

DS 

D6 

O 

D5 

Ds 

O 

Di 



Di 
Di 
Di 
O 
Di 
Di 

D2 

Di 

D2 

Di 

D2 



Di 



Remarks 



Ether was withdrawn, leaving 

layer of dye inside. 
Ether withdrawn. Solution of 

pigment in bag concentrated. 



Moderate diffusion. 
Diffusion slow. 

Rapid diffusion. 
Diffusion slow. 

Slow diffusion. 
Moderate diffusion. 

Slow diffusion. 

Rubber yellow; color not re- 
moved by ether. 



Denser Solution in bag than 
outside. (See 36.) 



Rapid diffusion of the Sudan 
III. Ether changed three 
times in 2 hours after which 
practically all Sudan III had 
been removed, leaving the 
methyl violet in the bag. 

Vary slight diffusion. 

Very slight diffusion. 

Very rapid diffusion. 

Rapid diffusion. 
Very rapid diffusion. 
Bag colored green. 
Slow diffusion. 
Very slow diffusion. 
Rapid diffusion. 

Pigment Solution inside concen- 
trated.« 

Pigment Solution inside concen- 
trated. 



• " Fat-soluble " chlorophyl was used in all the chlorophyl tests. 

* In these cases (36-44) the solvent diffused more rapidly than the solute. 



1912] 



II. 



George D. Beal and George A. Geiger 

SUMMARY OF DIFFUSION DATA (continued) 



8i 





Inside 












Outside solvent 


Result 


T< ATVl f l^e 


Solvent 


Pigment 


JxCularKa 


38 Ethyl 










acetate . . 


Alcannin 


Ethyl acetate 


Di 


Pigment Solution inside concen- 










trated. 


39 Ethyl 










acetate . . 


Martius yellow .... 


Ethyl acetate 


Di 


Pigment Solution inside concen- 
trated. 


40 Ethyl 










acetate . . 


Scarlet R 


Ethyl acetate 


Di 


Pigment Solution inside concen- 
trated. 










41 Ethyl 










acetate . . 


Brazil wood 


Ethyl acetate 


Di 


Pigment Solution inside concen- 
trated. 


42 Ethyl 










acetate . . 


Chrysoidin 


Ethyl acetate 


D3 


Pigment Solution inside concen- 
trated. 


43 Ethyl 










acetate . . 


Turmeric 


Ethyl acetate 


D3 


Pigment Solution inside concen- 








^-'O 


trated. 


44 Ethyl 










acetate. . 


Metanil yellow 


Ethyl acetate 


D3 


Pigment Solution inside concen- 
trated. 


45 Ethyl 










acetate . . 


Malachite green. . . . 


Ethyl acetate 


D6 




46 Ethyl 










acetate . . 


Naphthol yellow . . . 


Ethyl acetate 







47 Ethyl 










acetate . . 


Sudan I 


Ethyl acetate 


Di 


Rapid diffusion. 


48 Ethyl 










acetate . . 


Sudan G 


Ethyl acetate 


Di 


Moderate diffusion. 


49 Ethyl 






acetate . . 


Rhodamin 


Ethyl acetate 




D in about 2 days. 


50 Ethyl 






acetate . . 


Fast red A 


Ethyl acetate 


D3 


Diffusion very slight in each of 


51 Ethyl 








tests 50-56 inclusive. 


acetate . . 


Rose bengal 


Ethyl acetate 


D2 




52 Ethyl 










acetate. . 


Erythrosin 


Ethyl acetate 


D2 




53 Ethyl 










acetate . . 


Methylene violet . . . 


Ethyl acetate 


D5 




54 Ethyl 










acetate. . 


Phloxin red 


Ethyl acetate 


D3 




55 Ethyl 










acetate . . 


Auramine 


Ethyl acetate 


D5 




56 Ethyl 






acetate . . 


Orange G 


Ethyl acetate 




D in about 5 hours. 


57 Methyl 










alcohol. . 


Gold orange 


Methyl 










alcohol .... 


D8 


Color of diffusate very slight 
i week later. 


58 Methyl 










alcohol. . 


Naphthol yellow . . . 


Methyl 
alcohol. . . . 




No appearance of color in 8 hours. 


59 Meth 










alcohol. . 


Carmosin B 


Methyl 










alcohol .... 


D8 


Color of diffusate very slight 
i week later. 



82 Studics of Diffusion through Ruhher Memhranes [Sept 
II. SUMMARY OF DIFFUSION DATA (continued) 



Inside 



Solvent 



60 Methyl 
alcohol . 

61 Methyl 
alcohol . 

62 Methyl 
alcohol . 

63 Methyl 
alcohol . 

64 Amyl 
alcohol . 

65 Amyl 
alcohol . 

66 Amyl 
alcohol . 

67 Amyl 
alcohol . 



68 Amyl 
alcohol . . 

69 Amyl 
alcohol. . 

70 Amyl 
alcohol. . 

71 Amyl 
alcohol. . 

72 Amyl 
alcohol . , 

73 Amyl 
alcohol. , 

74 Amyl 
alcohol . . 

75 Amyl 
alcohol. , 

76 Amyl 
alcohol . , 

77 Acetone. 

78 Acetone. 

79 Acetone. 

80 Acetone. 

81 Acetone. 

82 Acetone. 

83 Acetone. 

84 Acetone. 

85 Acetone. 

86 Acetone. 

87 Acetone. 

88 Acetone. 

89 Acetone. 



Pigment 



Ponceau, G. A.. 
Ponceau, 2 R. . 
Naphthol red S. 
Curcumin S . . . 



Fast red A . 
Safranin . . . 



Eosin A. 
Phloxin . 



Rose bengal . 
Rhodamin . . 
Erythrosin . . 



Chrysoldin 
Sudan I . . . 
Sudan G. . . 
Sudan III. 
Alcannin. . 



Chlorophyl . 
Alcannin . . , 
Auramine. . 
Barwood . . , 
Chlorophyl . 
Chrysoldin . 
Fast red A . 



Methylene violet. . 
Malachite green. . . 
Martins yellow. . . . 
Metanil yellow. . . . 
Naphthol yellow S. 

Picric acid 

Rhodamin 



Outside solvent 



Result 



Methyl 
alcohol. . . . 

Methyl 
alcohol. . . . 

Methyl 
alcohol. . . . 

Methyl 
alcohol. . . . 

Amyl alcohol 

Amyl alcohol 

Amyl alcohol 
Amyl alcohol 



Amyl alcohol 
Amyl alcohol 
Amyl alcohol 

Amyl alcohol 

Amyl alcohol 

Amyl alcohol 

Amyl alcohol 

Amyl alcohol 

Amyl alcohol 
Acetone. . . 
Acetone. . . 
Acetone . . . 
Acetone . . . 
Acetone . . . 
Acetone. . . 

Acetone . . . 
Acetone. . . 
Acetone . . . 
Acetone . . . 
Acetone. . . 
Acetone. . . 
Acetone. . . 



O 

O 

O 

O 
D7 
D7 

O 



O 

D7 

D8 

D3 

D2 
D2 
D2 

D3 

D3 
Di 
Di 
Di 
D3 
D5 
D4 

D4 
D5 
D3 
D8 
O 
Dl 
D7 



Remaxks 



Color of diffusate very slight 
I week later. 



D in about 3 days. Color of 
diffusate very shght i week 
later. 



Color of diffusate very slight 
I week later. 



Very rapid diffusion. 



Color of diffusate not very 
strong I week later. 



I9I2] 



George D. Beal and George A. Geiger 



83 



II 


SUMMARY OF DIFFUSION DATA {continued) 


Inside 












Outside solvent 


^Result 


Remarks 


Solvent 


Pigment 








90 Acetone.. 


Sudan I 


Acetone 

Acetone 

Acetone 

Acetone 


Dl 

Di 



D8 




91 Acetone.. 


Sudan G 




92 Acetone.. 


Fustic 




93 Acetone.. 


Rose bengal 


Color of dif?usate very slight 










i week later. 


94 Acetone . . 


Phloxin 


Acetone . 


D4 


Color of diffusate very slight 
I week later. 






A ^^^ V.. W^^ 4 Jk^h^ • • ■ • a 


95 Acetone.. 


Eosin W. gelblich . . . 


Acetone 


D7 


Color of diffusate very slight 
I week later. 


96 Acetone.. 

97 Acetone.. 


Eosin A 


Acetone 

Acetone 



D4 




Cape aloes 




98 Gl. acetic 










acid .... 


Sudan G 


Gl. acetic acid 


Di 


Slow diffusion. 


99 Gl. acetic 




fc..^**.^ TT *.A**Ä 1.4V^ A^..r AA # 


acid .... 


Sudan III 


Gl. acetic acid 


Dl 


Rapid diffusion. 


100 Gl. acetic 








acid. . . . 


Sudan I 


Gl. acetic acid 


Di 


Slow diffusion. 


loi Gl. acetic 






acid. . . . 


Alcannin 


Gl. acetic acid 


D3 




102 Gl. acetic 






acid .... 


Chlorophyl 


Gl. acetic acid 


D6 




103 Gl. acetic 










acid .... 


Rose bengal 


Gl. acetic acid 


DB 




104 Gl. acetic 










acid .... 


Phloxin 


Gl. acetic acid 




D in about 2 days. 


105 Gl. acetic 










acid. . . . 


Malachite green. . . . 


Gl. acetic acid 







106 Gl. acetic 










acid .... 


Methyl violet 


Gl. acetic acid 


D8 




107 Gl. acetic 










acid. . . . 


Scarlet R 


Gl. acetic acid 


D2 




108 Gl. acetic 






acid .... 


Methylene violet . . . 


Gl. acetic acid 




D in about 10 days. 


109 Gl. acetic 










acid .... 


Martius yellow 


Gl. acetic acid 


D8 




iio Gl. acetic 










acid .... 


Biebrich Scarlet. . . . 


Gl. acetic acid 


D8 




III Gl. acetic 










acid. . . . 


Erythrosin 


Gl. acetic acid 




D in about 4 days. 


112 Gl. acetic 










acid .... 


Oranee G 


Gl. acetic acid 







113 Gl. acetic 


^.—1' ^ v***^^ ^- ^-* ••••»• •«• ■ 


^t^H« * ■ fc*^-v* W^V^ ^.l>VrA^.J 






acid. . . . 


Tropeolin OO 


Gl. acetic acid 




Very slight color after 2 days, 
which did not increase after 
Standing about four days. 


114 Gl. acetic 










acid. . . . 


Aurainine 


Gl. acetic acid 







115 Gl. acetic 






acid. . 


Rhodamin 


Gl. acetic acid 


D8 




116 Gl. acetic 


^^.AA V.' X.A%4* AA * A^ ■«•■*«*«■ 




acid .... 


Eosin A. gelblich . . . 


Gl. acetic acid 


D8 




117 Gl. acetic 










acid .... 


Chrysoidin 


Gl. acetic acid 




D in about s days. 


118 Gl. acetic 










acid .... 


Eosin W 


Gl. acetic acid 




D in about 4 days, which did 
not increase during the suc- 




M rf\J\^i**-* »■• •••••■••• 










ceeding 3 days. 



84 Stndics of Diffusion through Rubber Membranes [Sept. 
II. SUMMARY OF DIFFUSION DATA (continued) 



Inside 



Solvent 



119 Gl. acetic 

acid. . . . 

120 Gl. acetic 

acid .... 



acetic 



acetic 



acetic 



121 Gl. acetic 

acid 

122 Gl 

acid . . 

123 Gl 

acid. 

124 Gl 

acid. . . . 

125 Gl. acetic 

acid .... 

126 Alcohol. . 

127 Alcohol. . 

128 Alcohol. . 

129 Alcohol. . 

130 Alcohol. . 

131 Alcohol. . 

132 Alcohol. . 

133 Alcohol. . 

134 Alcohol. . 

135 Alcohol. . 

136 Alcohol. . 

137 Alcohol. . 

138 Alcohol. . 

139 Alcohol. . 

140 Alcohol. . 

141 Alcohol. . 

142 Alcohol. . 

143 Alcohol. . 

144 Alcohol. . 

145 Alcohol. . 

146 Alcohol. . 

147 Alcohol. . 

148 Alcohol.. 

149 Alcohol. . 

150 Alcohol. . 

151 Alcohol. . 

152 Alcohol. . 

153 Alcohol. . 

154 Alcohol. . 

15s Alcohol. . 



Pigment 



Fast Red A 

Safranin 

Turmeric 

Metanil yellow 

Barwood 

Annatto 

Picric acid 

Auramine 

ChrysoTdin. .._..... 

Eosin A 

Eosin W ' 

Fast Red 

Methyl violet 

Methylene violet. . . 

Malachite green. . . . 

Martins yellow 

Metanil yellow 

Rose bengal 

Rhodamin 

Sudan G 

Sudan I 

Bismarck brown . . . 

Benzopurpurin 

Tropeolin OO 

Phloxin 

Safranin 

Naphthol yellow . . . 

Alcannin 

Chlorophyl 

Barwood 

Fustic 

Turmeric 

Cape aloes 

Curcumin S 

Naphthol green. . . . 
Orange G 

Carmosin B 



Outside solvent 



Gl. acetic acid 
Gl. acetic acid 

Gl. acetic acid 

Gl. acetic acid 

Gl. acetic acid 

Gl. acetic acid 

Gl. acetic acid 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 

Alcohol 



Result 



D3 



O 



D7 



O 



D7 
D7 
O 
D7 
D8 

D7 

DB 
D7 
D7 

O 
O 
D2 

D2 

O 

O 

D7 



O 
DS 
D7 

O 
O 
O 

O 
O 



Remarks 



D in about 4 days, which did 
not increase during the suc- 
ceeding 3 days. 



D in about 2 days. 



D in about 6 days. 
D in about 3 days. 



Color of diffusate very slight 

I week later. 
Color of diffusate very slight 

I week later. 



Color of diffusate very slight 
I week later. 



Color of diffusate very slight 

I week later. 
D in about 3 days. 
D in about 5 days. Slight 

I week later. 



Color of diffusate very slight 
I week later. 



D in about 2 days. 



D in about 2 days. 

I week later. 
D in about 2 days. 

I week later. 



Very slight 
Very slight 



I9I2] George D. Beal and George A. Geiger 85 

III. ATTEMPTS TO SEPARATE PIGMENTS BY DIALYSIS 

The outcome of Test 25 encouraged us to ascertain whether 
two dissimilar pigments like scarlet R and malachite green might be 
wholly separated from each other by dialysis thru rubber in a suit- 
able solvent, e. g., ethyl acetate (see tests 40 and 45). A mixture 
of the two pigments dissolved in ethyl acetate was accordingly sub- 
jected to the usual mechanical treatment, but the diffusate was re- 
peatedly replaced with fresh solvent. The results are indicated in 
the f ollowing summary : 
Continuous differential diffusion of scarlet R and malachite green. 

March 28 — ist diffusate ii-i p.m. Bright red. 

March 28 — 2nd diffusate 4 p.m. Bright red. 

March 28 — 3rd diffusate 11 p.m. Deep red. 

March 29 — 4ith diffusate 12.30 a.m. Deep purplish red. 

March 29 — 5th diffusate i a.m. Light purplish red. 

March 29 — 6th diffusate 10.45 ^■^- Deep red with decidedly 
bluish tinge. 

March 29 — 7th diffusate 12.30 p.m. Light blue. 

March 29 — 8th diffusate 9.15 p.m. Blue green. 

March 30 — 9th diffusate 11.50 a.m. Blue green. 

March 31 — loth diffusate 9.15 a.m. Blue green. 

April I — iith diffusate i p.m. Blue green. 

April 2 — I2th diffusate 11.50 p.m. Light green. 

April 3 — I3th diffusate 11.50 p.m. Light green. 

Altho scarlet R and malachite green showed widely different 
rates of diffusion when they were treated separately, the results 
detailed above made it evident that it would be difficult if not im- 
possible to obtain all the scarlet R from mixtures like the one em- 
ployed without removing some of the malachite green with the red 
pigment. 

By subjecting Solutions of scarlet R and malachite green of simi- 
lar concentrations independently to diffusion in the usual way, we 
duplicated the blue and green effects with malachite green and the 
red effects with scarlet R, but the purplish colorations could not he 
ohtained under such circumstances. That these purplish effects 
were due to early diffusion of the malachite green with scarlet 
R, and that the red pigment facilitated the passage of the green one, 
are clearly indicated by the results. 



86 Studies of Diffusion through Rubber Membranes [Sept 

Repeated removals of the diffusate in the independent scarlet R 
experiment, and replacements with fresh solvent for a period of 
about a weck, led to the Separation from the original pigment- 
product of all its red diffusible matter. The bag, at that stage of 
the treatment, contained considerable brownish-red, indifTusible 
material, which evidently was not scarlet R. This result, and simi- 
lar observations with other pigments, emphasized Dr. Gies' opinion 
that it might be possible to purify pigment preparations in this way 
and that their value as coloration agents, for histological staining 
especially, might thus be considerably enhanced. 

It will be noted that those pigments which diffused most rapidly 
were the so-called " fat colors," i. e., those soluble in, or staining, 
the common fats and oils. Again, with these pigments the dif- 
fusion is the most rapid, and therefore the most satisfactory, when 
the solvents are those which, in the State of vapor, soften rubber. 
It will thus be seen that apparently the membrane, as well as the 
solvent, exert selective action. This is true to a far greater extent 
in experiments of this kind than in the ordinary dialyses in aqueous 
media. 

When we arrived at this point in these experiments, to which 
we could give but a few hours weekly, our period of residence at 
Columbia University was about to close and, after completing some 
repetitions of previous observations in this connection, we were 
obliged to discontinue the work. It is Dr. Gies' intention to pro- 
ceed along lines suggested by the results already obtained in this 
preliminary investigation. 



THE COLLOIDAL NITROGEN IN THE URINE FROM 
A DOG WITH A TUMOR OF THE BREAST 

MAX KAHN AND JACOB ROSENBLOOM 

(Biochetnical Laboratory of Columbia University, at the College of 
Physicians and Surgeons, New York) 

In 1892 Töpfer^ found that the urine of patients suffering from 
Cancer contained a very large amount of "extractive substance." 
This " extractive substance " was calculated by first determining the 
total nitrogen, and then subtracting from this amount the sum of the 
nitrogen values for the urea, uric acid, and ammonia contained in 
the same urine. Bondzynski and Gottlieb,^ five years later, reported 
that the nitrogen in oxyproteic acid was 2 to 3 per cent, of the total 
urinary nitrogen. Salkowski,^ and Hess and Saxl,* using different 
procedures in their researches, came to the conclusion that the oxy- 
proteic acid or the alcohol-precipitable substances are increased in 
the urine of human beings suffering from Carcinoma. 

Salkowski and Kojo,^ in a preliminary communication, recently 
suggested several methods for the determination of colloidal nitro- 
gen in the urine. A year later Kojo published the results of a com- 
parative study of the various procedures suggested in this connec- 
tion.^ Einhorn, Kahn and Rosenbloom^ studied the zinc sulfate- 
precipitable, colloidal, nitrogenous material from the urine of nor- 
mal subjects as well as from the urine of carcinomatous patients, 
and came to the conclusion that the amount of colloidal nitrogen 
was invariably increased in subjects with carcinomatous growths. 

* Töpfer: Wiener klin. Wochenschrift, 1892, v, p. 49. 

'Bondzynski and Gottlieb: Zentralbl. f. d. med. Wissenschaften, 1897, xxxv, 

P- 577- 

* Salkowski : Berliner klin. Woch., 1910, xlvii, p. 1746. 

*Hess and Saxl: Beiträge zur Carcinomforschung, 1910, Part II. 
'Salkowski and Kojo: Berl. klin. Woch., 1910, xlvii, p. 2297. 
'Kojo: Zeitschr. f. physiol. Chem., 191 1, Ixxiii, p. 416. 

^Einhorn, Kahn and Rosenbloom: Amer. Journ. of Gastro-enterology, 1911, 
i, p. 2; and Archiv f. Verdauungs-Krankheiten, 191 1, xvii, p. 557. 

87 



88 Colloidal Nitrogen in Urinc from a Dog [Sept. 

The writers lately embraced an opportunity to study the colloidal 
nitrogen Output in the urine of a dog with a large tumor. 

The dog upon which this study was made had a hard calcified 
growth about the size of an orange in one of the breasts. The 
tumor involved the nipple and the breast tissue for some distance 
around the nipple. Several metastatic deposits were present along 
the "breast lines." Microscopic examination of sections of the 
original growth and of the metastatic infiltrations, according to 
several pathologists who examined them, indicated that the tumor 
was a chondroma which had undergone carcinomatous degenera- 
tion. Other pathologists, on the contrary, believed the growth to 
be of a benign nature, with the histological structure of a chon- 
droma. 

For the determination of colloidal nitrogen the alcoholic pre- 
cipitation method of Salkowski was used, with modifications, as 
follows :^ 

The total nitrogen was determlned in 5 c.c. of the urine by the 
Kjeldahl process. Two portions of 100 c.c. each of the urine were 
evaporated in a porcelain dish over a gently steaming water bath tili 
they were of the consistency of thin syrup. The residues were then 
taken up in 100 c.c. of alcohol (98.5 per cent.) and thoroughly 
stirred. The alcoholic extracts were then filtered through ashless 
filter papers, and the precipitates washed with alcohol. 

We determined the effect of dialysis upon this alcohol-precip- 
itable, so-called "colloidal," nitrogenous material. Most colloidal 
substances fail to dialyze through the very best grade of parchment 
paper. Only that fraction of the alcoholic precipitate which would 
remain indiffusible under suitable conditions of dialysis could be 
called "colloidal," at the present stage of our knowledge of the 
subject. Accordingly, the two precipitates on the ashless filter 
paper were treated as follows : 

The precipitate on one filter paper, together with the filter, was 
placed in a Kjeldahl flask, digested with sulfuric acid, and the 
nitrogen determined in the usual way. The second precipitate and 

*Before subjection to analysis the urine was first tested for protein, which, 
if found, was removed by means of heat coagulation aided by the addition of a 
few drops of dilute acetic acid Solution. 



I9I2] 



Max Kahn and Jacob Rosenbloom 



89 



filter paper were placed with water in a bag of the finest grade of 
parchment paper and dialyzed for forty-eight hours. The liquid in 
the bag was then analyzed quantitatively for nitrogen. 

The appended summaries present the results obtained for urine 
from the dog with the breast tumor and also for urine f rom several 
normal dogs. 

In the Salkowski method for the determination of " colloidal " 
nitrogen (as the results in the summary show), diffusible nitrog- 
enous substance is precipitated as "colloidal" nitrogen. It has 
not yet been shown that such diffusible nitrogenous matter in the 
colloidal precipitate is true colloidal material. 





A. Data pertaining 


to the urine of normal dogs 


ßpecimen 
No. 


Total Nitrogen 

in 100 c.c. of 

Urine 


Colloidal Nitro- 
gen in 100 o.e. 
of Urine 


Percentage of 

Total Nitrogen 

as Colloidal 

Nitrogen 


Indiffusible 
Colloidal 
Nitrogen 


Percentage of Total 
Nitrogen as Indif- 
fusible Colloidal 
Nitrogen 


I 

2 

3 

4 


Grams 
2.304s 
3-2051 
0.8590 
1.6436 


Gram 

0.0437 
0.0314 
0.0172 
0.0214 


1.85 
0.98 
2.00 
1.28 


Gram 
0.02775 
0.01634 
0.01202 
0.01841 


1.2 

O.S 
1.4 
i.l 



B. Date pertaining to the urine of the dog with a tumor of the breast 



Sa. 


4.0088 


0-3392 


8.40 


0.0939 


2.3 


5 b. 


6.3034 


0-3897 


6.10 


0.2293 


3.6 


SC. 


4-4591 


0.3210 


7.10 


0.0767 


1-7 


5d. 


3.6862 


0.3294 


8.10 


0.0817 


2.2 


Se. 


3-1414 


0.0958 


3-04 


0.0867 


2.7 


5 f. 


3.9642 


0.3566 


8.90 


0.1175 


2.8 


5 g- 


2.5139 


0.4617 


13.10 


0.1342 


3.6 



The results demonstrate that the " colloidal " nitrogen, both 
before and after dialysis, was greater in amount in the urine of the 
dog with the tumor than that in the urines from normal dogs. 

It is desirable to study the effect of dialysis upon the " colloidal " 
nitrogenous substances in the urine of Cancer patients. 



GENERAL ASPECTS OF FASTING^ 

PAUL E. HOWE 
(Department of Physiological Chemistry, University of Illinois, Urbana, III.) 

Fasting (starvation or inanition) is a State in which the dietary 
elements are withheld, either wholly or in part, so that the organism 
is compelled to draw upon its own resources to maintain its exist- 
ence. In discussing this subject it is my purpose to make a rather 
general survey of the changes which take place as the result of fast- 
ing; to show briefly how such results have been used to elucidate 
other scientific problems; and, also, to touch upon the therapeutic 
value of fasting, with relation to man. 

A distinction is made between physiological and experimental 
fasting. The first form is illustrated by the hibernation of mam- 
malia (hedge hog and bear), and cold blooded animals (frog), by 
the normal condition of the salmon during the spawning season and 
by the period of metamorphosis of the insects, these being natural 
phenomena for which the organism has made suitable preparation. 
In experimental fasting the animal is forced to live without sus- 
tenance, of one kind or another. Under this last State we may con- 
sider pathological fasting as a special case in which the organism is 
forced to fast as a result of impairment of some organ or of a gen- 
eral diseased condition. These forms of inanition present certain 
differences as evidenced in the effect upon the organism; yet it is 
quite probable that they are chiefly phylogenetic and we can conceive 
that any of the animals which do not experience these periodical 
physiological fasts might do so under the proper adverse circum- 
stances. 

In our discussion we will consider only the phenomena which 
take place as the result of experimental fasting. Here, too, we must 
distinguish between a number of forms of fast; such as the com- 

^ A lecture delivered at the College of Physicians and Surgeons, New York, 
May I, 1912, under the auspices of the Columbia University Biochemical Asso- 
ciation. 

90 



I9I2] Paul E. Howe 91 

plete fast in which there is total abstinence from both food and 
water; a modification of this, in which the subject is permitted to 
take water "ad libitum" or caused to ingest a uniform quantity 
from day to day ; and the incomplete fast, in which one or more of 
the food principles or chemical elements contained therein is with- 
held, such as a diet lacking in protein, fat, carbohydrate, water, salt 
or certain amino acids. 

There is not a marked distinction between complete fasting 
and fasting with water taken "ad libitum," for under the latter con- 
ditions the quantity of water taken decreases as the fast progresses 
until finally there is a natural abstinence from water. Some hold 
that the desire for water returns just before death. The ingestion 
of water causes a lengthening of the life of the animal and the 
severity of the fast is lessened. If at any time the quantity of water 
given is increased there will be for a time an increase in the metab- 
olism (14). This condition also holds for the well nourished 
animal (8), i. e., under all conditions when the water ingestion of 
the animal is sufficiently increased the general metabolic processes 
of the organism are stimulated. 

The length of a fast which would result in death depends upon 
the size, the species, the age, the nutritive condition, the external 
surroundings (e. g., temperature, humidity, etc.) and the intrinsic 
rate of metabolism. In general, we may say that the smaller the 
subject the shorter will be the time it can live without food; but this 
does not hold in all cases, for certain of the lower animals can fast 
much longer than the higher forms, e. g., the Salamander, which is 
about 3-4 inches long, has been fasted for more than 125 days (19). 
Adult organisms can fast longer than the young of the same species. 
Thus, a young pup can fast but a few days, while a füll grown dog 
will fast from 20 to 60 days. Of the fasts on man and other warm 
blooded mammals, the longest on record is one of 117 days (15). 
This experiment was conducted in our laboratory, a Scotch collie 
dog being the subject. Subsequent to this long fasting interval the 
dog was fed, and it returned to its normal condition. 

A comparison of the results obtained by various investigators 
shows that death does not ensue until there is a loss of between 40- 
50 per Cent, of the original body weight. The real cause of death 



92 General Aspects of Fasting [Sept. 

from fasting has not been determined. The probable reason is the 
failure of some organ or life process (27) and not the depletion of 
all possible nutritive material. From our experiments (10) it 
would appear that a certain definite minimal proportion of nitrogen- 
holding substance must be present in the body for life to exist. 

Fasts have been reported upon men covering periods of from 
2-50 days, upon dogs as long as 117 days and Salamanders for 125 
days. In each of the extreme cases, the subjects were subsequently 
fed and they returned to normal. The influenae of repeated fasting 
upon the resistance of the animal to subsequent fasts is a phenome- 
non which appears to be intimately associated with hibernation. As 
has been shown by Russian investigators (20), and more recently in 
our laboratory (10), repeated fasting decreases the rate of metab- 
olism in each succeeding fast. A French investigator (21) has 
shown that repeated fasting, in which the subjects were alternately 
fasted and fed during equal periods of about a week each, resulted 
in the ultimate death of the animals. From the experimental data 
Et band it seems that where the animal is permitted to recover 
completely from a fast before it is subjected to another, there will 
be an increased resistance to the ravages of the succeeding fast. 

The number of men who have made a study of the changes 
which take place as the result of fasting is so great that it is difficult 
to name those who have made the most important contributions 
upon this subject without doing an injustice to others. The inves- 
tigations of Cathcart (4) in England and of Benedict (2) in this 
country, upon men, are the most extensive that have been con- 
ducted with the more refined methods of analysis which we possess 
today. The names of Succi, Cetti and Breithaupt stand out in the 
literature as the subjects of important experimental fasts. 

What changes take place in an organism as the result of a fast? 
Outwardly the subject becomes emaciated, his body weight de- 
creases, he becomes weak and apathetic and, should the fast proceed 
long enough, he would probably die in a State of coma. In man it 
has been demonstrated that the brain retains its activities unimpaired 
during a fast and that hunger is evident only at the beginning of the 
ordeal. These facts are substantiated in the populär writings upon 
fasting and also by an experiment made by us (11) in which the 



I9I2] Paul E. Howe 93 

subject prepared for the preliminary examination for the degree of 
Doctor of Philosophy during a seven-day fast. 

The fasting State is indicated in the body by certain changes; 
such as a general decrease in the body metaboHsm, represented by 
variations in the nitrogen excretion and the respiratory exchange, a 
decrease in the fat and glycogen Stores, a decrease in the volume of 
muscle and in the size and weight of certain organs. The tempera- 
ture remains normal, for a time at least, but shows a tendency to 
decrease toward the end of the fast. 

The decrease in the general metaboHsm is well Illustrated by the 
data obtained from the respiration calorimeter experiments. It has 
been shown by the earlier investigators and more recently by Bene- 
dict (2) that, in a well fed man, the quantities of protein and fat 
which were utiHzed, and the energy change (calories) per day, 
decreased very gradually and tended toward a constant minimum. 
In addition to the excreted carbon dioxide, Benedict determined the 
amount of oxygen consumed. From these data it was shown that 
the glycogen consumption, which is most rapid on the first day, 
decreases as the fast progresses. It is probable that the glycogen 
Store is never depleted and that even in fasting there is a resynthesis 
of glycogen from the protein material present in the body. 

Decreased metaboHsm in fasting is also shown by the quantities 
of nitrogen-containing substances eliminated in the excreta. We are 
particularly concerned with the losses of nitrogen, for it is the 
protein material which is the most fundamental nutritive substance 
and which the body strives to protect. In fasting, the nitrogen- 
containing substances in the urine or feces arise from the tissues and 
hence the total nitrogen excretion is a measure of the quantity of 
muscular or organ tissue catabolized. The excretion of total nitro- 
gen in the urine decreases rapidly at the beginning of the fast and 
soon reaches a minimum, which is maintained for some time. This 
minimum of nitrogen excretion, representing a minimum protein 
disintegration, is Held to represent the " maintenance " metaboHsm 
of the individual, i. e., that amount of protein substance which if 
supplied, with sufficient fats or carbohydrates, in the form of food 
would sustain life. This minimum has variously been shown to be 
greater or less than the metaboHsm as represented by fasting ex- 
periments. 



94 General Aspects of Fasting [Sept 

The muscular disintegration is influenced by the factors already 
mentioned; the nutritive condition and the experience of previous 
fasts, or repeated fasting. The diet just before the fast influences 
the nitrogen excretion for a number of days. This has been demon- 
strated in the classical experiments of Voit (26), in which he fed 
varying amounts of meat and bread to a dog and showed that, when 
fasted the rate of nitrogen excretion varied, but that in each case 
the animal came to the same level of catabolism on about the seventh 
day of fasting. 

The fat available in the body exerts a marked efTect upon the 
protein metabolism and the Hfe of an animal. So long as there is 
sufficient fat in the organism to supply the energy requirements, 
the protein metabolism will remain at a minimum. When, how- 
ever, the fat deposits are depleted, the body is forced to use protein 
to furnish the necessary energy. The result is a more rapid protein 
consumption and an earlier death. This increased protein con- 
sumption, is, of course, accompanied by an increased nitrogen excre- 
tion, which has been designated as the " premortal rise." The feed- 
ing of carbohydrate or fat sufficient to supply the energy require- 
ment of the body would prevent this increased consumption of pro- 
tein and thus lengthen the life. 

Repeated fasting will also modify the rate of metabolism. This 
point is well illustrated by the results obtained on a subject in a 
repeated fast (10), in which there was a rapid and increasing con- 
sumption of the protein reserves of the body during the first fast, 
but a more gradual and uniform consumption during the second 
fast. The total body weight and nitrogen losses were practically 
identical in the two fasts and the data f rom the intermediate feeding 
period would indicate that an increased fat störe was not the cause 
of the more gradual utilization of the body resources. 

A study of the differential distribution of the nitrogen in the 
urine serves to bring out certain points with regard to the protein 
metabolism of fasting animals. The percentage of total nitrogen 
occurring as urea-nitrogen decreases in man and is accompanied by 
an increased ammonia-nitrogen excretion. This has been explained 
as due to the condition of acidosis, which may result, at least in 
part, from the accelerated utilization of the fat deposits and the 
decreased oxidative powers of the animal. 



I9I2] Paul E. Howe 95 

In the case of dogs there is a difference o£ opinion as to the 
relation between the urea-nitrogen and the total nitrogen. Schön- 
dorf (22) and others hold that the percentage of urea-nitrogen 
decreases, while in all of our experiments it has remained nearly 
constant, which fact, coupled with the failure to find marked quan- 
tities of organic acids in the urine, would show that dogs are better 
able to utilize their body Stores. This may be due to the fact that 
the dog is naturally a "high-protein" animal. 

The daily Creatinine excretion, which is a constant for any indi- 
vidual under normal conditions of feeding and is generally believed 
to be a function of the muscular metabolism, decreases gradually 
as the fast progresses and in correspondence with the decreasing 
amounit of protoplasm. 

Creatine, which does not occur in normal urines, or is found 
only in cases associated with muscular disintegration, appears dur- 
ing fasting and ordinarily becomes a constant constituent. It has 
recently been shown that the feeding of carbohydrate causes the 
excretion of creatine to stop (5, 17) ; while ingested fats may even 
cause an increase in the excretion of this form of nitrogen. In 
one of our dog experiments (15) there was a disappearance of 
urinary creatine from the iQth to the 59th fasting days. This phe- 
nomenon might be explained on the above basis. It is improbable, 
however, that the body could synthesize sufficient glycogen at this 
stage of the fast to cause the disappearance of the creatine. The 
real explanation is therefore not apparent. 

In connection with the repeated fast, previously mentioned, it is 
interesting to note that the excretions of creatine as well as of total 
nitrogen were practically the same during each of the two fasts, 
notwithstanding the fact that the second fast was twice as long as 
the first. • This would indicate an intimate relation between the 
total-nitrogen excretion and the quantity of creatine excreted. 

When the data representing the creatine and Creatinine excretions 
of a fasting animal are examined, it is seen that there is generally a 
progressive increase in the creatine Output and an accompanying 
decrease in the Creatinine elimination, until the output of creatine 
exceeds that of the Creatinine. In other words, when expressed 
graphically, the curve representing the course of the creatine excre- 



96 General Aspects of Fasting [Sept. 

tion crosses that representing the Creatinine Output. This phenome- 
non has been termed by us the "creatine crossing" and is believed 
to be very significant. It occurs with great uniformity a few days 
previous to the decrease in the total nitrogen excretion that precedes 
the pre-mortal rise of excreted nitrogen. By means of the " creatine 
crossing" the length of the subsequent Hfe tenure of the animal 
may be quite closely estimated. 

Certain pathological constituents, other than creatine, may ap- 
pear in the urine as the result of fasting, such as acetone, diacetic 
acid, lactic acid, bile pigments, albumin, etc. 

The processes in the large intestine during fasting have received 
but httle attention. Various authorities contend that it is difficult 
to make a Separation (2, 18) of fasting feces. Müller (19a) has 
shown that indican, which is now considered as an index of intes- 
tinal putrefaction, disappeared upon the third day of fasting. We 
have been able to make an undoubted Separation of fasting feces 
and have found indican present during the whole of a seven day 
fast on man {22,), Fasting feces are distinct from those of the 
normal individual in that they are of a peculiar brown color and are 
pasty in consistency. The percentage of nitrogen present is higher 
than in normal feces. The bacterial content of feces has received 
but little attention and only recently have results upon the bacterial 
content of fasting feces been determined. The results indicate a 
lovver percentage content of bacteria (3). 

There is not an equal wasting of all of the organs and tissues 
of the body, those organs most necessary for the maintenance of 
life show only a slight decrease in size and weight, while others are 
reduced to but a fraction of their original proportions. Thus the 
heart, lungs, and nervous System exhibit but little change while the 
muscles and fatty tissues exhibit a marked reduction of both volume 
and weight. The organs of regeneration are also resistant to the 
ravages of a fast. This fact is of especial significance for it demon- 
strates the tendency of nature to preserve the species. 

A histological examination of the tissues and organs of fasting 
animals shows a decrease in the volume of the cells as a whole and 
of the nuclei. MorguHs (19) has shown that the decrease in the 
volume of the cells of the Salamander is greater than that of the 



igi2] Paul E. Howe 97 

nuclei and further that the nuclei become elongated. In the case of 
the liver, the cell walls finally begin to disappear and the small 
masses of pigment to clump together. Such a condition does not 
necessarily result in death, for Salamanders of the same size have 
been caused to fast for even a longer time than those whose tissues 
demonstrated these changes ; and, af ter feeding, it was f ound that 
the cell walls again appeared and the liver returned to a normal 
condition. 

We will not take up the question of the localization of the 
degenerative changes, i. e., as to whether they occur in an organ as 
a whole or in localized portions. It is an interesting fact, however, 
that even when the organism is undergoing the degenerative effects 
of a fast, there are still evidences of mitotic division of the nuclei. 

What changes take place which enable one organ to waste away 
while another retains its normal condition? The explanation most 
generally accepted is that of the nourishment of the more vital 
Organs by transference of the nutritive material from the less im- 
portant tissues. Thus the less resistant tissues gradually give up 
their stores of fat and protein to the blood stream which in turn 
furnishes them to the actively functioning organs. 

This idea has received further proof from the researches of 
Hottes.^ This botanist (9) worked with beans and has shown that 
upon removing the cotyledons, and thus the food supply, from seed- 
lings, the meristematic tissue which would normally go to produce 
lateral roots is transferred to the tip of the root (meristem) and 
there used for growth ; and that at the end of from three to f our 
weeks all the cells in the Upper part of the root have lost the major 
portion of their protoplasm and the only actively functioning cells 
are those at the tip. Hottes has also shown that the decrease in 
size of the root is due rather to a reduction in the number of the 
cells than to a mere decrease in size. This is in Opposition to the 
findings of certain zoölogists who hold that the reduction in the 
weight and volume of the organs is due more to reduction in size 
than in number, altho they admit a small decrease in the number of 
contained cells. 

'I am indebted to Professor Hottes of the University of Illinois for this 
Information which was taken from some of his unpublished work. 



98 General Aspects of Fasting [Sept 

The blood of fasting subjects which are ingesting water shows 
in general a decrease in the number of erythrocytes and leucocytes, 
and of the percentage of hemoglobin. The differential distribution 
of leucocytes varies with the species. In the dog (12) there is a 
decrease in the percentage of polymorphonuclear leucocytes and a 
corresponding increase in the small lymphocytes. The changes in 
the other forms of cells are but nominal. 

Fasting studies have been of great importance in the study of 
the minimum of food necessary to maintain life and upon which 
to base the calculation of dietary Standards. Such studies have also 
been utilized in the explanation of phenomena occurring in patho- 
logical States and of metabolism in general. 

Underhill and Rand (25) have explained certain anomalies in 
the urinary changes which occur in pernicious vomiting of preg- 
nancy from their knowledge of fasting metabolism. 

Two agriculturalists have recently made use of the results of 
fasting studies to elucidate problems of importance to both the 
scientist and the farmer. McCollum (16), fed a nitrogen-free diet 
to pigs and studied the efficiency of individual grains as feeding 
stuffs, as well as the nature of the repair processes in protein metab- 
olism. He shows that the difference in the nutritive values of the 
wheat, oat and corn kerneis is not so great as would be expected 
from the difference in the chemical composition; and further, that 
the repair processes of the cell are of a different character from 
those of growth, and that the cellular catabolism and repair do not 
involve the destruction and resynthesis of entire protein molecules. 
This last Statement is not in entire accord with the most widely 
accepted theories of metabolism. Certain zoölogists have also 
shown that the changes of regeneration are unlike those which occur 
in growth. 

Dietrich^ shows (7) that fasting so reduces the plane of metab- 
olism that the quantity of food which was insufficient for main- 
tenance before fasting was afterward sufficient, not only for main- 
tenance, but to produce a positive nitrogen balance ; in other words, 
the animals were more efficient machines after fasting. 

'I wish to thank Professor Dietrich of the University of Illinois for per- 
mission to refer to his unpublished data. 



I9I2] Paul E. Howe 99 

Aron (i), in his studies upon nutrition and growth, subjected 
dogs to incomplete fasts. The results showed that a growing 
animal, receiving only enough food to provide for little or no in- 
crease in weight, " is in a condition of severe starvation." Under 
such conditions the skeleton grows at the expense of the flesh, the 
Organs retain their weight and the brain reaches its normal size. 
The fat and protein of the muscles are largely used up, altho this 
loss of material is balanced by gain of water and by the growth of 
the skeleton. 

The biologists have made use of the fasting subject in the study 
of the Problems of degeneration, of regeneration, and of growth, 
The work of Morgulis and of Hottes already considered was of this 
nature. 

The therapeutic value of fasting is realized in the preliminary 
treatment of some digestive disorders and in the partial fasts of 
obesity eures. These latter eures consist in supplying only the 
protein requirements of the body and thus forcing the individual to 
utilize the surplus fat deposits to make good the energy require- 
ments. 

The increasing populär literature upon fasting and the tendency 
to fast on the part of certain people, especially the pronounced 
physical culturists, and their general good health, would seem to 
indicate that there are some beneficial results to be obtained from 
fasting. The various books upon fasting, of which the superficial, 
yet interesting book of some six hundred pages by Carrington (6) 
upon " Vitality, Fasting and Nutrition " is the most complete, lend 
strength to the idea that fasting as a therapeutic measure is impor- 
tant. The chief contention of this fasting cult is that by depriving 
the body of food the digestive organs are given a chance to recu- 
perate and the body is enabled to rid itself more effectively of the 
waste products and toxic substances. 

Fasting for short and widely-separated periods may be a bene- 
ficial procedure in some individuals. This conclusion is supported 
by the observed effects on dogs, which acquire increased resistance 
from repeated fasting. This view is strengthened, also, by the 
foregoing data pertaining to pigs as well as by Seeland's (24) 
results on pigeons and chickens, which show that repeated fasts. 



loo General Aspects of Fasting [Sept. 

for periods of f rom one to two days, were foUowed by better growth 
and greater strength. 

It is probable, then, that fasting under proper conditions may be 
advantageous. Long fasts, however, seem to be devoid of benefit 
and may endanger health. 

BIBLIOGRAPHY 

1. Aron. The Philippine Journal of Science, 6, i, 1911, 

2. Benedict. Carnegie Publications, 77, 1907. 

3. Blatherwick, Sherwin and Hawk. Proc. Amer. Soc. of Biological Chem- 

ists, 2, 42, 191 1. 

4. Cathcart. Biochemische Zeitschrift, 6, 109, 1907. 

5. Cathcart. Journal of Physiology, 39, 311, 1909. 

6. Carrington. Vitality, Fasting and Nutrition. Rebman Co., N. Y. 1908. 

7. Dietrich. Unpublished data. Illinois Agricultural Experiment Station. 

8. FowLER and Hawk. Journal of Experimental Medicine, 12, 388, 1910. 

9. HoTTEs. Publication of the Carnegie Institution. (In press.) 

IG. HowE AND Hawk. Journal of the American Chemical Society, 33, 215, 1911. 

11. HowE AND Hawk. Proc. Amer. Soc. of Biological Chemists, 2, 65, 191 1. 

12. HowE AND Hawk. American Journal of Physiology, 30, 174, 1912. 

13. HowE, Mattill and Hawk. Journal of the American Chemical Society, 33, 

568, 191 1. 

14. Howe, Mattill and Hawk. Journal of Biological Chemistry, 10, 417, 1911. 

15. Howe, Mattill and Hawk. Journal of Biological Chemistry, 11, 103, 1912. 

16. McCoLLUM. American Journal of Physiology, 29, 215, 191 1. 

17. Mendel and Rose. Journal of Biological Chemistry, 10, 213, 191 1. 

18. Mendel and Fine. Journal of Biological Chemistry, 11, 5, 1912. 

19. MoRGULis. Archiv für Entwicklungsmechanik der Organismen, 32, 169, 1911. 

20. Quoted by Pashutin. Pathological Physiology, 1902. 

21. Richet. Comptes rendus de la Societe de Biologie, 61, 546, 1906. 

22. Schöndorf. Archiv für die gesammte Physiologie, 117, 257, 1907. 

23. Sherwin and Hawk. Journal of Biological Chemistry, 11, 169, 1912. 

24. V. Seeland. Biologisches Centralblatt, 7, 145, 1887. 

25. Underhill and Rand. Archives of Internal Medicine, 5, 61, 1910. 

26. Voit. Zeitschrift für Biologie, 2, 307, 1866. 

27. Voit, E. Zeitschrift für Biologie, 41, 188, 1901. 



THE PHYSICO-CHEMICAL BASIS OF STRIATED 
MUSCLE CONTRACTIONi 

2. Surface tension 

WILLIAM N. BERG 

(WITH PLATE l) 

If the physico-chemical basis of muscle contraction is ever to be 
understood or explained, it is almost certain that it will be brought 
about thru speculation and experiment of a quantitative, rather 
than of a qualitative nature. The mere Statement that muscle con- 
traction is caused by surface tension, or thru osmotic action, etc., 
unless accompanied by quantitative data of an experimental or 
theoretical nature, can add little toward the Solution of the problem 
of the transformation of energy by muscle. It is, perhaps, regret- 
able that so many of the " theories of muscle contraction " which 
have appeared in the recent literature belong to the qualitative class. 
Occasionally someone attempts to treat the subject quantitatively. 
From this point of view the works of Bernstein,^ and of Zuntz^ 
are particularly meritorious, even if the problem has not yet been 
solved by them. 

Among the latest qualitative contributions to the theory of 
muscle contraction, is that of Strietman and Fischer.^ They studied 
the contraction and relaxation of catgut strings immersed in various 
Solutions. By attaching the strings to the usual arrangement of 
iever and recording drum, they found that when a catgut string is 
immersed in water or physiological salt Solution, even for some 
time, no changes in length take place (p. 66). But if the string be 
immersed in Solutions of hydrochloric or lactic acids (w/8o to 

^ Berg, W. N. : Biochemical Bulletin, 1912, i, 535. 
^ Bernstein, J. : Arch. f. d. ges. Physiol., 1901, 85, 271-312. 
^Zuntz, N. : Die Kraftleistung des Tierkörpers. Festrede; Berlin, 1908. 
* Strietman, W. H., and Fischer, M. H. : Ztschr. f. Chemie und Industrie der 
Kolloide, 19 12, 10, 65-77. 

lOI 



I02 Physico-Chemical Basis of Striatcd Musclc Contraction [Sept 

n/20) it contracts. On replacing the acid Solution by water, the 
string relaxes. The relaxation is faster, however, when the acid 
is replaced, not by water, but by a Solution of some salt such as 
sodium bicarbonate, which can neutralize the acid. From their 
diagrams it would seem that a minute or more may be required for 
a Single contraction or relaxation, depending upon the strength of 
the acid, etc., etc. 

These observations are, no doubt, interesting in themselves. 
But before connecting them with muscle contraction, might it not 
be well to consider whether the conditions under which a catgut 
string can contract and relax are at all similar to those existing in 
muscle ? 

Strietman and Fischer State that because lactic acid is formed 
in a working muscle and because a catgut string will contract when 
immersed in a lactic acid Solution and will relax when the acid is 
removed, therefore, in the working muscle, the contraction is 
brought about by the formation of lactic acid. They quote several 
other investigators who have stated their belief in the same idea of 
the connection between lactic acid formation and contractility with- 
out, however, making any of the simple calculations that would 
naturally suggest themselves. 

Their theory is open to the following objections: (i) It is not 
likely that there is any free lactic acid in the working muscle, it is 
probably neutralized at once by the phosphates present in lymph. 
At least this would be inferred from the work of Henderson^ who 
showed that the mixture of phosphates and other substances in blood 
and various tissue fluids was such as to enable them to maintain 
an absolute neutrality in spite of the formation of even considerable 
quantities of acid or of alkali. This point was not overlooked by 
Zuntz (p. 20, 1. c.) when calculating the amount of energy made 
available by the transformation of dextrose into lactic acid. The 
heat of neutralization of the lactic acid by sodium, as well as the 
heat required to separate the sodium from its presumable combina- 
tion with protein, are given due consideration by Zuntz, who cal- 
culated that the heat liberated in the formation of lactic acid from 
dextrose is equivalent to 3.4 per cent. of the heat of combustion of 
dextrose. 

* Henderson, L. J. : Ergebnisse d. Physiologie, 1909, 8, 254-325. 



I9I2] William N. Berg 103 

A repetition of the experiments of Strietman and Fischer, in 
which catgut strings would be immersed in Solutions comparable 
with lymph containing lactic acid (not exceeding the maximal 
amount possible if all of the muscle glycogen were changed at once 
to lactic acid), would probably give results more decisive than those 
in which free acids were used. 

(2) And even if free lactic acid existed in muscle, or if com- 
bined lactic acid could induce proteins to swell, one such Observa- 
tion is only one of very many that are needed for a rational theory 
of muscle contraction. The Statement that lactic acid swells protein 
adds very little to our knowledge of the mechanism in muscle by 
which the potential energy of the food is transformed into the 
kinetic energy of the moving muscle and its load. 

It is to be regretted that the work of Brod^ on the swelling of 
fibrin in acid Solutions has received practically no attention in the 
recent literature. The paper can be profitably studied by those con- 
templating studies on protein swelling. A brief resume of Brod's 
results is given by Berg."^ 

A good example of a quantitative theory of muscle contraction 
is the calculation of Bernstein^ on the possible changes in the sur- 
face energy resident on the muscle fibrils. The method of making 
the calculations is, perhaps, unnecessarily complicated and, in one or 
two instances, the mathematical equations are of doubtful correct- 
ness. Bernstein finds that in order that a muscle may lift an ordi- 
nary load, the surface tension between fibril and sarkoplasm must 
have an improbably great magnitude. He nevertheless concludes 
that the principle, that energy is transformed in muscle thru changes 
in surface energy, is correct. 

There are several reasons why, to the Student at least, a proper 
understanding of some of the recent applications of physical chem- 
istry to biology should be so difficult, if not altogether impossible. 
First: The indefiniteness of certain Statements that the writer has 

' Brod : Beiträge zu der Lehre von der Eiweissverdauung. Dissertation, 
Würzburg, 1892. 

"Berg, W. N. : Amer. Jour. Physiol., 1909, 23, 427. Brod's method has 
recently been used by Tracy and Gies, Biochemical Bulletin, 1912, i, 468. 

* Bernstein, J. : Arch. f. d. ges. Physiol, 1901, 85, 271-312. 



104 Physico-Chcniical Basis of Striated Muscle Contraction [Sept. 

frequently seen in the literature. This is an example taken from 
Freundlich's Kapillarchemie, p. 4: 

Stirface energy = surface tension X area of surface. 

A similar Statement is made by Michaelis,^ and others. Nothing 
further was stated that would enable the reader to use such a 
formula in making calculations were it desired. Expressing the 
surface tension in dynes per centimeter and the area of the surface 
in cm.2, what is the surface energy? The answer is very simple 
after one has taken the time to look the matter up. After a formula 
such as the above, a numerical example ought to be given, so that 
it means more than so many words to the average reader. Suppose 
it is desired to calculate the amount of energy required to form a 
water-surface (in contact with air) of i sq. cm. area? Or, what is 
the same thing, how much energy is liberated or is available for 
external work when the above water-surface diminishes by i sq. 
cm.? According to Michaelis (1. c, p. 14), this will require (or 
liberate) 70 ergs or 7 X lo"'^ kilogram-meters. The method of using 
the formula to obtain this result, simple as it is, was not given by 
Michaelis, altho at least one example of the use of a formula is 
desirable because it will enable the reader to make many other cal- 
culations. 

Following is a numerical example of the kind mentioned above. 
How much energy is required to form a water-surface (against 
air) of I sq. cm.? In the formula it is assumed that the surface 
tension remains constant during the change in area : 

surface energy required ^surface tension X increase in area, 

or 

surface energy liberated = surface tension X decrease in area, 

(ergs) = (dynes per cm.) X (cm.^). 

Since the surface tension of water-air is about 70 dynes per 

cm., it is evident that-^^ X i cm.2^70 ergs = the amount 

cm. 

of energy required. The erg is a unit of work (or energy) and is 
the work done when a mass is moved i cm. by a force of i dyne. 

' Michaelis : Dynamik der Oberflächen, p. 13. Dresden, 1909. 



I9I2] William N. Berg 105 

The element of time does not enter into the definition of the erg. 
The vvork done (ergs) is equal to the product of the force (dynes) 
times the distance (cm.) thru which the force acts. Of course, 
other Units may be used. The surface tension may be expressed in 
grams per cm., and the area in Square cm. The work then is ex- 
pressed in gram cm. But on account of the unfortunate use of the 
Word ' gram ' to designate a certain mass or quantity of matter and 
also to designate a force, it is better, for the present, to use the erg 
and the dyne, and later to convert ergs into kilogram-meters, or 
any other of the customary units for expressing muscular work. 

It is, of course, absolutely necessary that the terms used in such 
calculations be consistent. Here is the second reason why some 
so-called appHcations of physical chemistry to biology are not easily 
followed. An equation will sometimes be given that is not correct 
in its dimensions. To State that 2 sq. cm. = 2 cubic cm. is obviously 
incorrect. Such an inconsistency is to be found in one of Bern- 
stein's^" equations : 'Wir werden daher in dem Falle des isometri- 
schen Tetanus, in welchem alle chemische Energie als Wärme er- 
scheint, dp — ar = c. Wp setzen können, wenn Wp die in einer Zeit- 
einheit erzeugte Wärmemenge, c eine Constante und «p und oLt die 
Oberflächenspannung im Tetanus und in der Ruhe bedeuten. Da 
wir nun oben (S. 296) gesehen haben, dass ctr gegen CLp verhältniss- 
mässig sehr klein ist, so können wir annähernd ap=:c. Wp an- 
nehmen. ' 

Here are two equations in which surface tension is equated with 
work (or heat). It makes no difference what units are used, on one 
side there is a force (surface tension expressible in dynes per cm.) 
and on the other a quantity of energy or work (ergs, or dynes X 
cm.). The constant above referred to is probably meant to be 
the mechanical equivalent of heat. 

These equations are interesting for another reason. It is true 
that in isometric tetanus, a muscle does work in the physio- 
logical sense of the word. But not in the physical sense. In 
physics (or mechanics) work is defined as a product of force 
times distance thru which the force has acted. If either factor 
is zero, the product, work, is zero. The columns that support 

^^ Bernstein, J. : Loc. cii., p. 307. 



io6 Physico-Chemical Basis of Striatcd Muscle Contraction [Sept. 

a biiilding do no work in the physical sense, n'or would a man 
who took the place of one of them; altho physiologically he would 
do a great deal of work. In this case the distance thru which the 
force acts (the weight supported or the upward thrust of the man's 
Shoulders) is zero and hence the work is zero. The above equa- 
tions of Bernstein could be made consistent if there were two 
factors on the left-hand side; one, the surface tension or change in 
surface tension (expressed in dynes per cm.), the other, the area or 
change in area (expressed in cm.^). The product (ergs) could be 
calculated to calories (gram-degrees C.) by dividing by 4.2 X 10''^, 
since i small calorie (gram-degree C.) is equivalent to 4.2 X 10'^ 
ergs. It is difficult to see what the other factor (omitted by Bern- 
stein) can be. In an isometric tetanus the muscle does not change 
its length. In what way can an internal diminution in area take 
place? If the contractu units — whatever their shape may be — do 
not change in length, how do their areas diminish? This difficulty 
does not arise in the case of the ordinary (isotonic) contraction. 
Here one can assume a decrease in the areas of contact between 
contractil unit and sarkoplasm caused by an increase in the surface 
tension between the same surfaces. The product of these two 
quantities, according to the theory, should be an amount of work 
sufficient to account for the external work done and perhaps also 
for the heat liberated at the same time. 

We have stated before that Bernstein's calculations on the 
magnitude of the surface energy changes in muscle are probably 
unnecessarily complex, involving, as they do, several pages of cal- 
culus. The same result is obtained in the following calculations, in 
which two simple quantities are calculated and then compared : 
(i) the amount of energy liberated in a working muscle thru in- 
crease of surface tension times diminution of area of contractil 
Units; and (2) the external work done in lifting a weight a known 
distance. 

Assume that in i c.c. of muscle a right section contains (as 
Zuntz assumes, 1. c, p. 24) 62 million rods, and that there are 800 
such layers, making a total of very nearly 5 X 10^^ rods in i c.c. of 
muscle. Assume the general structure of muscle to be that de- 
scribed by Hürthle (see diagram), and that the muscle rod is the 



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igi2] William N. Berg 107 

CONTRACTIL UNIT (Hürthle, p. 157). From the dimensions 011 the 
accompanying diagram, it is evident that the lateral area of a rod 
diminishes from 4.8 /a^ when relaxed, to 2.8 /^^ when contracted. 
(We omit the simple geometrical calculation.) The base areas are 
increased, but the energy apparently required for this work will for 
the present be disregarded — it is an additional load on a probably 
overloaded theory. Since the total area of the relaxed rod (5.2 /a^) 
is greater than that of the contracted rod (3.8 fi^), it follows that 
there is an increase in the surface tension immediately preceding 
the contraction, according to the requirements of the theory. To 
calculate the energy liberated as being equivalent to the diminution 
in area times the surface tension of water is probably incorrect, for 
without an increase in surface tension there seems to be no reason 
why the rods should contract against the external resistance — the 
downward pull of the weight lifted. The rod contracts presum- 
ably, because in the relaxed State the surface tension on the rod 
surface is low. How low can it be ? It may be as low, perhaps, as 23 
dynes/cm. if we assume the rod to be covered with a layer of pure 
acetic acid, and that the acid has the same surface tension in con- 
tact with the water that it has when in contact with air. Other fatty 
acids also give low values for the surface tension of their Solutions, 
and they have still lower surface tensions in the pure State. Pres- 
ently, the surface tension is raised, presumably by the removal of 
the fatty acid or other agent causing low surface tension, by instan- 
taneous combustion, let us say. How high can the surface tension 
be raised? It might be raised" to 85 dynes/cm. if we imagine the 
rod now to be covered with a layer of saturated sodium chlorid Solu- 
tion. The surface tensions of aqueous Solutions of salts cannot be 
raised^^ very muchbeyond that of pure water, which varies between 
y2 to y6 dynes/cm. (at 18° C), according to the method of measure- 
ment. The upper limit for any Solution that possibly could exist in 
the muscle might be assumed, then, to be the value for saturated 
sodium chlorid Solution, or any other concentrated salt Solution that 
might probably occur in living muscle. Of course, the existence of 
the films of pure acetic acid and of strong salt Solution over the 

" Freundlich, H. : Kapillarchemie, p. 27 and 62. Leipzig, 1909. 
*^ Heyd Weiler, A. : Ann. Physik., 1910, 33, 145-185. 



io8 Physico-Chemical Basis of Striatcd Musclc C ontraction [Sept. 

muscle rods is purely hypothetical. The surface tension theory 
requires that changes in surface energy take place, and from what 
follows it is apparent that these changes must be great — greater, in 
fact, than the probable actual change on the rod surface. For it 
seems hardly possible that such great changes in concentration and 
in surface tension could take place. The values for the surface 
tensions of pure acetic acid and concentrated salt Solution have been 
taken from the literature ; whether such limiting values are ever 
reached in living muscle is, for the present, purely hypothetical. 

If it be assumed that, during the chemical changes taking place 
in a working muscle, the inorganic ions in the rod-surface film 
rapidly change their concentrations, the film might be regarded as 
an electrical double layer or Helmholtz double layer. Without a 
doubt, changes in surface tension would result from the changes in 
ion concentration. The more ions in one of these layers covering 
a rod, the more they repel one another and the lower is the surface 
tension, and vice-versa. But as has been pointed out before,^^ it is 
not certain that such a double layer really exists between living 
particles and their surrounding medium. And even if there were 
such a layer, the total change in surface tension in such a layer is 
hardly significant for the present purpose. The small Variation in 
surface tension when the Variation is caused only by ions was prob- 
ably overlooked by Robertson^"* and others who advocated a capil- 
lary electric theory of muscle contraction. It is really a special 
case of surface tension in which the variations in surface tension 
are caused by the mutual repulsion of the ions in each of the layers. 
But insofar as small amounts of certain organic substances, such 
as fatty acids, can affect (depress) the surface tension of water 
very much more than even improbably large amounts of inorganic 
salts, the surface tension theory is given the benefit of the greatest 
possibilities by assuming the changes in concentration from pure 
acetic acid (23 dynes/cm.) to saturated sodium chlorid Solution 
(85 dynes/cm.). This is as large a difference as can be assumed 
from the experimental data on the surface tensions of Solutions. 

"Berg, W. N. : New York Med. Journal, 1907, July 20 and 27; and Ion, 
1910, 2, 161-188. 

"Robertson, T. Brailsford: Trans. Royal Soc. South Australia, 1905, 29; 
and Quarterly Jottr. Exper. PhysioL, 1909, 2, 303-316. 



I9I2] William N. Berg 109 

If in I c.c. of muscle there are 5 X lo^*' rods, the lateral area of 
each of which diminishes from 4.8 /a^ to 2.8 /^^ when the muscle con- 
tracts, the total reduction in area is 5 X 10^" X 2 /x,2__io^i fx^ = 
lO'^ cm. 2 (i ju. = 0.001 mm.). The calculations will be simplified 
if it be assumed that the increase in surface tension is instantaneous, 
giving the contracting muscle the largest surface tension during the 
entire contraction phase. Then since 
surface energy liberated = diminution in area X surface tension, 

(e.,s) (c..^) {^) 

the energy liberated is 1000 X 85 ergs. Let it be assumed that all of 

this is transformed into external work — lifting a weight — and that 

the resultant heat arises from the activity of a different mechanism ; 

in short, that the muscle is an engine having an efficiency of 100 

per Cent. How great a weight will this i c.c. of muscle lift? Since 

there are 800 layers of rods, and each layer shortens by 3 /-i during the 

contraction (see Plate i herewith), the muscle shortens by 2400 /a 

or 2.4 mm., lifting a mass of W grams 2.4 mm. The energy (ergs) 

expended in lifting a mass of W grams thru the distance D (cm.) 

is PF X ■C' X 981 ergs, since gravity = 98i dynes. Therefore the 

8s,ooo 
85,000 ergs will hft ^^3^ = 361 grams. 

According to Zuntz (1. c, p. 23) i gram of muscle substance 
can do 0.002 kilogram-meter of work in one contraction under 
favorable conditions. If this muscle shortened 0.24 cm. as the 
above muscle did, it would lift a trifle more than 800 grams. Bern- 
stein^^ mentions 600 grams at least, as the pull of i cm.^ of frog 
muscle in an isometric contraction. Insofar as i cm.^ of many 
kinds of muscle can support without lengthening (but not lift) 
several kilograms — about 6 kilograms for human, and probably 
more for certain types of insect muscle — the above figure of 361 
grams, as the weight a muscle could lift, is small, especially when it 
is borne in mind that it is an improbable maximum. 

The foregoing discussion may be summarized as f oUows : 

I. Too often there is a general lack of definiteness in the mathe- 

" Bernstein, J. : Arch. f. d. ges. PhysioL, 1905, 109, 326. 



HO Physico-Chemical Basis of Striated Muscle Contraction [Sept. 

matical treatment of a biological problem. Formulae are stated 
vvith no information as to their use or application to the problem 
under disciission. 

2. Bernstein's calculations on the surface energy changes in 
working muscle are criticized. A much simpler method of calcula- 
tion is used with a result similar to Bernstein's, namely, the energy 
expended by a working muscle is much greater than the probable 
changes in surface energy can furnish, Of course, future investi- 
gations may bring to light sources of surface energy within muscle 
as yet unknown. 

Washington, D. C. 



A STUDY OF SOME PROTEIN COMPOUNDS 

WALTER H. EDDY 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

Contents. (A) Morphin mucoid, 112; (B) strychnin mucoid, 114; (C) conin 
mucoid, 115; (D) piperidin mucoid, 115 ; (E) anilin mucoid, 115; (F) morphin 
nucleoprotein, 115; (G) morphin caseinogen, 116, strychnin caseinogen, 116, 
calcium caseinogen, 116; (H) strychnin ovo-mucoid, 117; (I) histon mucoid, 
118; (J) histon nucleoprotein, 121; (K) histon ovo-mucoid, 121. 

I. INTRODUCTION 

When I began Ph.D. work in this laboratory, six years ago, 
Dr. Gies was actively engaged in studies of the properties of various 
protein Compounds which he had prepared as early as 1904.^ He 
inaugurated that work from the Standpoint of his interest in the 
chemical composition of protoplasm, and the nature of the struc- 
tural and dynamic relationships of cell constituents and products. 
He believed that the knowledge gained from studies of artificial 
protein Compounds would pave the way for successful inquiry into 
the nature of the protein correlations in the cells — relationships of 
the most fundamental biological character. At his Suggestion, and 
in furtherance of this object, I have conducted the experiments de- 
scribed in this paper.^ 

The general plan of the research was: (A) The production of 
protein salts by combining organic hases, such as strychnin, mor- 
phin, conin, piperidin, etc., with acid-reacting proteins, such as 
tendo-mucoid, ovo-mucoid, yeast nucleoprotein, etc.; and {B) the 
production of protein salts by combining hasic proteins, such as 

^Gies: Proc. See. Path. and Physiol., Amer. Med. Assn., 1906, p. 121. 

''The detailed results of this work have been described in the writer's 
dissertation, On the synthesis of some protein salts, Columbia University, 1909 
(pp. 61). A preliminary report was published by Eddy and Gies in the Pro- 
ceedings of the Society for Experimental Biology and Medicine, 1907, iv, pp. 
I4S-6. 

III 



112 Some Protein Compounds [Sept. 

histons and protamins, with the acid-reacting proteins eiiumerated 

above. 

In developing the latter part of this plan certain anomalies arose 
in connection with the preparation of thymus histon, which led to 
a collateral investigation of histons. The results of the latter 
studies will be embodied in a future paper. (See page 169.) 

IL EXPERIMENTAL 

I. Salts of various proteins with organic bases. 'Ä. Mor- 
phin MUCOiD. Purification of the materials. The first step in the 
preparation of a typical product was the removal of free alkali f rom 
the base — and free acid from the protein. Chemically pure, pul- 
verized, morphin was washed with distilled water until the wash- 
ings were entirely neutral to litmus. Tendo-mucoid was prepared 
after the manner of Cutter and Gies^ but dehydration with alcohol 
and ether was omitted. The dry scales w^ere soaked in distilled 
water until they softened. The protein was then washed with dis- 
tilled water until the washings were entirely neutral to litmus. 

Union of base and protein. The base and the protein were then 
triturated together in a mortar, a very little water being added to 
ensure an intimate mixture. A mechanical excess of the base was 
used in every case. Evidence of chemical action was seen in the 
peculiarly viscid, smeary character of the mixture. Mucoid and 
w-ater give a thick, milky mixture, but it is not viscid or smeary. 
The mixture w^as finally treated with sufficient water in excess to 
dissolve the product. The viscid liquid was filtered through a wet, 
fluted, hardened, filter paper, but the first portions of iiltrate were 
returned to the paper until a clear opalescent liquid appeared. This 
filtrate was neutral to litmus. 

Purification of the product. The filtrate, preserved with tol- 
uene, was subjected to continuous dialysis in a parchment bag, im- 
mersed in frequently renewed distilled water, until the dialysate, 
even when concentrated to a very small volume at 40° C, gave no 
test for the base (morphin). The contents of the bag were then 
evaporated to dryness at 40° C, toluene being used and frequently 
renewed during the process. The resultant dry produot was then 

' Cutter and Gies : Amer. Journ. PhysioL, 1902, vi, pp. 155-6. 



I9I2] Walter H. Eddy . 113 

pulverized in a mortar and extracted three times with a large excess 
of ether for the removal of traces of admixed free base (morphin). 

Isolation of the product. The powder was next dissolved in a 
small amount of water and this Solution poured into a mixture of 
Yz ether and Yz alcohol. A copious precipitate resulted. The pre- 
cipitate was gelatinous and dissolved easily in water. After dissolv- 
ing the precipitate in water and filtering the Solution, the filtrate 
was precipitated with alcohol-ether. This process was repeated sev- 
eral times. The final product was dehydrated in the usual manner 
with alcohol and ether. 

Special difficulties in the preparation of morphin mucoid. The 
first Solutions of the Compound filtered very slowly. It was found 
that this was due to excess of mucoid. When a large excess of 
morphin was used there was less insoluble mucoid residue and filtra- 
tion became correspondingly more rapid. 

Precipitation of the purified product with alcohol became in- 
creasingly difficult with the increasing purity of the product. Am- 
monium Sulfate, in excess, precipitated the product from its aqueous 
Solution, but long dialysis was required to remove the salt. 

The purified product failed to respond to the iodic acid test for 
morphin^ This fact was carefully investigated. The results 
showed that the failure was not due to the quality of the iodic acid 
used nor to interference with the test by the mucoid. In the puri- 
fication of the product there seemed to be continuous loss of mor- 
phin. This was presumably due to hydrolytic dissociation. 

Evidence of the compound-naturc of the product. The product 
was water-soluble, demonstrating that it was neither mucoid nor 
morphin, nor a mechanical mixture of the two. The aqueous Solu- 
tion of the product frothed strongly on shaking and gave a good 
biuret test, indicating its protein character. Addition of a few 
drops of 0.2 per cent. hydrochloric acid Solution yielded a flocculent 
precipitate of mucoid. 

Conclusions regarding morphin mucoid. Morphin and mucoid 

*For the detection of morphin, the iodic acid test was applied as follows: 
I c.c. of the Solution to be tested was added to an equal volume of dilute 
sulfuric acid Solution. To this was added a few c.c. of iodic acid Solution and 
finally a little Chloroform. After vigorous shaking, the presence or absence of a 
violet coloration served to indicate the presence or absence of morphin. 



114 Some Protein Compounds [Sept. 

react to form a water-soliible protein Compound which, in aqneous 
Solution, yields mucoid on treatment with 0.2 per cent. hydrochloric 
acid. The morphin enters into combination in a proportion so small 
as to be incapable of responding to the iodic test, or is united in 
such a way as to fail to respond to the test. 

B. Strychnin mucoid. In view of the extreme delicacy of 
the dichromate test for strychnin, this base was selected for the 
second series of preparations. Care was taken to insure purity of 
the original materials as in the preparation of the morphin-mucoid 
products. 

Preparation. The method of preparation was identical with 
that for morphin mucoid (page 112) except in the following details: 
The final water-solution of the product was again evaporated to dry- 
ness at 40° C. and the dry matter, after pulverization, was extracted 
with Chloroform. Twenty-six voluminous washings were neces- 
sary to free the powder from admixed strychnin and to obtain a 
strychnin-free washing. In view of the insolubility of strychnin 
in water and its ready solubility in ether this result seemed difficult 
to explain on any other basis than partial dissociation by the 
Chloroform. 

Evidence of chemical combination. The protein character of 
the Compound was established by the following results : The water- 
solution gave a strong biuret test; was precipitable by Saturation 
with ammonium sulfate or magnesium sulfate; gave a flocculent 
precipitate with 0.2 per cent. hydrochloric acid and 4 per cent. acetic 
acid Solutions ; and f rothed strongly on shaking. The aqueous Solu- 
tion of the product was neutral to litmus. 

The presence of strychnin was shown by the intense bitter taste 
and by strong "dichromate tests." Filtrates from precipitates 
formed by addition of 0.2 per cent. hydrochloric acid Solution 
yielded, in every case, strong "di-chromate tests " for strychnin. 
Four physiological tests were also made to establish the presence 
of the strychnin. The results and methods follow : 

The lethal dose of strychnin sulfate is about 2.5 mg. per kilo of 
weight for frogs and 7.6 mg. per kilo of weight for dogs. Vol- 
umes of aqueous Solution of strychnin mucoid (0.575 ^S- P^^* ^■^•) 
containing quantities equal to the lethal dose of strychnin sulfate 
were injected subcutaneously in frogs and dogs. 



I9I2] Walter H. Eddy ' 115 

In the first of two experiments on frogs, the initial dose failed 
to produce any strychnin effects. An effect followed the second 
injection of an equal dose, but it required three doses to produce 
Opisthotonus. Recovery was complete. In the second frog, each 
of two doses injected successively produced Opisthotonus. The 
frog recovered. 

For the first dog a double dose was required to produce hyperes- 
thesia and tetanus. The results with the second dog duplicated 
those with the first. 

In all these physiological tests the strychnin appeared to be 
liberated slowly in the animal, the effects Coming on gradually and 
extending over a period of 3-5 hours, with complete recovery. 

Concliisions regarding strychnin mucoid. The results seemed to 
leave no doubt regarding the compound-nature of this product. 
Whether it is a true salt or an adsorption Compound can not be de- 
cided from the available data, but its neutrality, its water-solubility, 
and its power to yield both strychnin and mucoid, strongly suggest 
the production of a salt by a process directly comparable to the 
neutralization of base by acid. 

The physiological tests show that the Compound evidently con- 
tains a much smaller proportion of strychnin than that in the com- 
mon Sulfate. The quantitative examinations have not yet been 
completed. 

C. Conin mucoid. Conin combines with mucoid very rapidly 
and yields a Solution which filters easily. The product, af ter purifica- 
tion by dialysis and alcohol precipitation, is water-soluble and biuret- 
reacting. As in the case of morphin mucoid, however, it was im- 
possible to demonstrate the presence of the alkaloid. All tests 
were negative with the potassio-mercuric iodid and phospho-tung- 
stic acid reagents. 

D. PiPERiDiN MUCOID. The purified piperidin product gave the 
protein tests and also a test for piperidin with platinic chlorid. 

E. Anilin mucoid(?). A water-soluble product of mucoid 
and anilin was obtained but the anilin disappeared early in the puri- 
fication process. 

F. Morphin nucleoprotein. Two attempts were made to 
produce a Compound of morphin with yeast nucleoprotein. The 



ii6 Sonic Protein Compounds [Sept. 

method of preparation was similar to that described on page 112. 
Neither attempt was successful in establishing the presence of mor- 
phin in the final product. A water-soluble protein of different char- 
acter from the nucleoprotein resulted in each case. 

G. Morphin caseinogen, strychnin caseinogen, and cal- 
cium CASEiNOGEN. Studies were made of the effects of morphin, 
strychnin and calcium hydroxid on caseinogen. In each case water- 
soluble, biuret-reacting products were obtained. Rigorous purifi- 
cation was not attempted. 

Salts of ovomucoid. Neumeister^ investigated a glucoprotein 
in eggs which he named " pseudopeptone." This Compound was 
studied by Salkowski,^ Mörner/ and Eichholz,^ and called by them 
" ovo-mucoid." As a " cell-protein," this substance seemed to offer 
good material for our experiments. A pure product was prepared 
by Mörner's''' well-known process. 

Preparation of ovo-mucoid. (From eggs.) With increasing 
purity, precipitation with alcohol became correspondingly difficult. 
Alcohol-ether did not remove this difficulty but the addition of a 
few drops of sodium chlorid Solution brought about precipitation in 
every case. The final water-solution was freed from chlorid by 
dialysis in a parchment bag in the presence of toluene. The Solu- 
tion, which then was acid to litmus, was evaporated to dryness at 
40° C, yielding yellow flakes which were ground to a white powder. 

Properties of the ovo-mucoid product. This ovo-mucoid was 
readily soluble in water and gave a good biuret test. The water- 
solution frothed on shaking, but was not viscid. Phosphotungstic 
acid, 0.2 per cent. hydrochloric acid, 4 per cent. acetic acid and 
tannic acid Solutions precipitated the aqueous Solution, which was 
acid to litmus. 

(From shad roe.) The roe was ground in a mortar with sand 
and this mixture poured into boiling, slightly acidulated, water. 
The remaining Steps were identical with those for the preparation of 
ovo-mucoid from eggs and the product responded to the same tests. 

These two products were used in the following studies. 

' Neumeister : Zeitschrift für Biologie, 1890, xxvii, p. 331, 

'Salkowski: Centralhlatt f. d. med. Wissensch., 1893, xxxi, pp. 513 and 706. 

^ Mörner : Zeitschr. f. physiol. Chem., 1893, xviii, p. 525. 

' Eichholz : Journ. Physiol., 1898, xxiii, p. 163. 



igi2] Walter H. Eddy 117 

H. Strychnin ovo-mucoid (egg). The method of prepara- 
tion followed the lines of the morphin-mucoid process (page 112) 
with the following abbreviation : After dialysis the Solution was at 
once precipitated with absolute alcohol. No other methods of puri- 
fication were used. 

The filtrate f rom the original mixture of ovo-mucoid and strych- 
nin was turbid and acid to litmus, but became neutral on standing, 
in the presence of toluene. On dialysis, and consequent dilution 
with water, the Solution clarified. The dialysate on the other band 
became turbid but failed to give a protein or strychnin test, 

When the dialyzed liquid was treated with absolute alcohol, in 
excess, a mixed, cheesy and gelatinous precipitate was produced. 
The alcoholic filtrate from this precipitate was acid and gave a 
strychnin test, suggesting dissociation. The precipitate dissolved 
readily in water and the Solution was then filtered. It was nozv acid 
in reaction and gave no strychnin test. Precipitated again with 
alcohol, the solid product failed to give the strychnin test, was acid 
and resembled in every way the original ovo-mucoid. 

A portion of this precipitate was dissolved in water and the So- 
lution evaporated to dryness at 40° C. A new trituration with 
strychnin was made with this product. The results were the same 
as with the first preparation, viz., a turbin Solution that cleared on 
dilution with water by dialysis and gave in this condition both 
strychnin and protein tests. Alcohol again dissociated it into 
strychnin and ovo-mucoid (?). 

From the above results it was deemed desirable to make a care- 
ful study of the reactions of the product and a second preparation 
was conducted for this purpose. The turbid filtrate obtained from 
the initial mixture of strychnin and ovo-mucoid was found to be 
actually amphoteric to litmus, though acid to Phenolphthalein. Its 
alkalinity to litmus was not increased by retriturating it with strych- 
nin. Dilution with water resulted again in a clear Solution, giving 
both strychnin and protein tests. On standing for a considerable time 
in a parchment bag, in the presence of toluene, the amphoteric reac- 
tion gradually disappeared and the Solution became distinctly acid to 
litmus. It also finally yielded a precipitate in the bag and lost its 
power to respond to the strychnin test. The turbid dialysate grad- 



ii8 Sonic Protein Compounds [Sept. 

ually acquired protein material, but the frequent renewals of water 
and large voliime made it impossible to determine the presence of 
strychnin. Apparently complete dissociation resulted, but neither 
the character of the dissociation products nor the manner in which 
the strychnin separated was determined. 

Strychnin ovo-niucoid (roe). The results with ovo-mucoid 
from shad roe were identical with those in the case of tgg ovo-mu- 
coid except that the disappearance of the strychnin on dialysis was 
much slower. It was ten days before the contents of the bag failed 
to give the strychnin test. Concentration of the dialysates in this 
case before applying the strychnin test failed to make its detection 
possible. 

Evidence of the Compound natnre of the ovo-mucoid products. 
The clear amphoteric Solution, with its response to strychnin and 
protein tests, indicates a chemical combination, especially in view 
of the water-insolubility of strychnin. The dissociability in alcohol 
of the shad roe product, and the results of dialysis, indicate that it 
is more stable than the strychnin product with &gg ovo-mucoid. 
Again the question of whether we are here dealing with a true 
chemical Compound or with an adsorption product remains open for 
further investigation. 

2. Protein-protein Compounds. The foregoing experiments 
were preliminary to attempts to bring about combinations between 
acid-reacting and basic-reacting proteins, such as protamins and 
histons. 

/. HiSTON MucoiD. Preparation of histon hydrochlorid. His- 
ton was prepared by the method of Huiskamp.^ Thymus glands 
from freshly killed calves were freed from fat with a knife and 
minced in a meat chopper. The hash was then placed in a large 
bottle and extracted in an ordinary ice box for 24-48 hours with 
distilled water. About 300 c.c. of water were used with each 100 
grams of thymus. The extract was filtered through wet fluted filter 
papers. Nucleohiston was precipitated from the filtrate with 5 c.c. 
of IG per Cent, calcium chlorid Solution per 100 c.c. of extract. The 
precipitate was then filtered off and redissolved in water to which a 
little ammonia had . been added. This Solution was filtered and 
reprecipitated with calcium chlorid Solution in the usual way. The 

* Huiskamp : Zeitscjir. f. physiol. Chemie, 1901, xxxii, p. 145. 



igi2] Walter H. Eddy 119 

precipitate was then extracted with 0.8 per cent. hydrochloric 
acid for the production of the hydrochlorid. This extract of histon 
hydrochlorid was finally dialyzed in a parchment bag against dis- 
tilled water until neutral to litmus. This Solution of histon hydro- 
chlorid was used for the preparation described below. 

Preparation of potassium mucoid. Acid-free mucoid was dis- 
solved in 0.3 per cent. potassium hydroxid Solution and the liquid 
filtered. The filtrate was then dialyzed in a parchment bag against 
distilled water (in the presence of toluene) until neutral to litmus. 
The product in this neutral Solution was presumably potassium 
mucoid. 

Preparation of histon mucoid. Histon hydrochlorid Solution 
was added drop by drop to the potassium mucoid Solution. A pre- 
cipitate formed immediately, and sedimented quickly beneath the 
clear supernatant liquid. Excess of the histon hydrochlorid Solu- 
tion dissolved the precipitate. The product was then filtered off 
and washed with water until the washings no longer gave precipi- 
tates with ten per cent. ammonium hydroxid or 0.2 per cent. hydro- 
chloric acid Solution. 

Evidence of the Compound nature of the histon mucoid product. 
A portion of the precipitate was triturated with 0.05 per cent. so- 
dium carbonate Solution. A colloidal Solution was obtained. Its 
filtrate gave a heavy precipitate with 0.2 per cent. hydrochloric 
acid Solution and a distinct precipitate with ammonium hydroxid 
Solution. 

These results did not determine whether the sodium carbonate 
merely dissolved the histon mucoid, or dissociated it into a histon So- 
lution and a sodium mucoid Solution. To ascertain these points 
the following tests were made : 

(a) A portion of the sodium carbonate Solution was poured into 
95 per cent. alcohol. It failed to precipitate at once or on Standing. 

(b) A portion of the sodium carbonate Solution was poured into 
95 per cent. alcohol, to which one drop of 10 per cent. sodium 
chlorid Solution had been added. A precipitate appeared on 
Standing. 

(c) Alcohol-ether failed to precipitate the Solution but with the 
addition of a drop of salt Solution a precipitate appeared. 



120 Some Protein Compounds [Sept. 

{d) The precipitates obtained in {h) and (c) failed, in this first 
set of tests, to dissolve in water and the washings gave no hydro- 
chloric acid or ammonia precipitate. The precipitates dissolved in 
0.05 per Cent, sodium carbonate Solution and the filtrates gave both 
the ammonia and hydrochloric acid tests. 

{e) Histon hydrochlorid Solution was not precipitated by alcohol 
even when sodium chlorid was present. Alcohol also failed to pre- 
cipitate potassium mucoid Solution but did so in the presence of a 
trace of sodium chlorid. 

These results Warrant the inference that the sodium carbonate 
acted as a solvent rather than as a dissociant. They also indicate 
that precipitation by alcohol in the presence of salt served to dif- 
ferentiate the histon mucoid from the histon hydrochlorid, and that 
our product was a Compound and not a mixture. 

A Solution of histon hydrochlorid was tested with an excess of 
alcohol in the absence of salt. A similar Solution of potassium 
mucoid was made. When these two clear Solutions were mixed a 
precipitate of histon mucoid formed at once. This histon mucoid 
was then dissolved in 0.05 per cent. sodium carbonate Solution and 
the filtered Solution precipitated with alcohol in the presence of a 
little salt. This precipitate, unlike that above (d), dissolved readily 
in zvater. The water-solution gave both the ammonia precipitate 
and the hydrochloric acid precipitate. This and similar results indi- 
cated the formation of a soluble histon mucoid Compound. 

Finally a new histon mucoid product w^as made by the original 
method. This product was washed free from excesses of both his- 
ton hydrochlorid and potassium mucoid, as before, and then treated 
as f ollows : 

A portion was macerated in a mortar with o.i per cent. hydro- 
chloric acid Solution and a second portion in another mortar with 
0.1 per cent. potassium hydroxid Solution. These liquids were fil- 
tered. The acid filtrate gave a precipitate with ammonia but not 
with hydrochloric acid. The alkali filtrate gave a heavy precipitate 
with hydrochloric acid but none with ammonia. 

These results suggest that mixtures of (a) histon hydrochlorid 
and potassium mucoid Solutions yield a precipitate of histon 
mucoid; (&) pure histon mucoid and o.i per cent. hydrochloric acid 



I9I2] Walter H. Eddy 121 

Solutions yield histon hydrochlorid and insoluble mucoid; (c) pure 
histon mucoid and o.i per cent. potassium hydroxid Solutions yield a 
potassium mucoid histon complex. 

The failure to get a histon precipitate with ammonia in the 
potassium hydroxid extract may have been due to the small amount 
of resultant histon mucoid or to the formation of an insoluble form 
o£ histon, such as ammonia produces. Whatever the explanation of 
this failure, there seemed to be no doubt of the power of histon 
to combine with mucoid to form a Compound different in proper- 
ties from either histon hydrochlorid or potassium mucoid. 

/. Histon nucleoprotein (yeast). Neutral potassium nu- 
cleoprotein (obtained by dissolving yeast nucleoprotein in o.i per 
cent. potassium hydroxid Solution and dialyzing free from hydroxyl 
ions) combines with histon hydrochlorid in the same way as potas- 
sium mucoid (page 119). Much more of the Solution of histon 
is necessary for the production of the salt. The product was similar 
to histon mucoid in being insoluble in water; in dissolving readily in 
0.05 per cent. sodium carbonate Solution but incompletely in 0.5 
per cent. sodium carbonate Solution ; and in forming, with sodium 
carbonate, a water-soluble sodium-histon nucleoprotein complex. 

K. Histon ovo-mucoid. Preparation. The ovo-mucoid {tgg) 
was purified to such a degree as to be practically soluble in salt-free 
alcohol (page 116). A similarly pure Solution of histon hydro- 
chlorid was used. 

When the water Solutions of these two substances were com- 
bined, a precipitate formed slowly. A slight excess of the histon 
Solution dissolved the precipitate. The precipitate dissolved to a 
turbid Solution in 0.05 per cent. sodium carbonate Solution. This 
turbid fluid was filtered and divided into two portions. One por- 
tion was poured into 95 per cent. alcohol to which 3 drops of 10 
per cent. sodium chlorid Solution had been added. The other por- 
tion was saturated with ammonium sulfate. Both portions gave 
heavy precipitates, which were soluble in water; the Solutions were 
precipitated in part by ammonia. When purification of the ammo- 
nium sulfate precipitate by dialysis was attempted, the Compound 
broke down. The "alcohol precipitate" was hydrolyzed with 
hydrochloric acid. The resultant liquid, neutralized with potassium 



122 Some Protein Compounds [Sept. 

hydroxid, rediiced the Fehling-Benedict reagent. This result, with 
the precipitation by ammonia, seemed to show the presence of both 
glucoprotein and histon in the precipitate. 

When alcohol Solutions of ovo-mucoid and histon hydrochlorid 
were poured together, a precipitate formed at once that gave both 
the ammonia test and the reduction test. The latter process is the 
simplest and quiekest method of obtaining this product. 

The results of these researches have shown that the methods for 
the preparation of histon, as outlined in the literature, are in serious 
need of revision. In fact, the results suggest that so-called histon 
is a protein salt rather than a simple protein. In a future paper will 
be presented the findings in regard to histon preparation. 



EFFECTS OF INTRAPERITONEAL INJECTIONS OF 

EPINEPHRIN ON THE PARTITION OF NITRO- 

GEN IN URINE FROM A DOG 

JACOB ROSENBLOOM and WILLIAM WEINBERGER 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

I. INTRODUCTION 

The action of epinephrin on nitrogenous metabolism has been 
the object of investigation by several authors. The experimental 
results of Kraus and Hirsch/ and Quest,^ indicate that intravenous 
or subcutaneous injections of epinephrin exert very Httle influenae 
on the nitrogenous metaboHsm of healthy dogs, the insignificant 
increase of eliminated nitrogen being caused both by the glycosuria 
and (after subcutaneous injections) by skin necrosis. Fasting ani- 
mals seem to be differently affected. Falta and Rudinger,^ and 
Underhill and Closson* were able to show an accelerating influ- 
enae, on protein metaboHsm, of subcutaneous and intravenous injec- 
tions of epinephrin. 

Underhill and Closson have shown that the subcutaneous injec- 
tion of " adrenalin chlorid " Solutions into dogs is not attended by 
any significant change in the proportions of the urea-, ammonia- and 
creatinin-nitrogen of the urine, in partial disagreement with Paton,^ 
who also found that, although on a sufficient diet, the catabolism of 
proteins is not interfered with, there is a markedly increased produc- 
tion of ammonia. 

In all the above mentioned experiments the epinephrin was in- 
jected into veins or into subcutaneous tissues. The intraperi- 
toneal way has not been utilized by previous observers in this con- 

^ Kraus and Hirsch. Cited by Kraus and Friedenthal : Berl. klin. Woch., 
1908, xlv, p. 1709. 

^Quest: Zeit. f. exp. Path., 1908, v, p. 43. 
' Falta and Rudinger : Central, f. klin. Med., 1908, Ixvi, p. i. 
* Underhill and Closson : Anier. Journ. Physiol., 1906, xvii, p. 42. 
' Paten : English Journ. of Physiol., 1903, xxix, p. 286 ; 1904, xxxii, p. 59. 

123 



124 Epinephrin Effects on Metabolism [Sept. 

nection, althongh it is possible that this mode of administering 
epinephrin has a different effect on nitrogenous metabolism, as is the 
case in carbohydrate metabolism (Löwi).® 

IL DESCRIPTION OF THE EXPERIMENTS 
This investigation consisted of two metabolism experiments. 
One animal was used for both experiments. An intermediate pe- 
riod served to allow the animal to recuperate from the effects of the 
first experiment before the second one was begun. The metabolism 
work was conducted by the general methods in use in this labo- 
ratory.'^ 

We determined the nitrogen content in the several ingredients 
of the food. Urinary nitrogen, in the leading forms, was deter- 
mined as f ollows : ammonia, each day ; total, urea, creatin and Crea- 
tinin (as Creatinin), every third day; purins, at the end of each 
period. The urine was preserved with thymol. Total nitrogen was 
determined by the Kjeldahl process ; ammonia and Creatinin by the 
Polin methods f iirea^ by Benedict's method ;^^ purin nitrogen by a 
combination of the Arnstein" and Salkowski^^ methods. 

The authors used two specimens of the colorless "adrenalin 
Chlorid" (i:i,ooo) of Parke, Davis and Co. They were pur- 
chased in the open market. Each was tested for its pressor action 
at the conclusion of the corresponding injection experiments and 
was then found to be practically as active as ever. Varying 
amounts and concentrations of "adrenalin chlorid" were injected 
into the peritoneal cavity ; in the first experiment the concentration 
was 1 : 10,000 — in the second, i : i,ooo. In one injection period of 

*Löwi: Von Noorden's Metabolism and practica! medicine, 1907, iii, p. 1181. 

' Mead and Gies : Anier. Journ. Physiol., 1901, v, p. 106 ; also Gies and collab- 
orators: Biochemical Researches, 1903, i, Reprint No. 21; Gies: Amer. Journ. of 
Physiol., 1905, xiv, p. 403; Gies: Amer. Journ. of Physiol., 1901, v, p. 235; also 
Gies and collaborators : Biochemical Researches, 1903, i, Reprint No. i ; Gies : 
Proc. Amer. Physiol. Soc, Amer. Journ. of Physiol., 1904, x, p. 22; Hawk and 
Gies: Amer. Journ. of Physiol., 1904, xi, p. 177. 

* Polin : Amer. Journ. of Physiol., 190S, xiii, p. 45. 

" No glycosuria occurred. Examinations were made repeatedly. 

^^ Benedict : Journ. of Biol. Chem., 1910, viii, p. 405. 

" Arnstein : Zeit. f. physiol. Chem., 1897, xxiii, p. 417. 

" Salkowski : Salkowski's Manual of physiol. chem. and path., 1904; Arch. d. 
ges. Physiol., 1897-98, Ixix, p. 268. 



I9I2] 



Jacoh Rosenhloom and William Weinherger 



125 



eighteen days, a total of 62 c.c. o£ i : 10,000 Solution was given 
intraperitoneally ; in another injection period of six days, a total of 
29 c.c. of 1 : 1000 Solution was administered. The accompanying 
tables contain the metabolic data obtained in this study. 



TABLE I. FIRST METABOLISM EXPERIMENT (jUNE I2-JULY l8, I9I2) 

A. Daily Records 
I. Fore Period. Normal Condition 



Number of the day 


I 


2 


3 


4 


5 


6 


7 


8 


9 


10 


Body weight (kilos) 


6.3 

270 
1,017 


6.33 

210 
1.025 


6.3 

270 
1,020 


6.3 

190 
1,024 


6.31 

192 
1,024 


6.3 

212 
1,021 


6.28 

275 

1,020 


6.27 

240 
1,018 


6.3 

188 
1,021 


6.33 


Urine, volume (c.c.) 


170 
1,026 


TJrine. so. er 







II. Dosage Period. Intraperitoneal Injections 



Number of the day 



Body weight (kilos) . , 
Urine, volume (c.c.) . , 

Urine, sp.gr , 

Adrenalin Solution (i 



10,000) c.c. 



6.34 
135 
1,027 

S 



6.27 

157 
1,031 

5 



6.28 

80 
1,042 
5 



6.26 
100 

1,040 
5 



6.35 
165 

1.030 
8 



6.25 

224 
1,020 



6.33 
160 

1.030 
5 



8 



6.26 

245 
1,020 



6.31 

180 
I,02S 

5 



Number of the day 



Body weight (kilos) . . 
Urine, volume (c.c.) . . 

Urine, sp. gr 

Adrenalin Solution (i 



10,000) c.c. 



10 


II 


12 


13 


14 


IS 


16 


17 


6.26 


6.33 


6.41 


6.40 


6.42 


6.37 


6.35 


6.34 


200 


105 


153 


175 


255 


230 


205 


270 


1.023 


1.037 


1.025 


1,023 


1,019 


1,017 


1,020 


1,020 


2 


3 


3 


— 


4 


— 


4 


4 



18 

6.43 
167 
1,023 

4 



III. After Period. Normal Conditions 



Number of the day 


I 


2 


3 


4 


5 


6 


7 


8 


Body weight (kilos) 

Urine, volume (c.c.) 

Urine. so. sx 


6.34 
135 
1,027 


6.40 

145 
1,020 


6.44 

200 
1,022 


6.40 

245 
1,020 


6.44 

220 
1,018 


6.41 

185 
1,022 


6.40 

214 
1,019 


6.46 

214 
1,020 











B. Analytical Totais and Daily Averages of Urinary Data for Each Period 







3 
> 


- 4) 

IS 


Nitrogen 


UreaN 


Ammonia N 


Creatin and 
Creatinin N 


Purin N 


Period 


2S 
H So 


Daily 

average, 

grams 


2 S 
E-i dt 


Daily 

average, 

grams 


H bo 


Daily 

average, 

grams 


Total, 
grams 


Daily 

average, 

gram 


15 H 


Daily 

average, 

gram 


Fore 

(10 days). 
Dosage 

(18 days). 
After 

(18 days). 


2,217 
3.206 
1.558 


221 
178 
195 


41-594 
74.225 

33-077 


4-159 
4.123 

4-134 


36.85 
63-58 
28.63 


3.685 
3-532 
3.578 


I.5II 

2.945 
I-I95 


0.1511 
0.1636 
0.1493 


1.046 
I.714 
I.IOI 


0.1046 
0.0952 
0.1376 


0.067 
0.147 
0.081 


0.0067 
0.0081 
O.OIOI 



126 



Epinephrin Effects on Metaholism 



[Sept, 



TABLE 2. SECOND METABOLISM EXPERIMENT (jULY 2I-AUGUST 9,. I912) 

A. Daily Records 
I. Fore Period. Normal Conditions 



Number of the day 

Body weight (kilos) 

Urine, volume (c.c.) 

Urine, sp.gr 



6.52 

190 
1,020 



6.53 

190 
1,023 



6.53 

230 
1,019 



6.54 

200 
1,020 



6.57 

193 
1,021 



IL Dosage Period. Intraperitoneal Injections 



Number of the day 


I 


2 


3 


4 


5 


6 


Body weight (kilos) . . . 
Urine, volume (c.c). . . 
Urine. so. er 


6.54 

ISO 
1.028 

4 


6.69 

134 
1,028 

5 


6.68 

215 
1,019 

5 


6.63 

275 
1,018 

5 


6.70 

185 
1,020 

5 


6.71 

240 
1,016 


Adrenalin Solution 
(i : 1,000) c.c 


5 



III. After Period. Normal Conditions 



Number of the day 

Body weight (kilos) 

Urine, volume (c.c.) 

Urine, sp.gr 



I 


2 


3 


4 


5 


6 


7 


8 


6.68 

245 
1,019 


6.72 

232 
1,018 


6.72 

230 
1,018 


6.76 

200 
1,019 


6.78 

202 
1,017 


6.77 

240 
1,018 


6.78 

200 
1,019 


6.83 

223 
1,017 



6.80 
23s 

1,019 



B. Analytical Totais and Daily Averages of Urinary Data for Each Period 







E 
3 

"o 

> 


>> 

> 
> " 


Nitrogen 


UreaN 


Ammonia N 


Creatin and 
Creatinin N 


Purin N 


Period 




— B E 


— « 

2 E 

rt 


r: n E 


ü E 


V 

:= t« E 


S E 
« 

H So 


CS 


u 


cd >" f^ 


Fore (5 days) . . . 
Dosage (6 days) 
After (9 days) . . 


1,003 
1,199 
2,007 


201 
199 
223 


20.997 
24.678 

34-937 


4.199 

4-II3 
3.882 


18.22 
21.46 
29-39 


3-643 

3-5766 

3.266 


0.767 
0.967 
1.499 


0.1533 
0.1611 
0.1664 


0.671 

0.7314 
1.266 


0.1342 

O.I2I9 
0.1407 


0.039 
0.039 
0.059 


0.0076 
0.006s 
0.0066 



TABLE 3. PARTITION OF THE URINARY NITROGEN 



Period 



I. Fore period 

Injection period .... 
Post injection period 

II. Fore period 

Injection period .... 
Post injection period 



Urea N, 
per Cent. 



88.5 
85-7 
86.5 

86.7 
87.0 
84.2 



Ammonia 

N, 
per Cent. 



3-64 
3-75 
3-61 

3-65 
3-92 
4.29 



Creatin and 

Creatinin N, 

per Cent. 



2.51 
2.50 
3-33 

3-20 

3-73 
3-62 



Purin N, 
per Cent. 



0.16 
0.20 
0.24 

0.19 
0.16 
0.17 



Undeter- 
mined N, 
per Cent. 



5-19 
7.86 
6.32 

6.26 

5-19 
7.72 



I9I2] Jacob Rosenhloom and William Weinherger 127 

III. CONCLUSIONS 

The results of these experiments show conclusively that intra- 
peritoneal injections of "adrenalin chlorid" Solutions were without 
appreciable effect on the proportions of nitrogen (in the forms of 
Urea, ammonia, creatin and Creatinin, purins, and undetermined sub- 
stances) in the urine of a healthy dog. 



/THE BIOCHEMICAL SOCIETY, ENGLAND 

In a previous note, which appeared in the Biochemical Bul- 
letin (i: 484), it was stated that the recently founded Bio- 
chemical Club would probably develop into a society with a Journal 
of its own. 

This is now an accomplished fact, and the Biochemical Society 
of England has been launched into being. It has been instituted 
for the purpose of facilitating intercourse between those biologists 
and chemists who are interested in problems common to both, such 
as the chemical questions connected with agriculture, brewing, animal 
and vegetable physiology and pathology, etc. Meetings are held at 
different centers throughout the country for the communication of 
papers and demonstrations. 

The Honorary Secretary is Dr. R. H. A. Flimmer, University 
College, London, W. C, from whom further Information can be 
obtained. 

The Bio-Chemical Journal, which has hitherto been under the 
editorship of Professor Moore, F.R.S., of Liverpool, will in the fu- 
ture be conducted by the Biochemical Society, and will be issued by 
the Cambridge University Press, Fetter Lane, London, E. C. 

The editors are Professor W. M. Bayliss, F.R.S., University 
College, London, W. C, and Professor A. Harden, F.R.S., Lister 
Institute, Chelsea Gardens, London, S. W. The first issue of the 
Journal under these editors is expected in January next. The price 
is £1.1.0 ($5) per volume. 

One f€els sure that our American confreres will heartily support 
the new enterprise. 

W. D. Halliburton 

King's College, London. 



128 



MEETINGS OF THE SECTION (II) ON DIETETIC 

HYGIENE AND HYGIENIC PHYSIOLOGY OF 

THE FIFTEENTH INTERNATIONAL 

CONGRESS ON HYGIENE AND 

DEMOGRAPHY, WITH AB- 

STRACTS OF SOME 

OF THE PAPERS 

Proceedings reported by THE Secretary, 
LAFAYETTE B. MENDEL 

The meeting o£ the Congress was noteworthy for the unusual 
opportunity which it afforded to American men of science to meet 
some of their foreign colleagues, particularly those from the Con- 
tinent, in a personal way. It can scarcely be said that the proceed- 
ings of the Section on Dietetic Hygiene and Hygienic Physiology 
were unique in any way; nor could they be expected to attract the 
Chief interest where so many important disciplines and conflicting 
or overlapping scientific fields were involved. The Symposium on 
the specific dynamic action of foodstuffs deserves special comment, 
however, both on account of the new views which were forcefully 
presented there for the first time, and the preeminent part played 
by all of the referees in the development of this field of study. 

I. OFFICIAL LIST OF PRESIDENTS AND VICE-PRESIDENTS 

OF THE SECTION 

Honorary Presidents : Dr. Max Rubner, Professor of Phys- 
iology and Director of the Physiological Institute, Berlin, Germany ; 
Dr. Artur Schattenfroh, Professor of Hygiene in the Univer- 
sity of Vienna, Austria ; Dr. Axel Holst, Professor of Hygiene, 
University of Christiania, Norway ; Dr. A. B. Macallum, Profes- 
sor of Biochemistry, University of Toronto, Canada. 

President: Dr. Russell H. Chittenden, Professor of Phys- 
iological Chemistry, Sheffield Scientific School of Yale University, 
New Haven, Conn. 

129 



130 Biochcmical Proceedings, Hygienic Congrcss [Sept. 

Vice-presidents : Dr. Graham Lusk, Professor of Physiology, 
Cornell University Medical College, New York City; Dr. David L. 
Edsall, Professor of Clinical Medicine, Harvard Medical School, 
Boston, Mass. 

II. OFFICIAL PROGRAMM 

All the meetings of the section were held on Sept. 23 to Sept. 27, 
inclusive, in Washington, D. C, at the new National Museum, 
Room 376. 

1. Monday afternoon, September 23. The physiological 

SIGNIFICANCE OF SOME SUBSTANCES USED IN THE PRESERVATION OF 

FOOD : Dr. John H. Long, professor of chemistry, Northwestern Uni- 
versity Medical School, Chicago, 111. (page 132) ; Dr. Artur Schat- 
tenfroh, professor of hygiene in the University of Vienna, Austria. 

2. Tuesday morning, September 24. The specific dynamic 
ACTiON OF FOODSTUFFS : Dr. Max Ruhner, professor of physiology 
and director of the Physiological Institute, Berlin, Germany. {A) 
The work of digestion and specific dynamic action : Dr. N. Zuntz, 
Direktor des tierphysiologischen Laboratoriums der landwirtschaft- 
lichen Hochschule, Berlin, Germany {presented by Prof. F. G. Bene- 
dict). — (5) The influence of the Ingestion of food upon metabolism: 
Dr. Francis G. Benedict, director of the Nütrition Laboratory of 
the Carnegie Institution of Washington, Boston, Mass. (page 134). 
— (C) The influence of foodstuffs and their cleavage products upon 
heat production : Dr. Graham Lusk, professor of physiology, Cornell 
University Medical College, New York City (page 135). 

3. Tuesday afternoon, September 24. Nutrition and 
GROWTH. (A) An anatomical analysis of growth : Dr. Henry H. 
Donaldson, The Wistar Institute of Anatomy, Philadelphia, Pa. — 
(B) Nutrition of the embryo: Dr. John R. Murlin, assistant pro- 
fessor of physiology, Cornell University Medical College, New York 
City. — (C) The nutrition and growth of bone: Dr. Francis H. Mc- 
Crudden, chemist at the Hospital of the Rockefeiler Institute for 
Medical Research, New York City (page 137). — {D) The role of 
proteins in growth : Dr. Lofayette B. Mendel, professor of physi- 
ological chemistry, Shefiield Scientific School of Yale University, 
New Haven, Conn. (page 138). — (E) The influence of the quantity 

* Abstracts of the papers appear on the pages indicated by the numerals in 
parenthesis. 



I9I2] Lafayette B. Mendel 131 

and quality of food upon the growing organism : Dr. Hans Aron, 
director of the scientific laboratory of the University Children's 
Clinic, Breslau, Germany {presented by Prof. Lafayette B. Men- 
del). — (F) Direct calorimetry of infants, with a comparison of the 
results obtained by this and other methods: Dr. John Howland, 
Professor of pediatrics, Johns Hopkins University, Baltimore, Md. 
(page 139). 

4. Wednesday morning, September 25. The röle of in- 

ORGANIC SUBSTANCES IN THE NUTRITION OF MAN. {A) The antag- 

onistic action of salts : Dr. Jacques Loeb, head of the department of 
experimental biology, Rocke feller Institute for Medical Research, 
New York City. — (B) The distribution of soluble salts in living 
cells and the forces Controlling it: Dr. Archibald B. Macalliim, pro- 
fessor of biochemistry, University of Toronto, Canada (page 140). 
— (C) The röle which common salt and water assume in the nutri- 
tion of man : Dr. Hermann Strauss, professor of clinical medicine, 
University of Berlin, Germany (page 141). 

5. Thursday morning, September 26. Practical dietetics, 
{A) Cost and nutritive value of f oods : Dr. C. F. Langworthy, ex- 
pert in charge of nutrition investigations, U. S. Department of 
Agriculture, Washington, D. C. — {B) The influence of the prepara- 
tion of food on its nutritive value : Dr. Max Riibner, professor of 
physiology and director of the Physiological Institute, Berlin, 
Germany. — (C) The choice of foods, with regard to disease: Dr. 
Carl von Noorden, professor of internal medicine and director of 
the First Medical Clinic, Vienna, Austria (page 143). — {D) Diet 
in relation to disease, chiefly in relation to some forms of partial un- 
derfeeding (beriberi and scurvy) : Dr. Axel Holst, professor of hy- 
giene, University of Christiania, Norway. — {E) Diet and metabo- 
lism in fever : Dr. Warren Coleman, Cornell University Medical 
College, New York City (page 145). 

6. Thursday afternoon, September 26. Ventilation in its 
HYGiENic ASPECTS, {A) Organic matter in the expired air: Dr. 
Milton J. Rosenmi, professor of preventive medicine, Harvard Med- 
ical School, Boston, Mass. — (B) A consideration of the unknown fac- 
tors in the ill-effects of bad Ventilation: Dr. Yandell Henderson, 
professor of physiology, Yale Medical School, New Haven, Conn. 
(page 146). — (C) The hygienic physiology of work in compressed 



132 Biochcmical Proceedings, Hygienic Congress [Sept. 

air: Dr. J. J. R. Macleod, professor of physiology, Western Re- 
serve Medical School, Cleveland, Ohio (page 147). 

7. Friday morning, September 27. The hygienic physiol- 
ogy OF EXERCISE. {A) The influence of exercise on the nervous 
System : Dr. Leon Asher, a. o. professor of physiology, Bern, Swit- 
zerland {presented by Prof. L. B. Mendel). — {B) The influence of 
exercise on the heart : Dr. R. Taif McKenzie, director of physical 
education, University of Pennsylvania, Philadelphia, Pa. — (C) 
Certain aspects of the influence of muscular exercise upon the respi- 
ratory System : Dr. Theodore Hough, professor of physiology, Uni- 
versity of Virginia, Charlottesville, Va. (page 148). — {D) Physical 
training in the United States Naval Service: Dr. J. A. Murphy, 
surgeon, U. S. N., U. S. Naval Academy, Annapolis, Md. 

Additional papers. The prevention of arteriosclerosis and 
heart disease in otherwise healthy individuals past middle life: Dr. 
Louis F. Bishop, New York City, — Tuberculosis and metabolism: 
Dr. Diesing, chief physician, Recreation and Convalescent Home, 
Gross-Hansdorf, Hamburg, Germany. — On the nature and impor- 
tance of the diet as the most important factor of causal therapy in 
severe diseases of the stomach and intestines, in nervous and mental 
diseases, and in disorders of the circulation and of the metabolism : 
Dr. W. Plönies, Hanover, Germany. — Public baths: Dr. Simon 
Baruch, president, American Association for Promoting Hygiene 
and Public Baths, New York City. — The significance of hydrother- 
apy for hygiene, therapeutics and medical Instruction: Prof. Dr. 
L. Brieger, Hydrotherapeutische Universitäts-Anstalt, Berlin, Ger- 
many. — The importance of the nutritive salts for healthy and sick 
people : Dr. R. Peters, Hanover, Germany. 

III. ABSTRACTS OF SOME OF THE PAPERS = 

The physiological significance of some substances used in the 

preservation of food 

JOHN H. long 

This paper dealt with the action on the human organism of a 
number of substances employed as food preservatives, or otherwise, 
in the preparation of food. 

* Reprinted f rom the official pamphlet containing " abstracts of papers to be 
read at the congress," Sept. 23-28, 1912 (pp. 11-28). 



igi2] Lafayette B. Mendel 133 

Something of the history of food preservatives was recited, and 
it was shown that a considerable number of substances are added to 
food largely because of their preservative properties, rather than 
because of flavors they may impart. Some of the so-called " na- 
tural " preservatives come under this head. Modern conditions of 
living and modern scientific advances have called for the introduc- 
tion of more efficient substances, the so-called " chemical " or " arti- 
ficial " preservatives. Many of these substances have been con- 
demned, and perhaps properly, but frequently the condemnation is 
solely on the ground of their origin. This basis of condemnation 
has no justification in fact, as all preservatives are as truly chem- 
ical as are those of recent introduction made by industrial processes. 
The active principles in cloves, cinnamon, allspice, etc., are true 
chemical Compounds, and in their action on the body and final dis- 
position are much like benzoic acid, now made largely by laboratory 
processes. 

A number of important investigations on the physiological action 
of sodium benzoate have been carried out in the last few years, and 
the results of these were discussed. The effects of large and small 
amounts of benzoic acid are known, and it has been clearly shown 
that the use of the small quantities employed in the ordinary pro- 
tection of the condimental foods is quite unobjectionable. Such 
small amounts are normally disposed of in the human body without 
ill effects. 

The use of copper salts in coloring vegetables was next discussed. 
There is an enormous literature on the subject, especially from 
France and Germany, where copper has long been used in the can- 
ning Industries. Several commissions have pronounced in favor of 
permitting the use of copper salts, although others have opposed it. 
But all authorities have come to agree that the toxicity of these 
salts is much less than was at one time assumed. This toxicity 
depends somewhat on the combinations in which the salts are in- 
gested. The effects of copper as used in young peas or string beans 
are far less marked than are those of its inorganic salts. It is, there- 
fore, not quite justifiable to draw conclusions as to the behavior of 
copper from experiments with copper sulfate alone. If only very 
young and fresh vegetables, with plenty of chlorophyl, were treated 



134 Biochemical Proccedings, Hygienic Congress [Sept. 

with copper, and if the amount were strictly limited, there might be 
but little fault foiind. But with older vegetables the combination 
is far less stable and the effects approach those of the inorganic 
salts. The amounts of copper taken up by the liver and other 
Organs from inorganic salts may be considerable, and such absorp- 
tion cannot be held free from danger. The use of these salts serves 
no real good purpose and should be condemned. 

The paper touched also on the employment of sulphurous oxide 
and sulphites in certain food Industries. 

The influence of the ingestion of food upon metabolism 

FRANCIS G. BENEDICT 

Three interpretations of the increase in metabolism following 
the ingestion of food are current : first, the theory in which the 
mechanical work of the digestive processes plays the most prom- 
inent röle; second, the less sharply defined theory in which the con- 
ception of the development of free heat unavailable to the cells is the 
dominant note, and, finally, the opinion expressed by Friedrich 
Müller, that there is absorbed out of the food certain substances 
w'hich are carried by the blood to the cells and there stimulate the 
cells to a greater metabolic activity. The evidence used for the 
evaluation of these views in this paper is based almost exclusively 
upon experiments made upon men in our laboratory. 

It was found that although the ingestion of sodium sulfate 
produced a powerful peristalsis, no measurable increase in the me- 
tabolism as measured by the oxygen consumption was noticed. 
Similarly, the ingestion of large amounts of agar-agar produced 
very voluminous, bulky stools, but did not increase metabolism 
measurably. As subsidiary evidence, in unpublished experiments 
on dogs with deficient pancreatic secretion, it was found that al- 
though the decreased assimilation of protein and fat resulted in 
large, bulky, fatty stools, there was not an increase in the carbon 
dioxide production. The evidence points strongly to the fact that 
the ingestion of meat by depancreatized dogs, accompanied as it is 
with large, voluminous, bulky stools, results in absolutely a smaller 
increase in metabolism than is experienced with the ingestion of 
meat by normal dogs. 



I9I2] Lafayette B. Mendel 135 

Both of these pieces of evidence, therefore, can be taken as 
strongly contrary to the work of digestion as an explanation of any 
considerable proportion of the increased metabolism generally noted 
after the ingestion of food. 

Experiments both on dogs and on men show that foUowing the 
ingestion of food there is an increased muscle tonus as indicated by 
the pulse rate, and frequently by the respiration rate, showing that 
the animal is living on a higher metaboHc plane than formerly. 
The increased heat is thus a product of cell action, and the question 
as to its economic value acquires a new significance. A man asleep, 
with lowest heat production, is of little value to the world ; awake, 
with no external muscular activity, he has increased internal activity 
and is capable of intellectnal life. 

The ingestion of protein alone stimulates metabolism with the 
possibility of some differences in the kinds of protein. Carbohy- 
drates show rapid effects, not so great as protein, and different car- 
bohydrates give different results. The metabolism 12 hours after 
the last meal of a carbohydrate-free, fat-rich diet, with moderate 
amounts of protein, is much greater than the metabolism under sim- 
ilar time-conditions after a mixed diet with the same amount of 
protein. Diabetics with varying degrees of intensity of the disease 
show marked differences in the total metabolism 12 hours after the 
last meal. A high acidosis is coincident with a high metabolism. 

A carbohydrate-free, fat-rich diet, eaten by a normal individual, 
is accompanied by the presence of an acidosis and an increased 
metabolism. The evidence suggests that coincidental with what 
is commonly termed a " State of acidosis " there is present in the 
blood a substance or substances, probably of an acid nature, that 
stimulate the cells to a greater metabolism. 



ö' 



The influenae of foodstuffs and their cleavage products upon 

heat production 

GRAHAM LUSK 

If meat in large quantity be given to a dog, the heat production 
rises in the second hour almost to its maximum, reaches its maxi- 
mum in the third hour and continues at this level through the tenth 



13Ö Biochemical Proceedings, Hygienic Congress [Sept. 

hour when it beglns to fall. In one instance the heat production 
during a morning hour was 22.3 calories, and after the ingestion of 
1,200 grams of meat it had risen in the second hour to 36 calories, 
reaching 40 calories in the third hour at which level it remained 
through the tenth hour, after which it gradually feil to 25 calories 
in the twenty-first hour. During the second hour the nitrogen elim- 
ination was one third the maximal nitrogen Output as evenly main- 
tained between the third and tenth hours. The second hour also 
showed that the calculated non-protein respiratory quotient ranged 
between 90 and 99, which indicated that that part of the metabolism 
which was not due to protein, as calculated from urinary nitrogen, 
originated largely from carbohydrate. During the later hours, the 
increased heat production is proportional to the nitrogen in the urine. 
During a period of 15 hours, protein carbon was retained in the or- 
ganism and when the oxygen absorption as computed on the basis 
of such retention in the form of dextrose is compared with the 
actual oxygen absorption, the two agree within 0.9 per cent., 
whereas computed on the basis of carbon retained as fat, there is a 
discrepancy of 10 per cent. between the calculated and actual value. 
Administration of 50 grams of dextrose in 150 c.c. of water to a 
dog causes a rise of heat production from 16.2 to 20 calories, at 
which level it is maintained during the second, third and fourth 
hours, falling nearly to the basal level in the fifth hour. The skin 
temperature rises to a greater extent than the rectal temperature. 
The absorption from the intestine is completed in the fourth hour. 
The urine is scanty until the fourth hour when 100 c.c. are suddenly 
eliminated. The sugar content of the blood in per cent. rises in 
the first hour but becomes normal after that. After the first hour 
the percentage of hemoglobin in the blood falls but returns to nor- 
mal subsequent to the fourth hour. Hence, after sugar ingestion, 
osmotic phenomena cause an increased volume of blood. When the 
absorption is complete, the glycogenic function removes the dex- 
trose from the blood, and the blood returns to its normal composi- 
tion through the elimination of water by the kidney. Water alone 
or a Solution of salt or of urea have no effect on the metabolism, 
hence the increase in metabolism is probably due to the increased 
number of molecules of dextrose carried to the cells and not to 



I9I2] Lafayette B. Mendel 137 

changes due to osmosis. Liebig's extract of beef is without influ- 
ence on the metabolism. Fifty grams of olive oil cause a consider- 
able increase in heat production. GlycocoU causes a very great in- 
crease in heat production, alanin also acts powerfully, leucin and ty- 
rosin less so and glutamic acid not at all. 

It is concluded that the heat production may be increased by 
increasing the quantity of sugar and fat reaching the cells, or it may 
be increased through the direct Stimulation of the cells by amino- 
acids, notably glycocoll and alanin. 

Nutrition and bone growth 

FRANCIS H. MCCRUDDEN 

The question of the nature of bone metabolism in health and 
disease is one that until recently can hardly be said to have been 
attacked experimentally. The pathologists, with one or two excep- 
tions, have generally considered bone as a dead tissue not undergo- 
ing metabolism, once it is laid down. This opinion has, of course, 
colored their views regarding the nature of the process in various 
bone diseases. In osteomalacia, for example, a disease in which 
there is a decrease in the mineral content of the bone, it has been 
supposed that the process is due to the action of an acid which dis- 
solves out the mineral constituents. 

Numerous investigations, chemical, histological and clinical, dur- 
ing the last few years have shown that these views regarding the 
nature of the process in osteomalacia and the nature of normal bone 
metabolism cannot be correct. Bone, like the other tissues, under- 
goes metabolism throughout life. Old bone is continuously being 
resorbed and new bone laid down. If the new bone laid down is 
not qualitatively of the right composition, the result may be rickets, 
osteomalacia, osteoporosis, or Osteitis deformans, depending on the 
age of the patient and other factors. The bones act as a störe of 
lime salts to be called on in time of need just as the subcutaneous 
tissue acts as a störe of fat and the liver as a störe of glycogen; and 
a flux of calcium from the bones started by a growing fetus, a 
hardening callus, metastatic bone formation, etc., may, under cer- 
tain circumstances, lead to decalcification enough to result in osteo- 



138 Biochemical Proceedings, Hygienic Congress [Sept. 

malacia and similar conditions. An important factor is the degree 
to which overproduction takes place, a factor involved also in im- 
munity and, in fact, in all tissue repair. 

Other disturbances which may be said to involve quantitative dis- 
turbance in bone growth, — the rate of growth, — rather than dis- 
turbances in the qualitative character of the bone produced, are the 
various types of dwarfism. In some of these the failure to grow 
seems to depend on an absence of the " growing tendency " on the 
part of the bones; in others, some disturbance in the supply of 
lime salts available for bone growth seems to be at fault. 

The role of proteins in growth 

LAFAYETTE B. MENDEL 

Some of the views held in the past regarding the interrelation of 
the food supply and growth are no longer tenable. Growth has 
often been associated in a causal way with the relative abundance of 
protein in diet. The parallelism between the protein content of the 
milk of various species and rate of growth may, in the familiär cases 
be an example of correlation rather than of causation. Recent in- 
vestigations have shown that the assumed association of growth 
with high protein intake is not confirmed by the evidence at band. 

Growth is a function of the cells. This inherent capacity ap- 
parently cannot be exaggerated by feeding ; but growth can be held 
in abeyance by various conditions. These include inadequacy of the 
food supply in respect to both quantity and quality of the nutrients. 
Attention must be directed to the chemical as well as the energetic 
aspects of the problems involved. In the past physiologists have 
largely disregarded the relative values of the individual members of 
different groups of food substances in nutrition, owing to an igno- 
rance of the chemical characteristics of the individuals. 

In considering the uses of protein in the organism, the distinc- 
tion between the requirement for maintenance and that for growth 
must be clearly kept in mind. The development of a successful 
method of investigation by Osborne and Mendel has made it easy to 
approach some of the problems experimentally. The method was 
explained in detail. Normal rate of growth has been induced in 



I9I2] Lafayette B. Mendel i39 

rats with dietaries containing various Single purified proteins. But 
not all proteins suffice to promote growth under otherwise favorable 
conditions. Some suffice for maintenance without growth, whereby 
a prolonged period of stunting, or suppression of growth, can be in- 
duced ; still other proteins are alone insufficient for the maintenance 
requirement. 

The capacity to grow is not lost even after comparatively long 
periods of dwarfing and a subsequent normal unimpaired rate of 
growth may be attained with a suitable protein dietary. Aside 
from the apparent nutritive inequalities of the different proteins, 
other incidental findings, such as the synthetic features in growth, 
and diverse questions raised thereby, present a multitude of view- 
points which may serve to direct further research in this field. 

Direct calorimetry of infants, with a comparison of the results 
obtained by this and other methods 

JOHN HOWLAND 

A discussion of the various methods for determining the ex- 
change of energy in infants with their advantages and disadvantages 
and their limitations. The results of direct calorimetry obtained 
with four children by means of a modified Atwater-Rosa-Benedict 
calorimeter. The carbon dioxide excretion, oxygen consumption 
and heat production of two essentially normal children. The effect 
of the Ingestion of food, and especially of an excess of protein. The 
effect of eighteen hours' fasting. The heat production and respira- 
tory exchange of two extremely emaciated children. 

Difficulty of comparing results with those obtained by other 
methods on account of the different conditions under which the 
experiments have been conducted, the different Information that has 
been supplied and also on account of the unsatis facto ry formulas for 
determining the surface areas of infants, which give errors of 20 
per Cent, and more. A new, more accurate and simple formula for 
determining this. 



140 Biochcmical Proceedings, Hygienic Congress [Sept. 

The distribution of soluble salts in living cells and the force^ 

Controlling it 

ARCHIBALD B, MACALLUM 

1. The distribution of salts in living matter is held to be due to 
the forces that make the distribution of salts uniform in an ordinary 
Solution. These forces are the same as those which determine the 
distribution of the molecules of a gas in an enclosed space. Conse- 
quently, in a living cell the salts are supposed to be uniformly dis- 
tributed throughout the fluid of the cytoplasm, that is, the osmotic 
force, or the pressure exercised by the molecules and ions through- 
out the fluid, is due to this uniform distribution of the solute 
throughout the System. The quantity of a soluble salt present in 
living matter is, therefore, a measure of the osmotic pressure therein 
and hence exchange between the salts within and those without 
would, in all cases, if the cell membrane were permeable, develop 
so as to ad just the pressure equally within and without the cells. 

2. This conception leaves wholly out of account the action of 
surface tension. Every particle of the colloid of which living mat- 
ter is composed presents to the fluid in which it is suspended an In- 
terface where the surface tension of the fluid is lower than such 
a fluid has at its free surface. In consequence the Gibbs-Thomson 
principle comes into Operation and there results a condensation of 
the molecules and ions of the solutes on the Interfaces of all par- 
ticles. As the united interfacial surfaces must, in relation to the 
total volume of the Solution or fluid, present a very great area, a 
very large proportion of each of the solutes must be so Condensed, 
and the general concentration is, accordingly, greatly reduced or 
brought to the vanishing point. This would reduce the osmotic 
pressure due to such solutes to a very low value or even to nil. 

3. The degree to which concentration on surfaces or on In- 
terfaces obtains depends on the degree of diminution of the tension 
of the fluid at an Interface, but it also depends on the nature of the 
solute, for the concentration in case of certain salts greatly exceeds 
the value demanded by the Gibbs equation, while in other salts the 
ascertained value approximates the theoretical value. 

4. Such surface condensation of the salts of living matter can. 



I9I2] Lafayette B. Mendel 141 

in a number of cases, be demonstrated microchemically. In the case 
of potassium salts this Is especially feasible. They are in this way 
found Condensed in interfaces inside of living cells, and also on sur- 
faces in tissues, i. e., on the external surfaces of nerve cells, of renal, 
pancreatic and salivary tubules, and thereby relations are established 
which determine the processes of excretion and secretion of these 
salts. In such cases the potassium salts in the tissue fluids elsewhere 
than at the surfaces or interfaces are scarcely detectible microchem- 
ically. 

5. Surface tension is, therefore, an all-important factor in de- 
termining the distribution of salts in living matter. 

The role which common salt and water assume in the nutrition 

of man 

HERMANN STRAUSS 

Common salt plays an important part as a regulator of the 
osmotic processes in the human organism, whereby the latter with 
greatest tenacity holds fast the percentage concentration of its fluids. 
Man can get along on relatively small quantities ( jE^ gm. ) of " salt 
required by the tissues." But the majority of civilized men con- 
sume much greater quantities of common salt, and the principal 
quantity taken in food plays the part of a " seasoning salt." There- 
fore, the reduction, in the diet, of common salt has its limits, since dis- 
turbances may ensue f rom too great a reduction. Where the supply 
is too abundant, the excess is excreted. As a result of reduction of 
ingested common salt, a diminution in the secretion of gastric juice 
has been noted in dogs. In diseases of the stomach in men, it has 
been proposed, in the case of lack of hydrochloric acid in the gastric 
juice, to introduce copious quantities of common salt ; in the case of 
increase in the secretion of gastric juice, to decrease the quantity 
of common salt in the food. But, in practice, with such a proce- 
dure, it has been possible to obtain only inconstant results. 

I myself have pointed out, that an excretory insufficiency of the 
renal function may be traced to a retention of common salt. 
Through the retention of water, this condition favors the develop- 
ment of dropsy, since the principal amount of the retained salt finds 



142 Biochemical Proceedings, Hygienic Congress [Sept. 

lodgement in the organism in the form of a " seroretention," while 
only a small part is deposited in the form of a " historetention." In 
consideration of these established opinions, for a decade, I have 
recommended a limitation of the supply of salt in the food, and a 
medicinal Stimulation of salt-elimination, in the prevention and 
treatment of hydronephrosis. 

The Situation, with regard to uncomplicated diseases of the heart 
(as well as incipient compensation disturbances in subjects of heart 
disease) is different from that in cases of parenchymatous nephritis. 
Also, in inflammatory discharges, and in ascites resulting from cir- 
rhosis of the liver, the circumstances are otherwise. In these con- 
ditions, the results of a deprivation of chlorin are very inconstant. 
For alimentation in diabetes insipidus, there have been established 
certain correct requirements similar to those laid down for parenchy- 
matous nephritics with an inclination toward dropsy. The signifi- 
cance of a limitation of salt as a means of lessening the thirst, in 
all cases in which there is a question of a decrease of fluid in the 
aliment, is now more highly appreciated than formerly. 

The question of dry retention of chlorin is now not wholly clear. 
At present, exact investigations as to the salt-content of the skin are 
lacking. Also, the relation of salt-retention to the development of 
uremia has not yet been fully explained. I should be inclined at 
this time to State only that, ccoteris parihus, uremia occurs more 
readily in the nephritic organism which is poor in water, than in 
one where water is abundant, and I may also State that, in the vom- 
ited matter of uremics, an extraordinary quantity of common salt 
is found. 

Through recent researches, a marked relationship between bro- 
min and chlorin has also been brought to light. Bromid poisoning 
may be success fully treated by means of an abundant supply of 
common salt. 

In complete deprivation of salt, and, likewise, in thorough lim- 
itation of fluids, an increase in the disintegration of protein may be 
noted. On the other band, an increase in the combustion of fat 
cannot be shown. As a rule, salt-equilibrium is restored in 24-48 
hours. On the contrary, following a previously sharp decrease in 
the supply of salt, it requires several days for the restoration of salt- 



1912] Lafayette B. Mendel 143 

equilibrium ; and, in extreme retention of salt, increased elimination 
may be checked for many days. 

It cannot be denied that many healthy persons consume too great 
quantities of common salt. Moderate amounts are not injurious. 
A certain quantity of salt, as seasoning, is permissible for civilized 
individuals accustomed to substances which stimulate the sense of 
taste. 

The choice of foods, with regard to disease 

CARL VON NOORDEN 

The discussion pertained to the lessons dietetlc therapy holds for 
US in its connection with various diseases and disease groups, and 
to the foods that are serviceable or a hindrance to the attainment of 
the end desired. Only the major groups of food substances, such 
as proteins, fats, carbohydrates, spices and salts were considered. 

1. Obesity. Principle : Decrease in the caloric value of the food. 
This is best attained through a decrease or total exclusion of the 
supply of fat. Carbohydrates, where relatively plentiful in the diet, 
should be barred out. In anti-fat treatments, the amount of con- 
tained protein should, where possible, amount to not less than 100 
grams. The supply of water must be curtailed only if the obesity is 
accompanied by disturbances of the circulation. 

2. Forced alimentation. Principle : Increase of the caloric sup- 
ply over the diet for maintenance. Theoretically, it is all the same, 
whether the center of gravity rests upon a large supply of carbohy- 
drates or fat. In reality, 250 gm. of carbohydrate is seldom ex- 
ceeded, because most carbohydrate foods possess a very great vol- 
ume. The supply of protein may not ordinarily be increased be- 
yond 100-120 gm. By means of these, approximately 1,300 calo- 
ries, no satisfactory alimentative results may be obtained. The 
practical results depend always upon the increase in the supply of fat. 
In most cases, the latter may be increased to 250 or 300 gm. daily, 
and then increases in weight of about 2 kilos per week may be 
gained. 

3. Gout and uric acid diatheses. Principle : Decrease in animal 
foods; eventually total exclusion of the same. It is useful, in the 



144 Biochemical Proceedings, Hygienic Congress [Sept. 

case of every gouty patient, to undertake a separate "test of tolera- 
tion," and, from the result of this test, to establish the patient's diet 

4. Diabetes mellitus. Principle : Avoidance of such foods as in- 
cite the organ of sugar-production, the liver cells, to increased for- 
mation of sugar. Every undue Stimulation of the sugar-forming 
organ has, as its result, not only an immediate lavish production of 
sugar, but also increases f or the f uture its morbid excitability ; while 
systematic care of the organ renders its recovery possible. There- 
fore, decrease, and, under some circumstances, total exclusion of the 
carbohydrates. Moreover, decrease of protein substances. It is 
essential, in every case of diabetes, to exactly determine under what 
dietetic regime and manner of living the least amount of superfluous 
sugar is formed. That order of diet is best under which the patient 
continues free from superfluous sugar. 

5. Feverish diseases and morbus Basedowii. Principle: In both 
these diseased conditions there occurs an abnormal increase in caloric 
production. Simultaneously ensues a heightened sensibility in re- 
lation to the specific dynamic influence of proteins. In order to 
limit as far as possible the caloric production and the loss in weight, 
practical empiricism and theory likewise call for a scanty protein 
supply, while weight is gained by an ample provision of carbohy- 
drates. 

6. Diseases of the digestive organs. Principle : A food supply 
which is sufficiently nourishing, while imposing as little tax as pos- 
sible upon the diseased organs. A discussion of the injurious effect 
of certain combinations of foods. Report upon enterotoxic neuritis. 
Attack by means of protracted pure milk diet. 

7. Kidney diseases. Principle : As much rest as possible for the 
kidneys. The amount of the intake of those nutrient media whose 
products of metabolism leave the body through the kidneys, 
should be reduced. The proteins conie first in this regard. But 
this limitation should not be carried too far, since patients with 
chronic kidney diseases become anemic and weak if strict curtail- 
ment of the proteins is too long continued. Many spices irritate 
the kidneys, and indulgence therein must be limited ; the same is the 
case with regard to alcohol. Common salt and water severely tax 
the kidneys. 



I9I2] Lafayette B. Mendel i45 

Final words : Warning against schematic employment of dietary 
precepts. An effort must be made, on the one band, to hold fast to 
the basic rules of nutrition-therapy, but, on the other, to duly take 
into account the individuality of the patient. 

Diet and metabolism in fever 

WARREN COLEMAN 

Empiricism has been a signal failure as a basis for the fever diet. 
Through studies of metabolism the Solution of this problem appears 
to be at band. With diets containing large amounts of carbohy- 
drate it is possible to bring typhoid fever patients into nitrogen 
equilibrium, or nearly so. The apparently excessive quantities of 
food required for the purpose are almost completely absorbed. 

The heat-production in typhoid fever, as determined by indirect 
calorimetry, averages about 35 calories per kilogram at absolute 
rest. Diets furnishing only sufficient energy to cover the heat-pro- 
duction do not Protect the body against nitrogen or weight loss. 
The explanation of this discrepancy has not yet been found. 

Respiratory quotients in typhoid fever below 0.65 to 0.70 appear 
to be due to errors of technique. The lowest quotient we obtained 
in the fasting state, during the febrile period, was 0.70. During 
the same period, patients on a füll diet gave quotients varying from 
0.75 to 0.95 at short intervals after food. The quotient rises dur- 
ing the later stages of the fever and reaches i.o to 1.15 early in con- 
valescence. During a relapse, a quotient of 1.04 was obtained while 
the patient had a temperature of 102° F. (37.7° C). 

The oxygen consumption during the febrile stage varies between 
4 and 6 c.c. per kilogram a minute. Compared with the amount 
used in the fasting stage, the oxygen consumption is not greatly 
increased by the quantity of food administered. 

The body burns carbohydrate by preference during fever as long 
as it is available. As indicated by a falling quotient, from 100 to 
120 grams of lactose is, for the most part, consumed, or deposited in 
the glycogen depöts, after 4 to 5 hours. The optimum amount of 
carbohydrate in the fever must be determined for each patient in- 
dividually, but is always large. The optimum amounts of fat and 
protein have not yet been determined. 



146 Biochcmical Proceedings, Hygienic Congress [Sept. 

A consideration of the unknown factors in the ill-effects of bad 

Ventilation 

YANDELL HENDERSON 

The facts regarding Ventilation present an extraordinary contra- 
diction. Fresh air, sunlight and dry cool climates exert a decidedly 
beneficial effect upon health. Ill-ventilated dwellings decrease 
vitality. In some persons under certain conditions even a few min- 
utes in a crowded room may produce acute ill-effects. As to how 
these effects are produced physiology has up to the present time 
afforded no satisfactory explanation. The evidence is almost en- 
tirely negative. The ill-effects of bad Ventilation can not be due 
to lack of oxygen. It is probable that they are not due in any con- 
siderable degree to excess of CO2. The idea that they are due to 
some poisonous substance contained in the expired air has in recent 
years been regarded as untenable. Recently this conception has been 
revived in a novel form by the brilliant work of Rosenau. Even 
Rosenau's investigations do not appear, however, to afford the Solu- 
tion of this problem. The recent investigation of Hill in England 
and of Flügge and his pupils in Germany makes it highly probable 
that the effects of fresh or vitiated air are brought about not by a 
direct action upon the lungs but indirectly through the skin. It 
appears probable that the temperature and moisture of the air sur- 
rounding the body are the essential Clements. 

According to the explanation to be suggested in this paper the 
condition of the skin exerts a potent influence upon the lungs. This 
may be in part a vaso-motor reflex acting upon the pulmonary circu- 
lation. More probably it is a chemical or hormone influence upon 
certain pulmonary processes. The evidence accumulated during 
recent years indicates that the lungs are not mere passive organs 
through which gases diffuse as through non-living membranes. 
The investigations of Bohr, of Haidane and his co-workers and of 
the recent Pikes Peak expedition all tend to indicate that the lungs 
are the seat of vital activities of great importance to health. Thus 
under certain conditions the lungs secrete oxygen into the blood, and 
it appears that considerable oxidation may take place in the blood 
during its passage through the pulmonary vessels. The evidence 



I9I2] Lafayette B. Mendel 147 

available, although still far from complete, suggests that these pul- 
monary activities are indirectly but powerfiilly influenced through 
conditions affecting the skin, and that it is in this manner that Venti- 
lation influences health. 

The hygienic physiology of work in compressed air 

J. J. R. MACLEOD 

Although it is now a well-established fact that the Symptoms of 
Caisson disease and diver's palsy are due to the sudden liberation o£ 
bubbles of nitrogen in the blood and tissue fluids, on account of too 
sudden decompression, there are several peculiarities regarding the 
conditions which influence the safety of decompression about which 
there is still a certain degree of uncertainty. This is the case more 
particularly with regard to : ( i ) Whether the decompression should 
be uniform or in stages; (2) how long it should take in proportion 
to the time of the shift and the pressure employed; (3) the degree 
to which the breathing of oxygen increases the safety of decompres- 
sion. Although, as insisted on by Haidane and others, it is no 
doubt the case that " the absolute air pressure can always be reduced 
to half the absolute pressure at which the tissues are saturated 
without risk " yet, in practice, it has not been found that the method 
is in any way superior to that of gradual decompression. 

The time that should be taken in decompression depends on the 
length of the shift in the caisson, because the Saturation of the re- 
moter parts of the body with nitrogen continues for a long time 
after this has been attained in the blood and the more accessible tis- 
sues. Tables indicating what time should be allowed have been 
prepared by Haidane and by Japp. 

The advantages of breathing oxygen are not only that it accel- 
erates the diffusion of nitrogen out of the lungs and, therefore, out 
of the blood; but, if Symptoms have already appeared, it supplies 
enough oxygen to keep life going when the circulation is danger- 
ously obstructed by nitrogen bubbles. In using oxygen at higher 
pressures, its toxic action must however be kept in mind. 

Recompression, either by placing the caisson worker in a pres- 
sure Chamber or by having the diver descend again to a certain depth 



148 Biochemical Proceedings, Hygienic Congress [Sept. 

whenever the first Symptoms appear, is by far the most efficient treat- 
ment, as both experiment and the experience of engineers testify. 

On account of the heat and the high relative humidity of com- 
pressed air, the worker in a caisson is under conditions which tend to 
lower his efficiency. Not only this, but his appetite is likely to suf- 
fer and his general condition after some time to deteriorate so that 
he becomes liable to infections if not to caisson disease itself. The 
Caissons should therefore be well ventilated and the wet bulb ther- 
mometer kept as low as possible. Means for doing this were dis- 
ciissed. In the choice of men for caisson work attention should be 
paid to age, body weight and f atness and while engaged in the work 
the men should be kept in good training. 

Certain aspects of the influenae of muscular exercise upon the 

respiratory System 

THEODORE HOUGH 

Muscular activity increases the respiratory exchange from three- 
to tenfold, thus making demands on the System comparable only 
with those of the more severe forms of dyspnea. In meeting the 
respiratory needs of the tissues there are secondary effects of hy- 
gienic importance, such as the increased aspiration of the thora'x 
upon the return of venous blood to the heart and also upon the flow 
of lymph in the larger lymphatics ; this increased lymph flow is feit 
in the interstitial Spaces of every organ in the body, thus favorably 
influencing the environment of every cell. 

The introduction with more vigorous exercise of physiological 
strain makes it important to inquire into the exact condition of the 
organism revealed by the accompanying respiratory phenomena. 

Of the conditions known to increase the work of the respiratory 
Center Geppert and Zuntz have excluded, as exciting causes of the in- 
creased breathing movements of muscular activity, afferent Impulses 
from the working muscles and deficiency of oxygen in the arterial 
blood; their work also shows a decrease of the total {i. c, free and 
combined) carbon dioxid of the arterial blood; it does not establish 
a diminished tension of this gas in the respiratory center, and it is 
possible (Haidane) that there may be increased tension of this gas in 



igi2] Lafayette B. Mendel I49 

the Center along with a fall of the total amount in the blood. No 
direct determinations of the condition of the blood in this respect 
have been made. 

Determinations of the alveolar tensions of oxygen and carbon 
dioxid during and at varying periods after muscular activity show 
that with increasing intensity of work there is first a rise of CO2 
tension, then a fall to and below normal. In the latter case the CO2 
tension sinks still further after the cessation of the exercise and 
may remain subnormal for over half an hour or even an hour. 

Review of the evidence as a whole leads to the conclusion that 
during more vigorous exercise the main cause of the increase of 
breathing movements is some catabolite (other than CO2) of the 
working muscle. During moderate exercise the increase of CO2 
tension of the blood is probably an adequate explanation ; the respi- 
ratory condition of the organism would thus differ not only in 
degree but also in kind with moderate and with more vigorous 
exercise. 

Theory that the distress which is relieved by " second wind " is 
due to excessive CO2 tension not well established by the evidence at 
band, but worthy of further study. Bearing upon this question is 
the effect of previous Inhalation of oxygen in lessening the distress 
of maximal effort. 

Should not the measurement of respiratory power in physical 
examination be extended so as to include not only the anatomic 
features of ehest expansion and vital capacity, but also the ability of 
the respiratory System to meet successfully the conditions of the 
more vigorous forms of muscular activity? 

Yale University, 

New Haven, Conn. 



MEETINGS OF THE SECTION ON BIOCHEM- 

ISTRY, INCLUDING PHARMACOLOGY (VIII, D), 

OF THE EIGHTH INTERNATIONAL CON- 

GRESS OF APPLIED CHEMISTRY 

Proceedings reported by THE Secretary, 
JOHN A. MANDEL 

I. LIST OF OFFICERS OF THE BIOCHEMICAL SECTION 

President, John J. Abel; Vice President, William J. Gies; Secre- 
tary, John A. Mandel; Executive Committee: Reid Hunt, Thomas 
B. Osborne and the officers. 

IL SECTIONAL PROGRAM^ 

The meetings of the Section were held on September 6 to Sep- 
tember 12, inclusive, at Columbia University, in Room 301 of 
Havemeyer Hall. The morning sessions were opened at 10 o'clock 
and the afternoon sessions at i o'clock. 

Friday morning, September 6. In the chair: The Vice 
President. Julius Stoklasa: Ueber die photochemische Synthese 
der Kohlenhydrate unter Einwirkung der ultravioletten Strahlen. — 
L. Marchlezvski: The present State of our knowledge of the relation- 
ship of the chemistry of the blood coloring matter and Chlorophyll. — 
L. Marchlezvski and C. A. Jacobson: On the quality of Chlorophyll 
and the variable ratio of the two constituents, and on methods for 
determining this ratio. — *Guido M. Piccinini: II manganese del 
punto di vista delle funzioni enzimatiche. — ^ Jules Wolff: Sur la re- 
sistance de la Peroxydase ä l'ammoniaque et sur son activation par 
contact avec l'alcali. — *Jides Wolff: Sur une nouvelle fonction du 
catalyseur dit " Peroxydase" et sur le transformation biochemique 
de l'orcine en orceine (page 53). — Walter Jones: Some new phases 

*The asterisks indicate the papers which were actually presented. Some of 
the titles were received after the official program had been printed and are 
included here informally. Abstracts of most of the papers were published in 
Volume 19 of the preliminary report of the proceedings of the Congress. 

150 



1912] ■ John A. Mandel 151 

of the nuclein fermentation. — Carl Voegtlin: Further studies in 
biologic oxidations. — "^Walter R. Bloor: Fatty acid esters of 
glucose. — C. C. Guthrie: A comparative study of the action of Solu- 
tions on the preservation of the vitality of tissues. — *R. Delaimay 
and O. Bailly: Les pepsines fluides etude du sediment qui se produit 
dans certaines d'entre elles. — William J. Gies: Modified collodion 
membranes, with demonstrations. 

Saturday morning, September 7. In the chair: Prof. 
Mauthner, of Vienna. ^Gabriel Bertrand and F. Medigreceanu: 
Sur la presence normale du manganese chez les animaux. — *M, Lin- 
det: Sur les elements mineraux de la caseine du lait. — *P. Malvezin: 
La question de l'acide sulfureux dans les vins blancs. — *Z. Mimu- 
roto: Ueber das Vorkommen von Adenin und Asparaginsäure in 
Maulbeerblättern. — U. Suzuki and 5". Matsunaga: Ueber das Vor- 
kommen von Nikotinsäure (m-Pyridinkarbonsäure) in der Reis- 
kleie. — Zozo Sakaguchi: Ueber den Fettgehalt des normalen und 
pathologischen Harns. — W. N. Berg: Effect of sodium chlorid and 
cold storage upon the activities of proteolytic enzymes. — *Thonias 
B. Aldrich: The iodine content of the small, the medium and the 
large thyroid glands of beef, sheep and hogs. — *Lezvis W. Fetzer: 
The chemical changes taking place in milk under pathological con- 
ditions. — *Ma.Y Kahn: A study of the chemistry of renal calculi, 

Monday morning, September 9. In the chair : The Pres- 
ident. *M. Nicloux: Moyen de caracteriser de petites quantites 
d'alcool methylique dans le sang et dans les tissus. — Zennoshin 
Hatta: Zur Kritik der Zuckerbestimmungsmethode nach Ivar Bang. 
— Munemichi Taniura: Zur Prüfung der Kumagawa-Sutoschen Fett- 
bestimmungsmethode in Bezug auf die Oxydation der fettsäuren 
und unverseif baren Substanzen im Verlaufe des Verfahrens. — Yuji 
Sueyoski: Eine neue approximative Eiweissbestimmungsmethode bei 
Albuminurie. — "^ Franz Herles: Schnelles Verfahren zur Bestim- 
mung der Harnsäure im Harn. — W. Worth Haie and Atherton 
Seidell: The comparative estimation of epinephrin in suprarenal 
glands and in its Solutions, physiologically and by color tests. — 
Lyman B. Stookey: The Cammidge reaction. — ^Herbert H. Bunzel: 
Oxidase determinations. — /. P. Atkinson: On the Separation of cer- 
tain alkaloids from nerve tissue. — *W. H. Schidtz and Atherton 



152 Biochcmical Procccdings, Chemical Congrcss [Sept. 

Scidcll: The determination of thymol in dog feces. — "^Shiro Tashiro: 
A new apparatus for the detection and estimation of exceedingly 
minute quantities of carbon dioxide in biological materials. — "^Shiro 
Tashiro: Carbon dioxide production in the nerve fibre during an 
excitation. Its apphcation for detection of life in protoplasm. — 
*F. Klein: Die selenige Säure — ihr Verhalten gegen Eiweiss und 
tierische Haut. — ^Gabriel Bertrand and H. Agulhon: Sur la presence 
normale du bore chez les animaux. 

Monday afternoon, September 9. In the chair : The Pres- 
ident. "^ Felix Ehrlich: Ueber einige chemische Reaktionen der 
Mikroorganismen und ihre Bedeutung für chemische und biologische 
Probleme. — ^Gilbert T. Morgan and E. Ashley Cooper: The influ- 
ence of the chemical Constitution of certain organic hydroxyl and 
aminic derivatives on their germicidal power. — Takaoki Sasaki: 
Ueber den Abbau einiger Polypeptide durch Bakterien. II. Unter- 
suchungen mit nicht verflüssigenden Bakterien. — Naganiichi Shi- 
bata: Zur Frage der Fettzersetzung durch einige Saprophyten. — 
*A. Trillat: Influence des impuretes gazeuses de l'air sur la vitalite 
des microbes. — *M. Javillier: Influence exercee par le zinc sur Vas- 
pergilliis niger au point de vue de l'utilisative par la plante. — *Car/ 
L. Aisberg and O. F. Black: Biochemical and toxicological studies 
upon Penicillium stolonifernm. 

Tuesday morning, September 10. In the chair : The Pres- 
ident. *M. Mane: Relations de la plante avec les Clements fertili- 
sants; loi du minimum et loi des rapports physiologiques. — *M. 
Gerber: Etüde comparee des pressures des Vamanite phalloide et de 
Vamadoiivier. — *i?. Dubois: Sur l'atmolyse et sur l'atmolyseur. — 
*/?. Dubois: Recherches sur les vacuolides de la purpurase. — *i?. 
Dubois: La biophotogenese reduite a une action zymasique. — 
Oszvald Schreiner: The physiological röle of organic constituents in 
plant metabolism. — *C F. Langworthy : The study of problems of 
vegetable physiology by means of the respiration calorimeter; 
a progress report. — ^Hozvard S. Reed: The enzyme activities in- 
volved in certain plant diseases. — ^Ernest D. Clark: Origin and sig- 
nificance of starch. — William J. Gies: Studies of diffusion, with 
demonstrations. 

Tuesday afternoon, September 10. In the chair.- The 



jgi2] John A. Mandel 153 

President. *P. Carles: Les phosphates et le son de froment dans 
ralimentation animale. — *P. Carles: Entretien du tissu dentaire par 
une alimentation appropriee. — Minoru Maeda: Versuche über die 
Ausnutzung von " Konnyak " (einer japanischen Speise). — *PanlE. 
Hozve and Philip B. Hawk: The utilization of various protein 
foods by man after repeated fasting. — *L. F. F oster and Philip B. 
Hawk: A study of the utilization of ingested food when undermas- 
ticated ("bolted") and overmasticated ("fletcherized"). — S. P. 
Beebe: The influence of the thyroid on the excretion of ammonia. — 
^Andrew Hunter and Maurice H. Givens: Purin metaboHsm in the 
monkey. — William Salant and /. B. Rieger: The influence of alcohol 
on protein metabolism. — Jacob Rosenbloom: Chemical and pharma- 
cological studies of human duodenal contents. 

Wednesday morning, September 11. In the chair: The 
President. *M, Sauton: Nutrition minerale du bacille tubercu- 
leux. — * Walter J. Dilling: Charts of spectra representing visible 
and invisible bands of various hemoglobin derivatives, with explana- 
tory booklet. — *G. O. Higley: Some notes on the form of the curve 
of carbon dioxide excretion resulting from muscular work follow- 
ing forced breathing. — *G. 0. Higley: The influence of barometric 
pressure on the carbon dioxide excretion in man. — *Joseph L. Miller 
and Dean D. I.ezvis: Physiological action of the various anatomical 
components of the hypophysis. — Isaac Levin: Immunity and specific 
therapy in experimental Cancer. — Lafayette B. Mendel: The physio- 
logical behavior of lipoid-soluble dyes. — *5. B. Crohn: Experiences 
with duodenal and stool ferments in health and disease. — Hertnan 
M. Adler: Experimental production of lesions resembling pellagra. 
— William J. des: Studies of edema, with demonstrations. 

Wednesday afternoon, September 11. In the chair: The 
President. George W. Crile: Neuro-cytological changes resulting 
from the administration of certain drugs. — *G. A. Menge: Some 
new Compounds of the cholin type. — Reid Hunt: Physiological 
action of some new Compounds of the cholin type. — Arthur S. Loev- 
enhart: Further observations on the action of oxidizing substances. 
— W. H. Schultz: Pharmacological action of proteins and some of 
their derivatives. — */^. H. Schultz and Atherton Seidell: Subcu- 
taneous absorption of thymol from oils. 



154 Biochemical Proceedings, Chemical Congress [Sept. 

Thursday morning, September 12. In the chair: The 
President. */. M. Fortesciie-Brickdale: The arylarsonates : their 
pharmacology considered from the experimental and practical stand- 
points. — *//. A. D. Jowett, V. F. L. Pyman and V. F. G. P. Remfry: 
The relation between chemical Constitution and physiological action, 
as exempHfied by the glyoxahnes, isoquinolines and acid amides. — 
Walther Straub: Pharmakologische Bedeutung der Zellmembranen. 
— Charles Baskerville: Inhalation anesthetics. — "^Thomas B. Al- 
drich: On feeding young white rats the anterior and posterior parts 
of the pituitary gland. — /. A. E. Eyster: The relation of calcium to 
the inhibitory mechanism of the heart. — Clyde Brooks: On the 
action of alcohol on the circulation. 

Thursday afternoon, September 12. In the chair: The 
President. Giovanni Bufalini: Reazioni caratteristiche del veleno 
del rospo (Bufo vulgaris.) — Giovanni Bufalini: Meccanismo dell' 
azione narcotici del chloridrine. — *E. Fonrneau and V. K. Ochslin: 
Chlorure de l'acide dichloroarsinobenzoique ; ethers des acides benz- 
arsineux et benzarsinique. — *C R. Marshall: The pharmacological 
action of brom-strychnins. — *C R. Marshall: The influence of hy- 
droxyl and carboxyl groups on the pharmacological action of nitric 
esters. — Isaac Adler: Studies on chronic adrenalin, lead and nicotin 
intoxications. — *Ivo Novi: II calcio e il magnesio del cervello in varie 
condizioni fiziologiche e farmacologiche. — *L. Launoy: Action de 
quelques amines, en particulier du chlorure et de l'hydrate de tetra- 
methylammonium sur la secretion pancreatique. — *R. Delaunay and 
O. Bailly: Examen critique des conditions d'essai des pancreatines 
medicinales. 

III. ATTENDANCE 

Among the many in attendance at one or more sectional meet- 
ings, the Secretary noted the presence of the colleagues named 
below : John J. Abel, T. B. Aldrich, C. L. Aisberg, J. P. Atkinson, 
W. N. Berg, G. Bertrand (Paris), Samuel Bookman, Harold C. 
Bradley, H. H. Bunzel, Ernest D. Clark, F. C. Cook, F. Ehrlich 
(Breslau), Frank R. Eider, B. G. Feinberg, Lewis W. Fetzer, M. S. 
Fine, Harry L. Fisher, A. O. Gettler, Wm. J. Gies, A. J. Goldfarb, 
R. A. Gortner, Isidor Greenwald, M. L. Hamlin, G. A. Hanford, 



1912] John A. Mandel i55 

Robert A. Hatcher, Philip B. Hawk, G. O. Higley, B. Horowitz, E. 
M. Houghton, Paul E. Howe, Reid Hunt, Max Kahn, F. Klein, P. 
A. Kober, W. M. Kraus, P. A. Levene, Isaac Levin, Alfred P. Loth- 
rop, Wm. G. Lyle, John A. Mandel, Samuel Matthews, T. Mauthner 
(Vienna), F. Medigreceanu, G. M. Meyer, Jas. L, Miller, G. T. 
Morgan (Dublin), Max Morse, Victor C. Myers, W. A. Pearson, 
F. B. Power (London), Howard S. Reed, A. I. Ringer, C. J. Rob- 
inson, Anton R. Rose, Jacob Rosenbloom, William Salant, Emily 
C. Seaman, Atherton Seidell, B. Setlik (Prague), H. C. Sherman, 
Torald Sollman, Matthew Steel, M. X. Sullivan, Shiro Tashiro, 
Rodney H. True, H, Vieth (Ludwigshavn), Charles Weisman, 
Louis E. Wise. 

University and Bellevue Hospital Medical College, 
New York City. 



SIXTH SCIENTIFIC MEETING OF THE COLUMBIA 

UNIVERSITY BIOCHEMICAL ASSOCIATION, AT 

THE COLLEGE OF PHYSICIANS AND SUR- 

GEONS, NEW YORK, JUNE 3, 1912 

PrOCEEDINGS RePORTED BY THE Secretary, 

ALFRED P. LOTHROP 

The sixth scientific session (third "annual" meeting) of the 
Columbia University Biochemical Association was held at the Co- 
lumbia Medical School on the evening of June 3, 191 2. The execu- 
tive proceedings of this session were published on pages 570-573 of 
Volume I of the Biochemical Bulletin (June number). 

The scientific proceedings consisted of research Communications 
by members of the Association. Abstracts of the papers are pre- 
sented here (pages 158-187) in two groups: (I) Abstracts of papers 
on research by non-resident members^ and (II) abstracts of 
papers from the Columbia Biochemical Department and affiliated 
laboratories. The appended summary will faciliate reference tp 
the abstracts (1-44). 

A SUMMARY OF THE NAMES OF THE AUTHORS AND OF THE 
TITLES OF THE SUCCEEDING ABSTRACTS 

I Allan C. Eustis. On the physio- 

WiLLiAM N. Berg. The physico- logical action of some of the amins 

Chemical basis of striated-muscle produced by intestinal putrefaction. 

contraction. (i) (s) 

William N. Berg, with L. A. Rogers, Allan C. Eustis. Solubilities and 

C. R. Potteiger and B. J. Davis. action of ^-imidazolylethylamin and 

Factors influencing the flavors of the relation to asthma and anaphy- 

storage butter. (2) laxis. (6) 

Isabel Bevier, for Anna W. Wil- A. J. Goldfarb. On the production of 

liams. A study of ropy bread. (3) grafted multiple embryos. (7) 

Allan C. Eustis. On the toxicity of Max Morse. Non-toxicity of inor- 

guinea pig urine and its relation to ganic colloid Solutions upon protozoa. 

anaphylaxis. .(4) (8) 

^ Members of the Association who were not officially connected with the 
Columbia biochemical department when the research was conducted. 

156 



I9I2] 



Alfred P. Lothrop 



157 



Max Morse, for L. B. Ripley. Larvae 
of Lepidoptera obtained with sul- 
furic acid. (9) 

Anton Richard Rose. A study of 
the metabolism and physiological 
effects of certain phosphorus Com- 
pounds in milk cows. (10) 

II 

David Alperin. Contribution to the 
knowledge of nucleoprotein metab- 
olism, with special reference to uri- 
colysis and to the properties of 
uricase. (11) 

George D. Beal and George A. Geiger. 
The comparative diffusibility of vari- 
ous pigments in different solvents. 
(12) 

Stanley R. Benedict. The occur- 
rence and estimation of Creatinin in 
urine. (13) 

Louis E. Bisch. An endeavor to pre- 
pare Phrenosin from protagon. (14) 

Louis E. Bisch. Mucoid-silver prod- 
ucts. (15) 

Sidney Born. Protein-copper prod- 
ucts. (16) 

J. J. Bronfenbrenner and Hideyo 
NoGUCHi. A biochemical study of 
the phenomena known as comple- 
ment Splitting. (17) 

Ernest D. Clark. Notes on the 
chemical natura of Lloyd's " tannin 
mass." (18) 

Walter H. Eddy. A study of some 
protein Compounds. (19) 

Walter H. Eddy. The preparation of 
thymus histon. (20) 

Frank R. Elder and William J. Gies. 
The influence of proteases on the 
swelling of collagen and fibrin par- 
ticles in alkalin and acid media con- 
taining a biological electrolyte. 
(21) 

William J. Gies. A convenient form 
of apparatus for demonstrations of 
osmotic pressure exerted by lipins. 
(22) 



William J. Gies. Some interesting 
properties of thymol. (23) 

William J. Gies. A convenient 
method of preparing starch that 
swells rapidly in water. (24) 

R. f. Hare. A study of the carbo- 
hydrates of the prickly pear and its 
fruits. (25) 

Henry H. Janeway and William H. 
Welker. The relation of acapnia 
to shock. (26) 

Max Kahn. Biochemical studies of 
sulfocyanate. (27) 

Max Kahn. The chemical Constitu- 
tion of renal calculi. (28) 

Max Kahn and Jacob Rosenbloom. 
The colloidal nitrogen in urine from 
a dog with a tumor of the breast. 
(29) 

Max Kahn and Frederic G. Good- 
ridge. A non-protein, colloidal, ni- 
trogenous substance in milk. (30) 

John L. Kantor. A biochemical test 
for free acid, with a review of the 
methods for estimating the various 
factors in gastric acidity. (31) 

Marguerite T. Lee. A study of modi- 
fications of the biuret reagent. (32) 

Alfred P. Lothrop. A chemical study 
of salivary mucin. (33) 

C. A. Mathewson. A study of some 
of the more important biochemical 
tests. (34) 

Jacob Rosenbloom. A quantitative 
study of the lipins of bile obtained 
from a patient with a biliary fistula. 

(35) 

Jacob Rosenbloom and William 
Weinberger. Effects of intraperi- 
toneal injectionsof epinephrin on the 
partition of nitrogen in urine from 
a dog. (36) 

Oscar M. Schloss. A case of allergy 
to common foods. (37) 

Carl A. Schwarze. The comparative 
enzyme content of green and varie- 
gated leaves of Tradescantia. (38) 



158 



Proceedings CGlumhia Biochcmical Association [Sept. 



Emily C. SE.^MAN. Biochemical 
studies of beryllium sulfate. (39) 

Clayton S. Smith. Chemical changes 
in fish during long periods of cold 
storage (40) 

William Weinberger. An attempt to 
sharpen the end point in Benedict's 
method for the quantitative deter- 
mination of sugar in urine. (41) 

William H. Welker. Diffusibility of 



protein through rubber merhbranes, 
with a note on the disintegration of 
collodion membranes by common 
ethyl ether and other solvents. (42) 
Charles Weisman. A further study 
of the Bardach test for protein. 

(43) 
Harold E. Woodward. A study of the 
surface tension of dog blood-serum 
by the drop-weight method. (44) 



I. ABSTRACTS OF PAPERS ON RESEARCH BY NON- 
RESIDENT MEMBERS^ 

1. The physico-chemical basis of striated-muscle contrac- 
tion. William N. Berg. {Washington, D. C). Part I was 
published in the June issue of the Biochemical Bulletin; part 
II is presentcd in this issue} 

2. Factors influencing the flavors of storage butter. Wil- 
liam N. Berg, with L. A. Rogers, C. R. Potteiger, and B, J. 
Davis. {Dairy Division Research Laboratories, Bureau of Aninial 
Industry, Washington, D. C.) The official government bulletin on 
this subject is in press. 

3. A study of ropy bread. Isabel Bevier, for Anna W. 
Williams. {Research Laboratory, Department of Household Sci- 
ence, University of Illinois, Urbana, III.) Published in fidl in the 
June issue'^ of the Biochemical Bulletin. 

4. On the toxicity of guinea pig urine and its relation to 
anaphylaxis. Allan C. Eustis. {Laboratory of Clinical Med- 
iane, Department of Nutrition, Tidane University, New Orleans, 
La.) The nrine of guinea pigs, fed on Kohlrabi or cabbage, con- 
tains a great excess of indican, which readily oxidizes to indigo. 
Such urine also contains excess of putrefactive amins. Tests for 
/8-imidazolylethylamin, as well as efforts to isolate it, have been 
negative. Experiments on fifteen guinea pigs weighing 300 grams 
each, with different specimens of guinea pig urine, indicate that 1.5 
c.c. constitutes a lethal dose when injected intravenously. In these 

^ Members of the Association who were not officially connected with the 
Columbia biochemical department when the research was conducted. 
^Berg: Biochemical Bulletin, 1912, i, pp. 535-7; ii, pp. loi-io. 
* Williams : Biochemical Bulletin, 1912, i, pp. 529-534. 



I9I2] Alfred P. Lothrop i59 

animals, the Symptoms were identical with those observed after in- 
jections of /8-imidazolylethylamin. There was no delay in coagu- 
lation of the blood, but there was marked lowering of blood pres- 
sure and lowering of body temperature. 

After intravenous injections of filtered giiinea pig iirine into 
three dogs, Symptoms resembUng those of anaphylactic shock were 
exhibited, but the fall in blood pressure was not constant as it is 
after anaphylactic shock, and there was no delay in the coagulation 
of the blood. There is evidently some relation between the occur- 
rence of putrefactive amins and anaphylactic shock, but the writer's 
results do not bear out Pfeiffer's opinion regarding that relation. 

5. On the physiological action of some of the amins produced 
by intestinal putref action. Allan C. Eustis. (Laboratory of 
Clinical Medicine, Department of Nutrition, Tulane University, 
New Orleans, La.) Putrescin (tetramethylendiamin) and cada- 
verin (pentamethylendiamin), in doses as small as o.i mg., are in- 
stantly fatal when injected intravenously into guinea pigs. Non- 
fatal doses produce marked lowering of blood pressure, dyspnea 
from edem.a of the lungs, salivation and prostration. The pulse is 
quickened. 

Phenylethylamin is immediately fatal to a guinea pig weighing 
300 gm. when 0.05 gram is injected intravenously; 0.03 gram was 
fatal in two minutes when injected intravenously into a 300 gram 
guinea pig, with immediate prostration and paralysis of the respira- 
tory center ; 0.02 gm. produced a distinct chill in a 300 gram guinea 
pig, followed by prostration but with ultimate recovery. 

ß-imidasolylethylamin, in doses of o.oi gram intravenously, 
caused death in three minutes with typical anaphylactic Symptoms, 
the animals dying in attacks of forcible inspiratory effort, the heart 
continuing to beat after the respiration had ceased. 

Parahydroxyethylamin, as well as isoamylamin, produced marked 
rise in blood pressure. 

6. Solubilities and action of /?-imidazolylethylamin and the 
relation to asthma and anaphylaxis. Allan C. Eustis. (Lab- 
oratory of Clinical Medicine, Department of Nutrition, Tiüane 
University, New Orleans, La.) I. A specimen of chemically pure 
)S-imidazolylethylamin, obtained through the courtesy of Dr. Dale 



i6o Proccedings Columbia Biochemical Association [Sept 

of the research laboratory of Burroughs, Welcome & Co., was in- 
soliible in cold Chloroform, benzene, tolnene, amyl alcohol, but 
slightly soluble in xylol, easily soluble in methyl alcohol, and soluble 
in cold carbon disulfide and hot amyl alcohol. 

Aqneous Solutions were tested with several reagents, to dis- 
cover if possible some means of detecting the presence of /?-imida- 
zolylethylamin in the tissties or blood, as follows: Bromine water, 
no precipitate, no coloration; copper sulfate, negative; potassium 
ferrocyanid, negative; Paidy's reagent, cherry red coloration; pic- 
ric acid, yellow precipitate insoluble in water, alcohol, ether, xylol 
and toluene, but which gave the positive Pauly reaction; phospho- 
tungstic acid, gray-blue precipitate, soluble in barium chloride Solu- 
tion, and in barium hydroxid Solution, which gave a positive Pauly 
reaction; sodiiim nitrite, negative; magnesium sulfate, negative; 
niercuric chlorid, negative ; gold chlorid, negative. 

Efforts to detect, by microchemical means, the presence of ß-\m\- 
dazolylethylamin in the bronchioles of guinea pigs dying from ana- 
phylactic shock, were without results. 

IL Tests of the physiological action of /J-imidazolylethylamin 
were conducted upon rabbits, guinea pigs and dogs by intravenous, 
subcutaneous and intraperitoneal injections. Intravenous injections 
of 0.5 mg. in guinea pigs caused immediate respiratory embarräss- 
ment, lowered blood pressure and diminished body heat, the animal 
dying in six minutes from suffocation due to complete occlusion of 
the bronchioles ; it being impossible to either f orce air into the lungs 
or to withdraw air, after the contraction had become complete. 
The Symptoms were typical of anaphylactic shock, and the post- 
mortem examination revealed the presence of enormous emphysema, 
the heart continuing to beat long after respiration had ceased. In 
dogs and rabbits there was also a lowering of the blood pressure 
and some respiratory embarrassment, but the occlusion of the bron- 
chioles was not as complete as in guinea pigs. 

Stibctttaneons and intraperitoneal injections were much less 
toxic and, in some instances, were entirely negative, suggesting that 
the tissues are able to utilize /?-imidazolylethylamin. 

The writer has seen many cases of asthma relieved entirely 
along dietetic lines by a " low protein " diet, and empirically has 



I9I2] 'Alfred P. Lothrop i6i 

found that red meats predispose to asthmatic attacks. )8-imidazo- 
lylethylamin is produced in the putrefactioii of histidin, and hemo- 
globin yields a large percentage of histidin on decomposition. It is 
possible, therefore, that /3-imidazolylethylamin causes asthma. Un- 
like clinical asthma, however, experimental asthma produced by 
iß-imidazolylethylamin is not relieved by injections of epinephrin 
("adrenaHn chlorid"). 

7. On the production of grafted multiple embryos. A. J, 
GoLDFARB. (Marine Biological Lahoratory, Woods Hole, Mass., 
and the Department of Natural History, College of the City of 
New York.) Grafted multiple embryos were first successfully 
produced in considerable numbers by Driesch, with the eggs of 
either of two genera of echinoderms, namely, Echiniis and Sphaere- 
chinus. Though several investigators have endeavored to repeat 
these experiments with American echinoderms they have failed 
completely. By slightly modifying the Herbst-Driesch method as 
described below, an unusually large number of grafted multiple 
embryos and larvae were produced from the eggs of Arbacia punc- 
tidata. 

After removing the fertilization membranes, the eggs were 
placed either directly into a sodium hydroxid Solution, or first 
placed in calcium-free sea water, then in an alkaline liquid of the 
following composition: 4 to 20 drops of 0.5 per cent. sodium hy- 
droxid Solution in 200 c.c. of sea water. This treatment sufficed in 
Driesch's experiments with Echinus and Sphaerechinus, giving 
rise to about 4 per cent. of agglutinated and fused embryos. 
For Arbacia eggs it was necessary to Supplement this treat- 
ment by centrifuging the eggs in tubes with very narrow bores, so 
that the eggs whose outer surfaces had previously been gelatinized 
were compressed against one another. These eggs gave rise to 
about 40 per cent. of agglutinated and fused embryos and larvae. 

The multiple embryos of Arbacia, so produced, were of the same 
general character as those described by Driesch, such as true twins, 
incomplete fusions, and complete fusions of the respective embryos. 

8. Non-toxicity of inorganic colloid Solutions upon pro- 
tozoa. Max Morse. (Boardman Laboratories, Trinity College, 
Hartford, Conn.) Colloidal platinum prepared by the Bredig 



102 Proceedings Columbia Biochemical Association [Sept. 

method, in which the house current of iio volts was reduced to 70 
volts by lamps in parallel and passed through glass-distilled water 
by means of platinum electrodes, was used as a medium in which 
cultures of Paramecium and other protozoa were permitted to rest. 
Drop-ctilture slides were also made of these cultures in hanging 
drops of the platinum black. In all cases there was no augmenta- 
tion of division-frequence or size of the organism, nor any evi- 
dence of toxicity. Attempts with a Solution of mastic in ether and 
alcohol, which gave beautiful pictures under the Dunkelf eldtheleuch- 
tung of Zeiss, were not clear in their results. The colloidal Solu- 
tion was dialyzed for seven days in a fish-bladder, which freed it 
from the ether and alcohol, leaving a colloidal mass with excellent 
brownian movement. However, there is good reason to believe that 
this is not toxic in any way on protozoa. No attempt was made to 
" ultra-filter " the colloidal Solutions, in order to study the effects of 
small and larger colloidal particles upon protozoa, because of the 
apparent indifference of the organisms to the mixed Solution. 

9. Larvae of Lepidoptera obtained with sulfuric acid. Max 
Morse, für L. B. Ripley. (Boardjnan Laboratories, Trinity Col- 
lege, Hartford, Conn.) Larvae were obtained from unfertilized 
eggs of the moth, Cecropia, by painting them with Baker's conc. 
sulfuric acid (sp. g. 1.84) for from 3 to 6 seconds and immediafely 
washing in pure water until entirely free from the acid. They were 
then left to dry and to develop. Checks were made by treating one- 
half of the batch from a given female with the acid and leaving 
the other half untouched. The females had been raised and 
isolated, from cocoons. The typical blueing of the developing eggs 
could be observed in the early stages of the eggs treated with acid 
while the control eggs remained white. The larvae emerged sev- 
eral days later in the case of the artificially fertilized eggs than in 
those normally fertilized. The percentage of errors was low. 

The larvae after emerging from the eggs were fed upon wild 
cherry, but thus far they have not been carried to the adult stage. 
This is now being tried. Petrunkevitch, Tichomorow and others 
have succeeded in obtaining larvae from silk-worm eggs by artificial 
means, but Cecropia has thus far failed to yield larvae under arti- 
ficial conditions. Short exposure and thorough washing may be the 
key to the success obtained in the present case. 



I9I2] 'Alfred P. Lothrop 163 

10. A study of the metabolism and physiological effects o£ 
certain phosphorus Compounds in milk cows. Anton Richard 
Rose. (New York Agricultural Experiment Station, Geneva, 
N. Y.) The phosphorus requirement of a cow, aside from the 
milk phosphorus, would seem from the results of this experiment 
to be about 26 mg. per kilo of body weight. When the phosphorus 
supply is less than this amount, the physiological functions are con- 
tinued at the expense of the phosphorus previously stored in the 
tissues. Storage takes place when a greater amount than that indi- 
cated above is ingested. When the ingested insoluble phosphorus 
did not exceed 14 grams per day, there was approximate regularity 
in the phosphorus elimination in the feces independent of the In- 
gestion, suggesting that all the forms of phosphorus were digested, 
with liberation of phosphates; also that the fixed phosphorus of the 
feces was entirely due to the cellular matter from the mucosa and 
the intestinal flora. The soluble organic phosphorus in the feces 
was relatively slight in quantity, even in the periods when " phytin " 
was fed in liberal amounts. The calcium phytate added to the 
washed-bran ration was not utilized as economically as the " phytin " 
of the whole bran, and the "phytin" of the partially washed bran 
also gave a lower digestion coefficient. 

The addition of "phytin" to the "low phosphorus" ration in- 
creased the potassium Output in both feces and urine. The fecal 
potassium dropped in quantity when the "phytin" was withheld, 
but the urinary potassium did not. The amount of fecal mag- 
nesium was constant through the several periods except in the 
fourth, when it seems to have been influenced by the increased in- 
take of calcium phytate. At the beginning of the experiment the 
magnesium in the urine was equal to half that in the feces, but con- 
tinually decreased until the mobile magnesium of the body had been 
largely eliminated. The calcium in the urine increased remarkably 
when the phosphorus intake decreased. In the calcium phytate 
period, the calcium increase in the feces was approximately equiva- 
lent to the calcium increase in the rations. 

In all cases the addition of organic phosphorus to the " low phos- 
phorus " ration was followed by a decrease in the milk flow, and the 
withdrawal of this phosphorus from the ration was followed by a 



164 Proceedings Coliunbia Biochemical Association [Sept. 

larger yield of milk. The percentage of fat in the milk fluctuated 
regularly with the changing amount of phosphorus ingested. The 
response was immediate, but the quantities of milk-fat bear no con- 
stant ratio to the amount of phosphorus in the rations. Aside from 
those pertaining to the fat, there were practically no changes in the 
composition of the milk, not even in the percentage of phosphorus 
in the fat-free solid matter. 

The moisture relations in the problem seem significant, though 
the intake and outgo of water could not be accurately measured in 
this experiment. The margin, after allowing for the influence of 
temperature, leads one to suspect a large retention of water in the 
last two periods. 

Up to the sixtieth day there was no outward sign of any physio- 
logical disturbance, but about that time the appetite began to wane. 
On the seventy-seventh day the milk-flow declined rapidly and 
serious trouble developed. A few days later the cow was placed 
in a box-stall and fed alfalfa, silage and vvheat bran, which caused 
all signs of malnutrition to disappear in the course of a week and 
also increased the milk-flow. 

IL ABSTRACTS OF PAPERS FROM THE COLUMBIA BIOCHEMICAL 
DEPARTMENT AND AFFILIATED LABORATORIES 

11. Contribution to the knowledge of nucleoprotein metab- 
olism, with special reference to uricolysis and to the properties 
of uricase.^ David Alperin. The author studied the relative 
efficiency of the Wiener, Rosell, Croftan, Wiener and Wiechowski, 
and Galeotti methods for the preparation of uricase, and indicated 
the properties of the products. Wiener and Wiechowski have sug- 
gested that the subcutaneous or intravenous administration of 
uricase preparations is an effective procedure for the eure of gout 
and allied diseases. The author concludes that "practical demon- 
stration of the efficiency of this method of treatment has not been 
made." 

12. The comparative diffusibility of various pigments in 
different solvents. George D. Beal and George A. Geiger. 
(Piiblishcd in füll in this issue of the Biochemical Bulletin.)^ 

"Alperin: Dissertation, Columbia University, 1912. 

'Beal and Geiger: Biochemical Bulletin, 1912, ii, pp. ydr-^. 



1912] Alfred P. Lothrop 165 

13. The occurrence and estimation of Creatinin in urine/ 

Stanley R. Benedict, l'he work contemplates a thoroiigh inves- 
tigation of the question as to whether the Jaffe reaction in urine is 
due entirely, as is usually assumed, to the form of Creatinin which 
is ordinarily isolated from urine, or whether other substances may 
not be partially responsible for the reaction. The results indicate 
that there are two (or more) forms of Creatinin in urine, both of 
which yield the Jaffe reaction and also a zinc chlorid Compound, but 
which differ from each other in certain specific properties. A 
change in the ratio between these two forms of Creatinin in the 
urine has been observed in certain abnormal conditions. The most 
marked change was noted in inanition. There is probably a third 
substance contributing to the Creatinin reaction of urine which is 
in no wise related to Creatinin, but appears to be a weak acid. The 
study is in progress. 

14. An endeavor to prepare phrenosin from protagon.^ 
Louis E. Bisch. Thudichum's method^ of isolating phrenosin has 
apparently never been reviewed. It was assumed that this method 
could be applied with success directly to protagon. The author was 
unable to do so, however. Repetitions of each of the numerous 
Steps in the process, with as much as 1450 grams of protagon at 
a time (in faithful accord with Thudichum's description), failed 
to yield sufficient material with which to complete the directions. 
It is possible that losses, which seem to have occurred at all stages of 
the process, totally consumed any phrenosin that existed in the orig- 
inal protagon. It is Dr. Gies' Intention to study this possibility 
further. 

15. Mucoid-silver products.^^ Louis E. Bisch. Mo'ist, a-cid- 
free tendomucoid, triturated with a moderate amount of moist, 
alkali-free silver oxid, yields a brown to black mixture which be- 
comes very viscid when a small volume of ammonium hydroxid So- 
lution is stirred into it. A mechanical excess of 10 per cent. 
ammonium hydroxid Solution converts the viscid mass into a brown 

^ Under the auspices of the George Crocker Special Research Fund. 

* Bisch : Dissertation (Part I), Columbia University, 1912. 

* Thudichum : A treatise on the chemical Constitution of the hrain, 1884, pp. 
136-8. 

"Bisch: Dissertation (Part II), Columbia University, 1912. 



i66 Proceedings Columbia Biochemical Association [Sept. 

to black Solution, from which free alkali and free silver can be 
removed by dialysis. The neutral Solution thus prepared appears 
to contain argent-ammonium-mucoid, which may be obtained by 
precipitation with alcohol or by direct desiccation. The aqueous 
Solutions of these products are similar to those of argyrol in many 
respects yet appear to keep indefinitely. The purified material is 
antiseptic, and retards the growth of plants, but is seemingly non- 
irritant to the Cornea or other animal tissues. Fairly large quan- 
tities fail to induce toxic effects when injected subcutaneously or 
intravenously into dogs. The product in aqueous Solution is decom- 
posed by acidification. The purified material yields about i6 per 
cent. of ash. The silver content will be given special attention in the 
near future. 

i6. Protein-copper products. Sidney Born. Concentrated 
aqueous Solutions of various indiffusible proteins, when rendered 
slightly alkalin with sodium hydroxid Solution and treated with a 
moderate quantity of copper sulfate Solution, exhibit the typical 
biuret reaction in marked degree, but the excesses of alkali and 
copper may be removed by dialysis and, as the process continues 
(although no color may appear in the diffusate), the deep "biuret 
color" slowly changes until finally a blue or green persists. The 
resultant protein-copper product was isolated by precipitation of 
such a Solution with alcohol or by its direct desiccation. The Pro- 
portion of copper in six products made from edestin, gelatin, and 
serum protein ranged from 4.2 to 6.3 per cent. Injected subcuta- 
neously into frogs, the edestin and gelatin products (1.3 c.c. of 
concentrated aqueous Solution in each case) caused death in three 
hours. The properties of the dialyzed Solutions and the products 
therefrom will be described in some detail later. 

17. A biochemical study of the phenomena known as com- 
plement Splitting. J. J. Bronfenbrenner and Hideyo Nogu- 
CHi.^^ A. It is generally accepted that complement may be split 
into a mid-piece and an end-piece. The mid-piece is thought to be 
in the globulin fraction, and the end-piece in the albumin fraction. 



11 



Bronfenbrenner: Dissertation, Columbia, 1912; Bronfenbrenner and 
Noguchi : Journal of Experimental Mediane, 1912, xv, 598-643. Most of the 
work was conducted at the Rockefeller Institute for Medical Research. 



I9I2] 'Alfred P. Lothrop 167 

The restoration of complement activity by putting together the albu- 
min and globulin fractions does not prove, however, that each 
fraction contained a part of the complement, for the albumin 
fraction can be reactivated in the absence of the globulin fraction. 

Complement-splitting as brought about by hydrochloric acid, 
carbon dioxid, and dialysis, is really an inactivation of the whole 
complement by certain acids or alkalis, either added in the free 
State to the serum, or liberated as a result of the dissociation of 
certain electrolytes. 

That the whole complement, and not a part only, is present in 
the albumin fraction of the serum can be demonstrated by the re- 
moval of the inhibitory action of the acid or alkali. This can be 
effected by the addition, not only of alkali or acid, but also of any 
amphoteric substance. When hydrochloric acid, carbon dioxid, or 
dialysis are employed to produce the phenomenon known as com- 
plement Splitting, the complement is merely inactivated, not split. 

B. Thus far, most investigators have made but little distinction 
between the Splitting phenomenon obtained by chemical interference 
and that which takes place in the biological phenomenon known as 
complement fixation. In this study we have shown that these two 
sets of phenomena exhibit certain fundamental differences and that 
the so-called complement Splitting by physical conditions leading to 
chemical interaction, or directly by chemical means, is not a real 
Splitting of the complement, but an inactivation of the active prin- 
ciple of complement through an alteration in the reaction of the 
medium caused by an excess of either anions or cations. The modi- 
fication of the reaction of the medium may cause a more or less 
definite combination of the complement with the free ions, but the 
latter can readily be removed by an appropriate number of opposite 
ions, and render the complement active once more. The fluids that 
have hitherto been regarded as containing the end-piece of com- 
plement, contain, as a matter of fact, the whole complement tem- 
porarily deprived of its activity by certain ions derived either from 
the Salt constituents of the serum itself under a modified physical 
condition (dialysis against water or dilution with water) or intro- 
duced in the form of dissociable electrolytes. 

On the other hand, the Splitting of complement in the fixation 



i68 Proccedings Columbia Biochemical Association [Sept. 

reaction seems far more complicated than that caused by the phys- 
ical or chemical procedures. The supernatant fluid from the fixa- 
tion test differs from all the other end-pieces prepared by chemical 
methods in being active upoii persensitized sheep corpuscles only 
(not upon human corpuscles). The addition of various mid-pieces, 
obtained by different methods, to sensitized sheep corpuscles does 
not render the Wassermann supernatant fluid active. It is quite 
remarkable that the persensitized sheep corpuscles are, on the other 
hand, easily attacked, not only by the supernatant fluids of fixation 
tests, but also equally well by the other end-pieces. It is not at all 
improbable that in the fixation reaction, where so many factors come 
into play, there is a most complicated physical as well as chemical 
interaction leading to such an entangled mixture of factors that a 
substance carrying one set of ions alone cannot reverse the activity 
of complement, and hence the reversion takes place only when cer- 
tain electrolytes with both ions are employed. At all events there 
seems to be no doubt that the inactivation of complement is far 
more complicated in the Wassermann reaction or the Bordet-Gen- 
gou phenomenon than in the inactivation by physical or chemical 
means. Nevertheless, no one has as yet proved conclusively that 
the supernatant fluid of a fixation test necessarily contains the end- 
piece of complement. 

i8. Notes on the chemical nature of Lloyd's " tannin 
mass." Ernest D. Clark. Chemical studies were made upon 
"tannin masses" prepared by Lloyd from the fruit of the persim- 
mon. The original material dissolved in alkalies to form a purple 
jelly-like Solution. In dilute mineral acid Solutions the "masses" 
turned bright red in color and no swelling was observed. Upon 
hydrolysis with 0.2 per cent. and 2.0 per cent. hydrochloric acid 
Solutions, cherry-red colorations were obtained. Such Solutions 
contained both tannin and phloroglucin in considerable proportions. 
The presence of phenolic substances like vanillin was also indicated. 
An insoluble gelatinous substance was removed, by filtration, from 
the hydrolyzed acid mixture and seemed to be cellulose or a related 
material. Hydrolysis with 0.5 per cent. and 5.0 per cent. sodium 
hydroxid Solutions gave thick, dark-colored liquids and large 
amounts of insoluble gelatinous residue. Alkaline hydrolysis pro- 



igi2] 'Alfred P. Lothrop 169 

duced the same kinds of materials as those that resulted from acid 
hydrolysis. The "tannin masses" seem to be combinations of 
tannin and phloroglucin associated with cellulose-like substances. 
With ferric chlorid, phloroglucin gives a dark blue product but not 
the blackish precipitate characteristic of the tannin-ferric chlorid 
reaction. Theref ore, " iron reagents " do not detect tannin in the 
presence of phloroglucin. 

19. A study of some protein Compounds. Walter H. Eddy. 
(Published in füll in this issue of the Biochemical Bulletin. )^2 

20. The preparation of thymus histon. Walter H. Eddy. 
As outlined by Bang, the properties of histon may be summarized as 
follows : Water-soluble, non-coagulable by heat, precipitated by am- 
monia in the presence of salts, precipitated from neutral Solution by 
"alkaloidal reagents," produces precipitates of several soluble pro- 
teins from their aqueous Solutions. 

The current method of preparing thymus histon, as recom- 
mended in Standard handbooks such as Abderhalden's and Oppen- 
heimer's, may be summarized as follows : Extraction of the minced 
glands with water. Precipitation of the water extract by acid or 
calcium chlorid, and extraction of this precipitate with 0.8 per cent. 
hydrochloric acid Solution. Precipitation of the hydrochloric acid 
extract with ammonium hydroxid Solution, either before or after 
removing free hydrochloric acid by dialysis. Washing the "am- 
monia precipitate " free from ammonia with alcohol and ether. 

Kossei, who discovered histon in goose blood, obtained it by 
saturating the hydrochloric acid extract with sodium chlorid. He 
alone calls attention to the anomaly noted in our experiments, viz., 
that treatment with ammonium hydroxid Solution results invariably 
in the precipitation of a substance that is practically insoluble in 
water. In a series of many preparations, extending in time over a 
period of two years and involving materials obtained from many 
calves, we have come to the conclusion that the " ammonia-pre- 
cipitation" of a hydrochloric acid Solution of thymus histon results 
invariably in a water-insoluble product. Furthermore, our 
experiments show that of two fractions of the same hydrochloric 
acid Solution, the fraction saturated with sodium chlorid invariably 

^^Eddy: Biochemical Bulletin, 1912, ii, p. 111-22. 



I/o Proceedings Columbia Biochemical Association [Sept. 

yields a prodtict which (when free from sodium chlorid) is water- 
soluble, gives all the qualitative histon tests, and contains less 
nitrogen than the "ammonia precipitate" from the other fraction; 
the " ammonia precipitate " being water-insohtble and appar- 
ently a very different stibstance. Finally, when the " sodium 
chlorid precipitate " of histon is dissolved in vvater, and the aqueous 
Solution is treated with a few drops of ammonium hydroxid Solu- 
tion, a precipitate is produced which is insoluble in water. Quanti- 
tative studies now under way show marked differences in the nitro- 
gen content of the two products. 

The results suggest that " histon " as commonly prepared is an 
adsorption product or a salt, rather than a simple protein. 

The following method is suggested as a means of obtaining 
water-soluble histon from thymus : Mince f resh thymus glands and 
extract the hash with distilled water for 24 hours (best in the cold), 
Precipitate the aqueous extract with acetic acid Solution and ex- 
tract the precipitate with 0.8 per cent. hydrochloric acid Solution 
(after Lilienfeld) ; or add sufficient calcium chlorid to the aqueous 
extract to make its content of that substance 0.2 per cent. and 
extract the precipitate with 0.8 per cent. hydrochloric acid Solution 
(after Huiskamp) ; or add sufficient hydrochloric acid to the aqueous 
extract to make its content of the acid 0.8 per cent. and let stand 
24 hours (after Kossei and Kutscher). Filter off the hydrochloric 
acid extract, and either remove the free acid or precipitate the 
histon directly by Saturation with sodium chlorid. Remove ad- 
mixed sodium chlorid by dialysis. Filter the resultant salt-free 
Solution and evaporate it to dryness at 45° C. This material, 
ground to a powder, may be heated to 105° C. without loss of 
water-solubility. 

21. The influenae of proteases on the swelling of Collagen 
and fibrin particles in alkalin and acid media containing a bio- 
logical electrolyte. Frank R. Elder and William J. Gies. 
(Published in full in the June issue of the Biochemical 
Bulletin. )^^ 

22. A convenient form of apparatus for demonstrations of 
osmotic pressure exerted by lipins. William J. Gies. The 

" Eider and Gies : Biochemical Bulletin, 1912, i, pp. 540-545. 



I9I2] 'Alfred P. Lothrop 171 

writer repeated the demonstration described on page 59.^^ Instead, 
however, of using a thin rubber bag in a muslin sheath, he employed 
a i2-inch section of ordinary bunsen-burner tubing. The rubber 
tube had been swollen to its maximum extension by immersion in 
ether for about an hour previous to its use. It was then closed at 
one end by the Insertion of a short, tightly fitting, section of a thick 
glass rod, which was fastened by a ligature. After the swollen 
tube had been filled with olive oil and a narrow glass tube about 
10 feet in length (in two sections) had been tied into the open end 
and held upright, the rubber-oil portion of the vertical tubulär appa- 
ratus was completely immersed in ether in a tall, narrow cylinder. 
The oil began to rise in the tube almost immediately, and rapidly 
proceeded upward until the liquid emerged from the open top. 

23. Some interesting properties of thymol. William J. 
GiES. During the course of recent experiments on enzymes as pos- 
sible factors in the development of edema/^ we had occasion to 
study the effect of trypsin on elastin in ammonium hydroxid Solu- 
tions containing a biological electrolyte (NaCl). To our surprise 
we not only failed to obtain the swelling results which we had pre- 
viously observed under similar conditions/^ but the elastin particles 
in use gradually became green, ultimately blue. With repeated 
shaking, the elastin particles were more deeply colored, and the 
supernatant liquid slowly became green; finally, bluish green. The 
color of the particles slowly diminished in intensity as the pigment 
accumulated in the liquid. Unlike the elastin used in the previous 
experiments, this product had been prepared about 10 years before. 
The fresh-ligament hash had been put in water and preserved there 
with considerable alcoholic thymol Solution; later, had been put in 
alcohol; ultimately, had been dried and bottled. The main supply 
of the dry elastin smelled strongly of thymol. 

Some of the above-mentioned green and blue ammoniacal liquids, 
when shaken with ether or toluene, were quickly transformed into 
purplish, then reddish mixtures. The ether layer on the quiescent 
liquid was bright red — all green and blue had disappeared from the 

" Gies : Biochemical Bulletin, 1912, ii, p. 55. 
" Eider and Gies : Ibid., 1912, i, p. 540. 
*' Tracy and Gies : Ibid., 1912, i, p. 472. 



1/2 Proceedings Columbia Biochemical Association [Sept. 

alkalin liquid imderneath, which was colorless. By spontaneous 
evaporation, the ether extract yielded a purplish-red oily product, 
vvith a pronounced thymol odor. 

When a small quantity of thymol (Kahlbaum) was mixed with 
lO per Cent, ammonium hydroxid Solution, the liquid became green- 
ish in about 2 hours ; then gradually turned blue. Alcohol appeared 
to accelerate the transformation. Shaken with ether, the blue was 
wholly removed and a beautiful, red, ether-layer obtained. Such 
ether extracts yielded, by spontaneous evaporation, a purplish-red 
oily product, which dissolved readily in ether, toluene and alcohol, 
the Solutions being bright red. In some cases the oily product be- 
came crystalline, due apparently to the presence of unchanged thy- 
mol ( ?). The red alcoholic Solution was turned deeply bluish by a 
drop of n/io sodium hydroxid Solution; the red was restored by a 
drop of w/io hydrochloric acid Solution. These transformations 
could be elicited repeatedly in the same Solution. The changes were 
so sharp that the material may prove to be a valuable indicator for 
use in the titration of alcoholic liquids. Concentrated alcoholic Solu- 
tions yielded reddish white precipitates when they were diluted with 
water — a ready means of isolating the substance. The reddish 
white precipitate dissolved promptly in alcohol, ether and toluene, 
and formed a red Solution in each case. 

An excess of thymol, added to a green or blue ammoniacal Solu- 
tion in its original condition, completely changed the green or blue 
to red, and wholly dissolved the red material, behaving, in this 
respect, like toluene and ether. 

These phenomena did not appear to be due to impurities in the 
thymol. A general survey of thymol literature has not revealed 
the explanation of these results, although certain inferences are sug- 
gested by several color reactions of thymol. 

The chemical nature of the colored substances derived from 
thymol in these preliminary experiments, the possible Utility of the 
products — their probable antiseptic, pharmacologic and other rela- 
tionships, suggest numerous interesting biochemical inquiries which 
will be undertaken in the near future. 

24. A convenient method of preparing starch that swells 
rapidly in water. William J. Gies. For the purpose of study- 



I9I2] ^Alfred P. Lothrop 173 

ing the effects of amylases on the power of starch to imbibe water 
(prior to hydrolytic cleavage), the writer prepared markedly hydro- 
phylic starch in the following way: A very thick starch paste was 
speedily prepared by rapidly pouring a thicl<: potato-starch Suspen- 
sion through musHn into boiling water while the latter was being 
vigorously stirred. The vessel containing the paste was inimersed 
in ice water immediately after the last portion of starch Suspension 
had been added. By constant stirring of both liquids, and by the 
maintenance of a low external temperature, the paste was speedily 
cooled,^'^ when it was poured into, and thoroughly stirred in, a large 
excess of 95 per cent. alcohol. After the Sedimentation of the prod- 
uct, and the decantation of the alcoholic liquid, the snow white ma- 
terial was treated with fresh portions of alcohol until its viscidity 
disappeared and it became firmly granulär. After several washings 
with ether, to remove alcohol, the product was rapidly freed from 
ether in a current of air from an electric fan. Although somewhat 
hygroscopic, the material formed hard, snow-white masses which 
could be granulated easily in an ordinary pulverizer. 

Placed in water, the particles swell very rapidly into bloated 
glassy forms. " Starch paste " may be made almost instantly from 
the product. The powder can easily be freed from its soluble car- 
bohydrate impurities by dialysis. The material promises to be of 
special Service in many connections. Mr. Nathan Rosenthal has 
undertaken a study of the effects of amylases on the swelling of 
material of this kind in various anti-hydrophylic media, such as 
dilute alcohol. 

25. A study of the carbohydrates of the prickly pear and its 
fruits. R. F. Hare.^^ The difficulties encountered in the practical 
laboratory Separation of the sugars from the mineral matter, muci- 
lages, gums and dextrinoid substances have been numerous, and the 
Operations time-consuming. Many attempts to obtain the sugars 
free and in crystalline form have usually resulted unsuccessfully; 
so that it became necessary to make the individual tests not on the 
sugar crystals, but on the syrups previously purified as much as pos- 
sible by different methods. 

" The Operations were conducted rapidly in order to prevent undue hydrol- 
ysis. It is probable that satisfactory results can be obtained by pouring the 
hot paste directly into alcohol. 

"Hare: Dissertation, Columbia University, 191 1. 



174 Proceedings Columbia Biochemical Association [Sept. 

The Juice of the ripe fruit contains 1.57 per cent. of pentosans 
and only traces of galacfan. After precipitation with lead acetate, 
the Juice gave the anihne acetate reaction for pentose, but none for 
galactose. The presence of fructose and gliicose in considerable 
amounts was quite definitely estabhshed by several reactions char- 
acteristic of these sugars. 

The dried mucilage of the prickly pear, when separated by pre- 
cipitation with alcohol from a two per cent. Solution, contained 15 
per cent. of galactan, 31 per cent. of pentosan and 12 per cent. of 
ash. The mucilage in the aqueous extracts could not be separated 
completely from cell fragments, starch, crystals of calcium Oxalate 
and other solid particles that caused opalescence and turbidity. A 
dilute Solution containing 1.5 per cent. of solid matter, rendered 
fairly clear by repeated filtration through silk, had no effect on 
polarized light. This was true of all the Solutions of mucilage ob- 
tained in this work, both before and after subjecting them to acid 
hydrolysis. Harley^^ reports having found a specific rotation of 
+ 38° for Opuntia mucilage, but places little confidence in his own 
results, since the reading was made on a very dilute opalescent Solu- 
tion and calculated from an observed rotation of + 6 minutes. 
Hydrolysis of the mucilage by digestion for several hours with 1,25 
per cent. sulfuric acid Solution produced a sugar that had properties 
similar to arabinose. When its osazone was formed, oily globules 
rose to the surface. The precipitate was darker than glucosazone, 
readily soluble in hot water and melted at about 160° C. 

A 95 per cent. alcoholic extract of the dried stems, previously 
treated with ether, contained a sugar with specific rotations made on 
three separate Solutions of — 6.6°, — 8.25°, and — 7.1°. The 
osazone produced from this sugar had properties similar to those of 
glucosazone. These results indicate the presence of glucose and 
fructose^ in this extract. 

A 60 per cent. alcoholic extract of the dried stems contained 
a suhstance apparently intermediate in character hetween mucilage 
and sugars. It did not reduce Fehling Solution before hydrolysis, 
but was very readily hydrolyzed by dilute acid Solutions. Alcohol 
stronger than 60 per cent. reprecipitated this material as a flocculent 

" Harley : Journal de Pharmacie, iii, pp. 6-193. 



I9I2] 'Alfred P. Lothrop 17 S 

mass, quite different in appearance and properties from the precipi- 
tate of the mucilage obtained with alcohol. The precipitate was 
readily soluble in water, but its Solution was not mucilaginous. 
When hydrolyzed, it gave a plus rotation to polarized Hght. 

The coloring matter can be concentrated and made into a mar- 
ketable product, of value for coloring certain foods, by first remov- 
ing mucilages and gums with alcohol, and precipitating the pigment 
from the filtrate with acetone. The pigment is evidently a gluco- 
side. When separated from the juice with alcohol and acetone, and 
then precipitated with lead acetate, the coloring matter liberated by 
sulfuric acid gave a glucose-like sugar on hydrolysis. The lead salt 
produced by precipitating the purified pigment with lead acetate con- 
tains 61.42 per cent. of lead. 

26. The relation of acapnia to shock.^*^ Henry H. Janeway 
AND William H. Welker. Henderson has published a number 
of papers on the relation of acapnia to shock. He maintains that a 
diminution of the normal amount of carbon dioxide in the blood to 
a sufficient degree, and maintained for a sufficient length of time, 
produces an irreparable disturbance of the normal balance of osmotic 
forces between the blood and the cytoplasm of the body cells, and that 
this disturbance leads to tissue asphyxia, acidosis, and fatal oligemia, 
accompanied by Symptoms indistinguishable from shock. He be- 
lieves that the essential cause of shock is acapnia. He supports this 
theory, not only by very thorough work on the relation of acapnia 
to shock from several different Standpoints, but also by furnishing 
control experiments, as it were, in which shock is prevented by con- 
servation of the animal's störe of carbon dioxide and also by suc- 
cessful treatment of animals, already in a condition of shock, with 
injections of Ringer Solution containing carbon dioxide. Whether 
this theory fails to stand in whole or in part, its originator deserves 
the greatest credit for calling attention to the possibility that härm 
may arise from neglect to conserve the body 's störe of carbon di- 
oxide, the important functions of which, in the body, have long 
oxide, the important functions of which, in the organism, have long 
been appreciated by physiologists. This theory has been of the 
greatest interest to one of us because of the relation of acapnia to 

^ Some of the work was done in the Surgical Research Laboratory of the 
College of Physicians and Surgeons. 



176 Procccdings Columbia Biochemical Association [Sept. 

artificial respiration, and to the production of shock in connection 
with intrathoracic surgery. It has prompted us to investigate the 
degree of acapnia and the associated shock produced by excessive 
artificial respiration. 

We soon found that the diminution of carbon dioxide in the 
blood in ordinary intrathoracic insufflation was neghgible. On the 
other hand, it has been quite an easy matter for us to reduce the 
amount of carbon dioxide in the blood to from one-third to one-half 
the normal amount by forced rapid inflation and deflation of the 
lungs. The artificial respiration was performed 45 to 90 times a 
minute and was continued for periods varying from 30 minutes to 
3 hours. These experiments have differed from those of Hender- 
son in that the animals were allowed to recover. The trachea was 
not divided but respiration was performed by inserting a large, 
rather tightly fitting, tube through the larynx into the uninjured 
respiratory tract. The blood pressures in our experiments were 
not accurately measured, the animals being left as nearly normal as 
possible after the Operations. The degree of shock was estimated 
entirely from the condition of the animals after the Operation and 
the manner in which they recovered from it. Judged in this manner 
there was nothing about these animals to indicate a serious degree 
of shock or any greater disturbance than could be accounted for. by 
three other factors to which we desire to call attention in connection 
with these experiments and which, unless guarded against, can alone 
cause considerable depression and even death. 

( I ) In all experiments in which excessive artificial respiration 
is employed there is a great reduction in the animal's body heat. 
The temperature can easily fall to 85° F. (2) There is a very 
evident possibility (which we believe to be a fact) that the rapid and 
complete filling of the lungs exercises a definite interference with the 
return of the blood to the heart. The fall of the blood pressure, as 
estimated with the finger, and the rapidity of the heart's actioncoin- 
cide closely with the pressures used to inflate the lungs; indeed, a 
scarcely perceptible pulse may be immediately improved by slightly 
lowering the latter pressures. (3) The duration of the apnea fol- 
lowing these experiments depends as much upon the amount of mor- 
phin and ether administered as upon any other factor. We do not 



I9I2] 'Alfred P. Lothrop i77 

believe that it is possible to produce death by apnea, caused in turn 
by acapnia, without the assistance of the toxic effects of morphin 
and ether. The toxic effects of these drugs must be included as 
factors contributing to the shock, 

This report deals with only one of the phases of the relation of 
acapnia to shock, namely the relation of acapnia, produced by ex- 
cessive artificial respiration, to shock ; and as it is only a preliminary 
report, it is not intended as an answer to Henderson's contention. 
Its purpose is mainly to record two general f acts : (A) That we 
have reduced the amount of carbon dioxide in the blood to nearly 
40 per Cent, of the normal amount, and have maintained this reduc- 
tion for a period of 3 hours, without producing Symptoms of shock; 
and (B) that there are other factors than depletion of the störe of 
carbon dioxid, which, unless properly guarded against, can in them- 
selves cause the death of the animal under experimentation. 

27. Biochemical studies of sulfocyanate.^^ Max Kahn. 
A. The ferric chlorid colorimetric test for sulfocyanate in saliva 
is inexact and unreliable. A negative result by the Bunting suction 
method is no evidence of the absence of sulfocyanate but a positive 
result is suggestive of the presence of a comparatively large amount. 
The pink color spontaneously disappears from the ethereal layer 
in positive tests by the Bunting suction method. Various medicinal 
substances, and also certain Compounds that result from biological 
transformations of proteins and carbohydrates, if excreted in the 
saliva, give a very marked red coloration in the ferric chlorid test, 
similar to that produced by sulfocyanate. 

B. Sulfocyanate occurs in the saliva and salivary glands of man, 
in the salivary glands of oxen, but apparently not in the salivary 
glands of dogs. It occurs in the blood, but the spieen, the pancreas, 
the thymus, the thyroid and the testicles of dogs do not contain it. 
The liver seems to be the gland in the body that contains most 
sulfocyanate, which is also present in bile and in the small intestines. 
The stomach contents of dogs on an ordinary diet were free from 
sulfocyanate. When, however, sodium sulfid was given, the gastric 
mixture contained sulfocyanate. 

'^Kahn: Dissertation, Columbia University, 1912. Conducted under the 
auspices of the Dental Society of the State of New York. 



lyS Proceedings Columbia Biochemical Association [Sept. 

C. Siilfocyanate is excreted in the urine and feces. Its elimina- 
tion in the urine is not dependent upon the amount in the saliva. 
Althoiigh dog saliva is apparently always free from sulfocyanate, 
dog urine invariably contains it. The ingestion of amino acids 
(alanin) and of nitriles (acetonitrile) increases the amount of sul- 
focyanate in the body, as well as in the excreta. Sulfocyanate seems 
to be produced in the body from protein. Results with a fasting 
dog harmonize with this conclusion. The ingestion of sulfur, so- 
dium Sulfid, thioacetic acid, thiourea and taurin did not increase the 
Output of sulfocyanate. 

D. Potassium sulfocyanate is toxic to both plants and animals. 
Its toxicity is so marked that indiscriminate dispensation of the sub- 
stance to people is dangerous. The growth of molds is enhanced 
by potassium sulfocyanate. Yeast fermentation is not affected or 
is stimulated by moderate proportions of potassium sulfocyanate. 
Biological proportions of potassium sulfocyanate have no inhibiting 
influence on the growth of bacteria. The souring of milk is inhib- 
ited by large proportions of sulfocyanate. 

28. The chemical Constitution of renal calculi. Max Kahn. 
Sixteen stones of nephric origin were analysed according to the 
method of Mackarell, Moore and Thomas. ^^ Most of the stones 
were composed mainly of salts of calcium. All of the stones -con- 
tained uric acid or urates in varying amounts, but no stone was 
wholly composed of urates. The shape, color and consistency of a 
stone are not criteria of its chemical composition. Three gouty 
tophi were examined by the murexid test for urates. A negative 
response was obtained in each case, showing that not all gouty 
deposits are composed of uric acid salts. 

29. The colloidal nitrogen in urine from a dog with a tumor 
of the breast. Max Kahn and Jacob Rosenbloom. (Published 
in füll in this issue of the Biochemical Bulletin ).2^ 

30. A non-protein, colloidal, nitrogenous substance in milk. 
Max Kahn and Frederic G. Goodridge. Since the figure ob- 
tained for " total " nitrogen in milk exceeds the sum of the values 
for the known nitrogenous constituents, unknown nitrogenous sub- 

^ Mackarell, Moore and Thomas : Bio-Chemical Journal, 1910, iv, p. 179. 
^ Kahn and Rosenbloom : Biochemical Bulletin, 1912, ii, p. 87. 



1912] "Alfred P. Lothrop 179 

stance must be present. The urines of man and dog contain col- 
loidal nitrogenous material.^* It was thought probable that such 
material is present in all the secretions. 

After a careful process, including the removal of protein without 
hydrolysis, substance was obtained f rom milk which is white, amor- 
phoiis, odorless and tasteless; insoluble in the lipin solvents, but 
forms in water an opalescent Solution which falls to flocculate on 
boiling. This material does not respond to any of the protein 
"color tests." It contained about 5.3 per cent. nitrogen; also car- 
bon, hydrogen, oxygen, and sulfur, but no loosely combined am- 
monia radicals. 

31. A biochemical test for free acid, with a review of the 
methods for estimating the various factors in gastric acidity.^^ 
John L. Kantor. The author presented details along the lines of 
our original publication on this subject.^^ The test is a microscopic 
one and depends upon the immediate expansion of moist collagen 
fibrils when they are immersed in aqueous Solutions containing free 
organic or mineral acids of the kinds that ordinarily appear in gas- 
tric Contents. " Combined " acid^^ and acid salts fail to induce such 
effects. The test may be satisfactorily conducted with a drop of 
liquid and a single collagen fibril. 

Comparative observations indicate that for free mineral acid 
(HCl) the collagen-fibril test is equal in delicacy to the Töpfer and 
Günzberg tests, but that for free organic acid (lactic), or for mix- 
tures of free mineral and organic acids, it is more delicate than the 
latter tests. Comparative studies of common factors of interfer- 
ence with the several tests indicate that the collagen-fibril test ex- 
hibits the greater delicacy. The color of the Solution under exam- 
ination had no effect on the test. Further details from the clinical 
Standpoint, and an abstract of the historical discussion, will be pub- 
lished at an early date. 

32. A study of modifications of the biuret reagent. Mar- 
GUERiTE T. Lee. This investigation was made in the endeavor to 

^ Kahn and Rosenbloom : Biochemical Bulletin, 1912, ii, p. 87. 

^ Kantor : Dissertation, Columbia University, 1912. 

^ Kantor and Gies : Proceedings of the American Society of Biological 
Chemists, 1911, ii, p. 20; Journal of Biological Chemistry, 1911, ix, p. xxvi. 

" Goodridge and Gies : Proceedings of the Society for Experimental Biology 
and Mediane, 1911, viii, p. 107. 



i8o Proceedings Columbia Biochemical Association [Sept. 

discover, if possible, a more effective alkali for the biuret reagent 
than the Standard sodium hydroxid — or a combination of alkalies 
that might be better. 

Fairly strong Solutions o£ the following alkalies, when substi- 
tuted for sodium hydroxid in the biuret reagent,^^ yield Solutions that 
give the biuret test when they are added to dilute Solutions of 
Witte peptone : potassium hydroxid, ammonium hydroxid, calcium 
hydroxid, sodium carbonate, conin, piperidin, ethylene di-amin, tri- 
methyl amin, piperazin, and tetra-ethyl ammonium hydroxid. 

Sodium hydroxid, potassium hydroxid, ammonium hydroxid, tri- 
methyl amin, and tetra-ethyl ammonium hydroxid are excellent as 
alkalies in the biuret reagent. Tri-methyl amin appears to be more 
effective than sodium hydroxid. Tetra-ethyl ammonium hydroxid 
is seemingly as effective as sodium hydroxid when the reagent is 
fresh, but the efficiency of the Solution decreases on standing. 
Piperazin and tetra-ethyl ammonium hydroxid give most satisfac- 
tory tests when an excess of copper is present. There is apparently 
an Optimum amount of copper (sulfate) for each alkali. The study 
is in progress. 

33. A chemical study of salivary mucin. Alfred P. Loth- 
ROP, Salivary mucin from the submaxillary glands of oxen.was 
prepared by the Hammarsten-Levene method. It is a white powder, 
insoluble in water, acid in reaction and readily soluble in dilute 
alkalin Solutions. 

The sodium salt can be prepared by dissolving mucin in nine 
parts of 0.5 per cent. sodium bicarbonate Solution plus one part of 
0.5 per cent. sodium carbonate Solution. The thick Solution is then 
dialysed until it no longer reacts alkalin to phenolthalein but is still 
alkalin to litmus. (Prolonged dialysis completely hydrolyses the 
salt and precipitates the mucin. ) The dialysed Solution may be pre- 
cipitated by the addition of about six volumes of alcohol, although 
electrolyte (NaCl) must be present for complete flocculation. The 
product, washed with alcohol and ether, dries to a fine powder. 

^ Gies : Proceedings of the American Society of Biological Chemists, 1910, 
i. P- ^7Z'> Journal of Biological Chemistry, 1910, vii, p. Ix. Also, Kantor and 
Gies : Biochemical Bulletin, 1912, i, p. 264. 



I9I2] ^Alfred P. Lothrop i8i 

The Salt, having an ash content of 2.7-3.3 per cent., is completely 
soluble in water. A 0.2 per cent. Solution is very much like a rela- 
tively thick natural saliva. The Solution is faintly alkalin to litmus, 
gives all the usual protein tests, including the Molisch test for the 
carbohydrate group, and is precipitated in stringy masses by acetic 
acid. 

Quantitative determinations of nitrogen and ash in mucin prep- 
aration III and its sodium salt gave the f ollowing typical results : 





Ask 


Nitrogen 




Per Cent. 


Found 
Per Cent. 


Calculated 
(Ash Free) Per Cent. 


Preparation III 
Sodium Salt III 


0.28 
3.27 


12.49 
12.20 


12.53 
12.61 



The potassium salt was prepared in the same manner, Fre- 
quent reprecipitations by alcohol render the salts decreasingly solu- 
ble in water. 

These products have been made preparatory to experiments on 
the possible relation of salivary mucin to dental caries, in contin- 
uance of our studies under the auspices of the Section on Stoma- 
tology and Research, of the First District Dental Society, State of 
New York. 

34. A study of some of the more important biochemical 
tests.^^ C. A. Mathewson. Representative substances from the 
following groups were studied in their influence on the tests named 
below: neutral inorganic salts, neutral organic Compounds, acids, 
acid salts, bases, basic salts, biological mixtures and miscellaneous 
materials. Over seventy-five substances or products were used in 
each case. It was found that the ten tests under examination could 
be arranged in the following sequence according to the percentage 
of factors causing interference with them: Sudan III, o; xantho- 
proteic, 4; Hopkins-Cole, 4; Seliwanoff, 5; MoHsch, 6.5; iodine, 
(for starch) 6.5 ; Fehling-Benedict, 10; biuret, 13; Millon, 22; Bar- 
foed, 60. 

The acid salts were the most potent interfering substances, the 
neutral organic Compounds the least potent. Of the salts, ferric 
chlorid was the most active agent of interference. An extension of 
the study is in progress. 

''Mathewson: Dissertation, Columbia University, 1912. 



i82 Procccdings Columbia Biochemical Association [Sept. 

35. A quantitative study of the lipins o£ bile obtained from a 
patient with a biliary fistula. Jacob Rosenbloom. Through 
the kindness of Dr. William Weinberger, of the Lebanon Hospital, 
there was placed at my disposal 3180 c.c. of human bile obtained 
from a patient with a biliary fistula. The fluid had the appearance 
of typical human bile. Its specific gravity was 1.020. The follow- 
ing data were obtained in a quantitative lipin analysis, the results 
being expressed in parts per thousand: Water, 970.2; total solids, 
29.8; cholesterol, 2.61; lecithans, 6.42; fat, 6.85; fatty acids, 1.2; 
soaps, 2.6. Total lipins, 19.68 (1.97 per cent.). 

36. Effects of intraperitoneal injections of epinephrin on the 
partition of nitrogen in urine from a dog. Jacob Rosenbloom 
AND William Weinberger. (Published in füll in this issue of the 
Biochemical Bulletin. )^^ 

37. A case of allergy to common foods.^^ Oscar M. 
ScHLOSs. In a boy now 8 years old marked urticarial lesions were 
caused by the Ingestion of eggs, almonds and oatmeal. The idiosyn- 
crasy to egg was not congenital but was acquired at some time be- 
tween the ages of 10 days and 14 months. Symptoms due to the 
ingestion of oats appeared some time after the child had first eaten 
oatmeal when he was 22 months old. As far as can be ascertained, 
the idiosyncrasy to almonds was manifested the first time this food 
was eaten. 

It was found that cutaneous inoculation of these and certaln 
related food substances produced an urticarial wheal at the site of 
inoculation. The cutaneous reaction was produced only by the 
protein constituents of eggs, almonds and oats. Different proteins 
from the same source varied in activity, some being incapable of 
causing a reaction. Some of the active proteins caused Urticaria by 
mere contact with the unbroken skin. It was possible passively to 
sensitize guinea-pigs to ovo-mucoid (one of the active proteins from 
eggs) by intraperitoneal injections of the patient's blood-serum. 
By feeding ovo-mucoid, in gradually increasing doses, the patient 
became immune to egg. At the same time immunity to oatmeal 
and an apparently decreased susceptibility to almonds occurred. 

^"Rosenbloom and Weinberger: Biochemical Bulletin, 1912, ii, p. 123. 
^ Schloss : American Journal of Diseases of Children, 1912, iii, p. 341. 



I9I2] 'Alfred P. Lothrop 183 

38. The comparative enzyme content of green and varie- 
gated leaves of Tradescantia.^^ Carl A. Schwarze. The re- 
sults of the experiments made to determine the relative enzyme con- 
tent of green and variegated leaves of Tradescantia show that there 
is a marked difference between Juices expressed from them. Etio- 
lated leaves are yellow in their rudimentary stage ; that is, an entirely 
yellow leaf presents this condition when first formed. The etio- 
lated leaves are free from chloroplasts and therefore possess no 
starch. The juice extracted from yellow leaves gives a negative 
Fehling test; that from green portions, a positive test. When yel- 
low leaves are ground in a mortar, and the juice is expressed 
through cheese cloth, a dark brown liquid results. Green leaves 
similarly treated yield a dark green liquid. Alcoholic extracts of 
crushed green and yellow leaves, when filtered, assume a brown 
color. The filtrate from yellow leaves is at first pink but the liquid 
gradually assumes a brown color. The filtrate from the green 
leaves comes through brown immediately. The juice of yellow and 
green leaves, when filtered, gives in both cases a brown filtrate, that 
from the yellow leaves being a reddish brown, When unfiltered 
green juice desiccates, a glossy dark green residue is deposited, 
at the periphery of which a few needle-shaped crystals are seen. 
The juice from the yellow leaves, upon desiccation, deposits a 
brown crystallin mass, the long crystals of which make a figure 
which resembles a polyaster seen in plant cells. Extracts in alcohol 
(80 per Cent.) deposit the greatest amount of crystals. The crys- 
tals from yellow leaves are darker than those from green leaves. 

Such reagents as guaiac and trikresol show the presence of 
oxidase and peroxidase in yellow and green Juices. The yellow 
juice seems to be richer in oxidase and peroxidase. When green 
juice was heated to 72° C, and tested the following day, oxidase 
proved to be present, that temperature having failed to destroy it. 
(Subjecting green juice to high temperatures results in the produc- 
tion of a flocculent precipitate, which Sediments promptly under a 
clear supernatant liquid.) 

Juice from yellow leaves was injected into the nodes and inter- 

** Conducted in the Botanical Laboratory under Dr. Gies' guidance. 



184 Proceedings Columbia Biochemical Association [Sept. 

nodes of healthy green Tradescantia stems. No discoloration or 
yellowing of the injected stem coiild be detected. 

39. Biochemical studies of beryllium sulfate.^^ Emily C. 
Seaman. The experiments vvith beryllium sulfate have shown very 
conclusively that the substance has a marked effect on biochemical 
processes. When administered with the food it produced in dogs 
decided nutritive disturbances, which manifested themselves in loss 
of body weight, total inorganic matter, nitrogen, sulfur and phos- 
phorus. When large doses were administered per os the substance 
caused vomiting before a sufficient amount was absorbed to pro- 
duce any other obvious toxic Symptoms. 

When the calculated lethal dose was administered by a single 
siihcutaneous injection, the substance produced edema and necrosis 
of the tissue extending over a large area. No other decided Symp- 
toms were produced by this method. 

Very gradual intravenous injections of the salt produced decided 
toxic effect. The action of the heart became irregulär — unusually 
rapid and very weak: the respiration also became irregulär and 
shallow. During the course of the injection, there was decided 
tremor but this disappeared soon after the Operation. As a direct 
effect of the injection the temperature increased, sometimes to 
105° F., but about 24 hours before the death of the animal the tem- 
perature began to decrease and steadily feil. After intravenous 
injections there was increased elimination of urine followed by re- 
tention. The feces became diarrheal and bloody. Vomiting began 
about the time the dog refused food or water. 

Beryllium sulfate had a decided inhibitory effect on the action of 
ptyalin, pepsin, and trypsin. It also retarded the action of sucrase 
but not to so great an extent. Solutions of the salt (i per cent. or 
less) did not precipitate proteins from neutral or acid Solutions. 
Below the concentration of M/512 Solution, beryllium sulfate did 
not inhibit the growth of lupin or timothy seedlings, but more con- 
centrated Solutions prevented growth. When present in propor- 
tions less than 0.5 per cent., beryllium sulfate had very little, if any, 
bactericidal action. 

40. Chemical changes in fish during long periods of cold 
storage. Clayton S. Smith. Fresh fish were delivered directly 

^Seaman: Dissertation, Columbia University, 1912. 



1912] 'Alfred P. Lothrop 185 

frort! the boat. Specimens of the same catch were immediately 
placed in storage and delivered to us, at intervals, in a frozen State, 
when they were thawed under uniform conditions and promptly 
subjected to analysis. 

Comparative data were obtained regarding moisture, organic 
matter, inorganic matter, and total solids; ammonia nitrogen, solu- 
ble nitrogen, insoluble nitrogen, coagulable nitrogen, non-coagulable 
nitrogen and total nitrogen; "proteose" nitrogen, both before and 
af ter autolysis ; f at content and f atty-acid number ; and the reducing 
power of the aqueous protein- free extract, as determined by the 
Benedict method. 

The flesh of fish which had been refrigerated not less than four 
months and not more than six months was unaltered in composition. 
After a period of nine months in cold storage there was a sHght, 
almost imperceptihle, increase in the content of ammonia nitrogen, 
but no other change was noted. The work is in progress. 

41. An attempt to sharpen the end point in Benedict's 
method for the quantitative determination of sugar in urine. 
William Weinberger. In Benedict's modification of FehHng's 
sugar titration method "instead of the reduced copper being pre- 
cipitated as the red sub-oxid, which of its own color obscures the 
end point of the reaction, the copper is precipitated as cuprous sulfo- 
cyanate, a snow white Compound, which is rather an aid than a hin- 
drance to accurate Observation of the disappearance of the last trace 
of blue color," However, in applying Benedict's method to urine 
of low sugar content (below 0.5 per cent., as it frequently occurs in 
cases of glycosuria), one is Struck by the fact that the blue color of 
the mixture does not persist until the reaction is ended, for the 
Contents of the porcelain dish assume a dirty brownish-green hue 
that gradually merges into brown. This renders the correct estima- 
tion of the end point very difficult if not impossible. 

Clarifying the urine by the addition of lead acetate previous to 
the titration might overcome the difficulty, but this procedure would 
require additional manipulations and calculations ; and there is also 
the danger of a chemical change in the copper Solution. None of 
these objections apply to the simple method proposed by the author. 
It consists in the addition, just before heating, of approximately 10 



i86 Proceedings Columbia Biochemical Association [Sept. 

grams (two heaping teaspoonsful) of powdered calcium carbonate 
to the Contents of the porcelain dish (25 c.c. of Benedict's Solution, 
5-10 grams of anhydrous sodium carbonate, and a small amount of 
powdered pumice). The titration is then made in the usual manner. 

The snow white calcium carbonate, insoluble and suspended in 
the alkalin Solution, appears to act like the copper sulfocyanate in 
that it efTectively obliterates all colors except the blue color of the 
copper Solution. The end point obtained is sharp, the blue color 
being visible up to the addition of the last two drops of urine that 
are necessary for complete reduction. A sufficient amount of cal- 
cium carbonate (10 grams) must be added, otherwise the precipitate 
will be gray and the end point less distinct. In order to prevent 
sudden ebullition of the concentrated Solution, it is advisable to 
dilute the latter with a little distilled water, Experiments have 
shown that the addition of the calcium carbonate does not introduce 
any noticeable error. 

The author demonstrated these facts. 

42. Diffusibility of protein through rubber membranes, with 
a note on the disintegration of collodion membranes by common 
ethyl ether and other solvents. William H. Welker. {Puh- 
lished in füll in this issue of the Biochemical Bulletin, )2* 

43. A further study of the Bardach test for protein. 
Charles Weisman. {Piihlished in fiill in the June issue of the 
Biochemical Bulletin ).2^ 

44. A study of the surface tension of dog blood-serum by 
the drop-weight method.^^ Harold E, Woodward. These ex- 
periments, about twenty in number, were planned to answer the 
question whether ordinary variations in the blood supply and nutri- 
tive condition of an individual affect the surface tension of the 
blood.^'^ Serum could be handled better than blood and serum from 

** Welker: Biochemical Bulletin, 1912, ii, p. 70. 

** Weisman : Ibid., 1912, i, p. 538. 

^The animals were fed and controlled, and the blood was withdrawn and 
the serum coUected, by Dr. Gies and Mr. Chris Seifert. The drop-weights were 
made by the author in the laboratory of physical chemistry under the direction 
of Prof. J. L. R. Morgan. 

^ These experiments were a logical preliminary to the work described else- 
where: Woodward, Dissertation, Columbia University, 1912. 



I9I2] 'Alfred P. Lothrop 1S7 

clotted blood was more satisfactory than serum obtained by centri- 
fuging defibrinated blood. 

The normal surface tension of dog serum (five dogs), from the 
blood of animals on the usual diet in metabolism experiments in this 
laboratory, is about 45.5 dynes per centimeter. A daily hemorrhage 
of 3 per Cent, or more of the body weight, on two successive days, 
was without material effect on the surface tension. ^^ Small addi- 
tions of salt to the food raised, whereas additions of sugar lowered, 
somewhat the surface tension. The Ingestion of extra quantities of 
meat, several hours before blood was withdrawn, caused a decrease 
of about 1.5 per cent. in the surface tension. Fasting (1-2 days) 
raised the surface tension about i per cent. Copious water drink- 
ing (2 hours before withdrawal of blood) and the administration 
of magnesium sulfate, with resultant marked diarrhea (a short time 
prior to removal of blood from another dog), were without appre- 
ciable effect on the surface tension of the serum. These results 
suggest that the nutritive State of a given individual must be defi- 
nitely established before accurate conclusions can be drawn regard- 
ing the significance of data for surface tension of the subject's 
blood (or serum). 

[The December issue of the Biochemical Bulletin will pre- 
sent abstracts of the scientific Communications at the meeting of the 
Biochemical Association to be held on December 6, at the Columbia 
Medical School.] 

Biochemical Laboratory of Columbia University, 
College of Physicians and Surgeons, 
New York. 

'^When the second bleeding occurred in much less than 24 hours after the 
first, the surface tension was above normal. 



BIOCHEMICAL NEWS, NOTES AND COMMENT 

Contents. I. General: Necrology, i88; in memoriam, i88; anniversary 
celebrations, 189; honors, 190; retirements, resignations and appointments, 190; 
prizes, grants, endowments and funds, 193; meetings of congresses and societies, 
194; buildings and general equipment, 195 ; acts of congress, 196; miscellaneous, 
197. IL Columbia University Biochemical Association: General notes, 200; pro- 
ceedings, 201 ; biochemical department, 201. 

I. General 

Necrology. Dr. W. W. Daniels, emeritus professor of chemis- 
try at the University of Wisconsin. — Thomas Doliber, president of 
Mellin's Food Co., and one of the best known manufacturing drug- 
gists in America. — Dr. Morris Loeb, professor of chemistry at New 
York University and president of the Chemists' Club. — Dr. Her- 
mann Munk, formerly professor of physiology at the veterinary Col- 
lege in Berlin. — Dr. E. A. Holmström, Sweden's foremost pharma- 
cist. — Dr. Edmund von Neusser, professor of internal medicine at 
Vienna. — Prof. Melville Amasa Scovell, director of the Kentucky 
Agricultural Experiment Station and dean of the College of Agri- 
culture of the Kentucky State University. — Dr. Henry Adam 
Weber, professor of agricultural chemistry, Ohio State University. 
— Dr. Thomas Winter, professor of agriculture in University Col- 
lege of North Wales, Bangor. 

In memoriam. Lord Lister. A memorial to Lord Lister will 
be established at University College Hospital. It was in 1843 that 
Joseph Lister entered the College as an arts Student and graduated 
bachelor of arts in 1847. He then became a Student of medicine 
and entered the hospital to complete his studies. A special commit- 
tee has been formed under the presidency of the Duke of Bedford, 
President of the hospital. The exact nature of the tribute will be 
largely decided by the amount of the subscriptions received, but it 
has been suggested that either a bust or a tablet should be placed in 
both the hospital and the College. It is understood that the memo- 
rial will be entirely local in character, and only those who have been 

188 



I9I2] General 189 

in some way connected with University College or the hospital are 
being asked to subscribe. 

The presidents of the Royal Society and the Royal College of 
Surgeons some weeks ago took the necessary steps for the forma- 
tion of a large and representative committee for the purpose of es- 
tablishing a memorial to the late Lord Lister. A meeting of the 
committee, which was largely attended, was held on July 22 at the 
rooms of the Royal Society, under the chairmanship of Sir Archi- 
bald Geikie. The following were appointed an executive commit- 
tee to recommend to a future meeting of the general committee a 
scheme for the memorial to Lord Lister and to organize an appeal 
for subscriptions : The Archbishop of Canterbury, the Lord Chan- 
cellor, Lords Iveagh, Rayleigh, Rothschild and Alverstone, the dean 
of Westminster, the Lord Mayor, the Lord Provosts of Edinburgh 
and Glasgow, the Master of the Rolls, Mr. Lewis Harcourt, M.P., 
Sir T. Barlow, Sir W. W. Cheyne, Sir R. J. Godlee, Sir H. Morris, 
Sir A. Geikie, Sir D. MacAlister, the Hon. Sir C. Parsons, Sir W. 
Turner, Sir J. Wolfe-Barry, Sir J. R. Bradford, Sir A. P. Gould, 
Sir A. Kempe, the Hon. W. F. D. Smith, Mr. F. M. Fry and Mr. 
Edmund Owen. Lord Rothschild and Sir W. W. Cheyne were 
appointed treasurers and Sir J. R. Bradford was appointed secretary 
of the Lister Memorial Committee. 

Dr. Paul C. Freer. The Bureau of Science of the Philippine 
Government has adopted resolutions in memory of Dr. Paul C. 
Freer, director of scientific work in the bureau, who died last April. 
The resolutions express the sense of his associates that " the Bureau 
of Science has suffered a very great loss and that the cause of sci- 
ence in the Philippine Islands has been deprived of one of its most 
zealous and conscientious advocates." 

Anniversary celebrations. June 30: Professor Gad, formerly 
director of the Physiological Institute at Graz, a pupil of Du Bois- 
Reymond, celebrated his seventieth birthday. — July i: Prof. Carl 
Binz, formerly director of the Pharmacological Institute at Bonn, 
celebrated his eightieth birthday. — August ß: Professor Bernstein, 
formerly director of the Institute of Physiology at Halle, celebrated 
the fiftieth anniversary of his doctorate. — September 14: Prof. W. 



190 Biocltemical News, Notes and Comment [Sept. 

O. von Leube, the distinguished clinician, celebrated his seventieth 
birthday. Professor Leubc has been living at Stuttgart since last 
year, when he resigned his directorship of the Würzburg medical 
clinic. 

Honors. 'Awards of prizes. Dr. Alexis Carrel has been 
awarded the Nobel prine in medicine, in recognition of his achieve- 
ments in the suture of blood-vessels and the transplantation of 
Organs. — The Vienna Academy of Sciences has conferred its Liehen 
prise for 191 2 on Dr. Oswald Richter for his work on the food of 
algse. 

Honorary degree. The University of St. Andrews, Dundee, 
Scotland, has conferred the degree of LL.D. on Dr. S. J. Meltzer. 

Foreign associates. Sir William Ramsay and J. Reverdin have 
recently been elected foreign associates of the Paris Academie de 
Medecine. 

Retirements, resignations and appointments. Retirements. 
Col. Martin V. Calvin, for the past six years director of the Georgia 
Agricultural Experiment Station. — Prof. H. J. Wheeler, former act- 
ing-president of the Rhode Island State College, at Kingston, R. I., 
and, during the past eleven years, director of the government agri- 
cultural experiment Station at that Institution. 

Leave of ahsence. Dr. W. P. Bradley, professor of chemistry at 
Wesleyan University, has been granted leave of absence for the year 
1912-13, to organize a department of research for the United States 
Rubber Goods Company. — Dr. A. F. Blakeslee has a year's leave of 
absence from the Connecticut Agricultural College. He has a tem- 
porary appointment on the staff of the Carnegie Station for Experi- 
mental Evolution at Cold Spring Harbor, L. L, where he will study 
lower fungi. 

Appointments have lately been announced, as follows '} 

Bryn Mawr College: Dr. Don R. Joseph (associate in physiology 
and pharmacology at the Rockef eller Institute), associate professor of 
physiology. 

Carnegie Institution, Boston Nutrition Laboratory: Mr. Joseph C. 
Bock (instructor in chemistry at Michigan Agricultural College), 
chemist. 

^In the appended summary, institutions from which resignations occurred 
are named in parenthesis. 



I9I2] General 191 

College of Agriculture and Mechanic Arts (Mayaguez, P. R.) : Dr. 
B. E. Ray (N. C. Experiment Station and College of Agriculture), 
Professor of chemistry. 

Columbia University: Mr. Ernest L. Scott (University of Kansas), 
instructor in physiology; Dr. Otto von Huffman (Cincinnati Hospital 
and Ohio Miami Medical College), instructor in clinical pathology. 

Commission for the study and prevention of malaria in the South : 
Dr. William S. Thayer, member. 

English Government Laboratory, London : Mr. E. Grant Hooper, 
deputy-government chemist (promoted), vice Mr. H. W. Davis, retired. 

Hamburg Botanical Institute: Dr. Hans Winkler (associate Pro- 
fessor of botany at Tübingen), director. 

Harvard University: Dr. Geo. R. Lyman (assistant professor of 
botany in Dartmouth College) will take the work of Professor Roland 
Thaxter during a sabbatical leave of absence. 

Institute for experimental research on Cancer, established by the 
Kaiser Wilhelm Society for the promotion of science: Prof. A. von 
Wassermann, director. 

Margaret Morrison School for Women of the Carnegie Institute 
(Pittsburgh) : Miss Mary D. MacKenzie (professor of biology at 
Western College, Oxford, Ohio), head of the department of biology. 

McGill University (Montreal) : Prof. Francis E. Lloyd (professor 
of botany in the Alabama Polytechnic Institute and plant physiologist 
to the Alabama Experiment Station), MacDonald professor of botany; 
Dr. F. R. Miller, lecturer in physiology. 

Medico-Chirurgical College (Philadelphia) : Dr. H. Lowenherg, 
assistant professor of infantile dietetics and also pediatrist to Mount 
Sinai Hospital, succeeding the late Dr. Edwin Rosenthal. 

Municli medical clinics : Prof. Friedrich Müller, instead of retain- 
ing the second clinic, has taken the first, left vacant by the death of 
Professor Bauer; Prof. E. v. Romberg (Tübingen) succeeds Professor 
Müller. 

N. Y. State Food Laboratory (Ithaca) : Mr. /. T. Ciisick (assistant 
in nutrition investigations, N. Y. Agricultural Experiment Station), 
analyst. 

N. Y. State School of Agriculture (Alfred University at Alfred) : 
Prof. W. J. Wright (Pennsylvania State College), director. 

N. C. Agricultural Experiment Station (West Raleigh) : Dr. Joseph 
F. Brewster, chemist. 

Ohio State University: Dr. W. G. Stover (Oklahoma Agricultural 
Experiment Station), assistant professor of botany. 



192 Biochemical Nezvs, Notes and Comment [Sept. 

Ontario Agricultural College : Mr. R. E. Stone, lecturer in the botan- 
ical department. 

Pennsylvania Chestnut-Tree Blight Commission: Dr. F. D. Heald 
(professor of botany in the University of Texas), pathologist; Miss 
Caroline Rumbold (Missouri Botanical Garden), physiologist in charge 
of tree medication ; Mr. Joseph Shrawder, chemist. 

Reed College (Portland, Oregon) : Dr. Harry Beal Torrey (asso- 
ciate professor of zoology in the University of California), professor 
of biology. 

Skin and Cancer Hospital of Maryland : Mr. /. M. Codd, chemist. 

State University of Oregon Medical College (Portland) : John M. 
Co;mo//y, Ph.D., M.D. (Harvard Medical School), professor of physio- 
logical chemistry. 

U. S. Bureau of Animal Industry : Dr. Frederick J. Birchard (as- 
sistant in chemistry at the Rockefeiler Institute), research chemist in 
the Dairy Division. 

U. S. Bureau of Mines (Pittsburgh) : Dr. /. K. Phelps (U. S. Bu- 
reau of Chemistry, Washington, D. C), chemist. 

U. S. Bureau of Plant Industry : Dr. R. Kent Beattie (professor of 
botany in the State College of Washington), expert in the office of 
forest pathology; Dr. Neil E. Stevens (assistant pathologist in Kansas 
Experiment Station), forest pathologist. 

University College, Reading: Dr. vS". M. T. Auld (lecturer in the 
chemical department of the Southeastern Agricultural College atWye), 
professor of agricultural chemistry; Mr. John Goding (Midland Agri- 
cultural College), research chemist in dairy ing. 

University of Bonn: Professor Johannes Fitting (director of the 
State Botanical Institute at Hamburg), successor of Professor Stras- 
burger. 

University of Illinois: Dr. /. Howard Beard, instructor in physi- 
ology (promotion). 

University of Maryland : Dr. Jsaac M. Macks, pathologist. 

University of Minnesota : Dr. Robert B. Gihson, assistant professor 
of physiological chemistry (promotion) ; Dr. Rodney M. West, assist- 
ant professor of agricultural chemistry (promotion). 

University of South Dakota : Mr. Herbert Otto Lussky (assistant 
in physiology at the University of Chicago), director of the department 
of physiology in the College of arts and sciences and the College of 
medicine. 

University of Vienna: Prof. Wilhelm Türk, temporary successor 
to Prof. E. von Neusser in the medical school (page 188). 



I9I2] General 193 

Washburn College : Dr. Edith M. Twtss, head of the department of 
botany ; Mr. James P. Poole, instructor in botany. 

Washington State College (Pullman) : Dr.IraD. Cardiff (professor 
of botany in Washburn College), professor of plant physiology. 

Prizes, grants, endowments and funds. Prizes. The Col- 
lege of Physicians, Philadelphia, announces that the next award of 
the Alvarenga prise, amounting to about $180, will be made July 14, 
191 3. Essays may be devoted to any subject in medicine but must 
not have been published, and shonld be received by May i, 1913, by 
the secretary of the College, Dr. Thomas R. Neilson, 1937 Chestnut 
Street, who will furnish particulars, on request. — Madame Dieula- 
foy, widow of the late clinician, has given to the Academy of Medi- 
cine, of Paris, in memory of her husband, the sum necessary to 
found the Dieulafoy prise of $400, which will be awarded every 
two years to the author of the best work on the subject of internal 
pathology. — The Riberi prise, amounting to $4,000, will be awarded 
by the University of Turin, after the close of the year 1916, for the 
work which is adjudged to have most advanced the science of 
medicine. 

Grants. Grants for research, at the recent meeting of the Brit- 
ish Association: Mr. A. D. Hall, plant enzymes, £30; Prof. E. A. 
Schäfer, the ductless glands, £40; Prof. E. H. Starling, oxy-hemo- 
globin, £15 ; Prof. F. Gotch, mammalian heart, £20; Sir W. Ramsay, 
for the International Commission on Physical and Chemical Con- 
stants, £40. 

Endowments and funds. The London School of Tropical Med- 
icine is making an appeal for $500,000 to provide for the equipment 
and more efficient conduct of its work. — The late Dr. J. E. Robinson, 
first governor of Kansas, bequeathed $100,000 to the University of 
Kansas. The gift will be used for the medical school. — Mr. James 
B. Brady, of New York, has given the sum of $220,000 to the Johns 
Hopkins Hospital, for the establishment of a ward for the treatment 
of diseases of the kidney. — The late Mr. Allan Octavian Hume, well 
known as an ornithologist and botanist, lately bequeathed about 
£14,000 to the South London Botanical Institute, to which in 1907 
he gave £10,000. — Under the will of the late Augustus W. Open- 
hym, Columbia University will receive a third of a trust fund of 



194 Biochemical News, Notes and Comment [Sept. 

$275,000 for the endowment of research into the cause, prevention 
and eure of Cancer. Mr. Openhym's will stipulates that if at any 
time further investigation of Cancer is not required, the income of 
the fund may be used for research in any brauch of medicine or sur- 
gery. The endowment under Mr. Openhym's will is to be known 
as the Openhym Research Fund, and the terms of the gift are sub- 
stantially the same as those of the Crocker Research Fund, which 
amounts to $1,440,777.13. 

Meetings of congresses and societies. The Fifteenth Inter- 
national Congress on Hygiene and Demography was officially opened 
in the Continental Memorial Hall on September 23 and continued 
until September 27. President Taft delivered an address at the 
opening exercises. The delegates numbered about 3,000, represent- 
ing 33 foreign governments, every American State and territory, 
over 300 American cities, and leading Colleges and universities and 
many scientific, medical and social institutions throughout the world. 
The congress was divided into eleven sections and four general ses- 
sions were held. President Taft was honorary president, Dr. 
Henry P. Walcott, of Massachusetts, was president, and Dr. John S. 
Fulton, of Maryland, was secretary-general, of the congress. A 
füll account of the proceedings is given in the Journal of the Amer- 
ican Medical Association, beginning at page 1207 (September 28). 
The proceedings of the biochemical section — " dietetic hygiene ; hy- 
gienic physiology" — are reported at page 129 of this issue of the 
Biochemical Bulletin. 

The Eighth International Congress of Applied Chemis'try was 
officially opened at Continental Memorial Hall, in Washington, on 
September 4, and continued in New York from September 6-13, in- 
clusive, where the work was centralized at Columbia University and 
the College of the City of New York. About 2,500 members were 
in attendance. Dr. Edward W. Morely was honorary president, 
Prof. William H. Nichols was president, and Dr. Bernhard G. 
Hesse was secretary, of the congress. The scientific work of the 
congress was organized in twenty-four sections. Among the gen- 
eral addresses was one by Prof. Gabriel Bertrand on " The part 
played by infinitely small quantities of chemicals in biological 
chemistry." 



igi2] General i95 

Professor W. H. Perkin delivered a lecture on " The polymeriza- 
tion of butadiene and isoprene," before the Sections on Organic 
Chemistry and India Rubber. Prof. Perkin outlined bis original 
method of making synthetic rubber,^ and then described the follow- 
ing new method : Take ethyl alcohol, which may be easily oxidized 
to acetaldehyde. This is Condensed by means of potassium carbon- 
ate to aldol and the aldol can be quantitatively converted into butyl- 
idine gycol. All the yields of these reactions are practically quan- 
titative. The butylidine glycol is then converted into a chlorid and 
passed over soda-lime, when practically the same product is pro- 
duced as the isoprene from isoamyl chlorid and, when treated with 
sodium, gives even better rubber than isoprene. Professor Perkin 
exhibited samples of what he called the first synthetic rubber ever 
made (the product of Tilden). 

A general review of the proceedings of the Congress will appear 
in the October issue of the Journal of Industrial and Engineering 
Chemistry (pages 706-719). The proceedings of the biochemical 
section are reported at page 150 of this issue of the Biochemical 
Bulletin. 

The eighty-second annual meeting of the British Association for 
the Advancement of Science, which opened at Dundee on Septem- 
ber 4, had a registration of 2,504 members, which is considerably 
larger than the average. At the opening session the President, 
Prof. E. A. Schäfer, delivered a notable address on the "Nature, 
origin and maintenance of life," which has been published in Nature 
(90: 7-19) and Science (36: 289-312). It was announced that 
Dr. J. K. Caird, of Dundee, had given £10,000 to the funds of the 
association. A general account of each sectional meeting will ap- 
pear in Science (36 : 446-452). 

The Royal Society recently celebrated its 25oth anniversary. 

The I4th meeting of the Australasian Association for the Ad- 
vancement of Science will be held in Melbourne in January, 19 13. 

Buildings and general equipment. The work of the Herriman 
Dispensary of the Brooklyn Hospital was inaugurated on July 17. 
The dispensary will be open daily. It is a two and one-half story 

* Biochemical Bulletin, 1912, i, p. 566. 



196 Biochemical News, Notes and Comment [Sept. 

brick and marble structure and was given by Mr. William H. Herri- 
man in memory of his wife. Mr. Herriman donated $100,000 for 
this purpose, $25,000 of which will be used as an endowment fund. 
— Messrs. Jacob H. Schiff, Sei. R. Guggenheim, Ferdinand Sulz- 
berger and Samuel Sach have each given $50,000 to a fund for the 
construction of a private hospital for persons suffering f rom chronic 
diseases, to be built by the Montefiore Home, in the Bronx, New 
York City. — The Medical Faculty of the University of Utah is re- 
questing the Regents of the University to ask the Legislature for a 
special appropriation of $25,000 for the medical school. It is not 
generally known that the State of Utah is doing better by its Uni- 
versity, proportionately, than any other State, in that this Institution 
receives 28 per cent. of the state's income in taxes. The State of 
Utah contains about 400,000 inhabitants. 

Acts of Congress. Public Health Service. The following is 
the text of the act of congress concerning the Public Health Ser- 
vice : Be it enacted by the Senate and House of Representatives of 
the United States of America in Congress assembled. That the 
Public Health and Marine-Hospital Service of the United States 
shall hereafter be known and designated as the Public Health Serv- 
ice, and all laws pertaining to the Public Health and Marine-Hos- 
pital Service of the United States shall hereafter apply to the Public 
Health Service, and all regulations now in force, made in accord- 
ance with law for the Public Health and Marine-Hospital Service 
of the United States, shall apply to and remain in force as regula- 
tions of and for the Public Health Service until changed or rescinded. 
The Public Health Service may study and investigate the diseases 
of men and conditions influencing the propagation and spread 
thereof, including sanitation and sewage and the pollution either 
directly or indirectly of the navigable streams and lakes of the 
United States, and it may from time to time issue Information in 
the form of publications for the use of the public. 

'Amendment to the food and drug act. Congress, before ad- 
journment, passed an amendment to the food and drug act which 
the President has signed, making it illegal " if its package or label 
shall bear or contain any Statement, design, or device regarding the 
curative or therapeutic effects of such article, or any of the ingredi- 



I9I2] General 197 

ents or substances contained therein, which is false and fraudulent." 
It will be remembered that the act of 1906 declared that a drug is 
misbranded "the package or label of which shall bear any Statement 
. . . which shall be false or misleading in any particular . . . " ; but 
the supreme court, by a majority of five to three, decided that this 
did not refer to false Statements regarding the curative effect of a 
drug. 

Miscellaneous items. Proposed State medical service in Eng- 
land. During the recent meeting of the British Medical Associa- 
tion at Liverpool, a State Medical Service Association was formed 
under the inspiration of Dr. B. Moore, professor of biochemistry at 
the University of Liverpool. Prof. Moore lately produced a book 
entitled " The dawn of the health age," in order to demonstrate the 
necessity for entirely remodeling the present System of medical prac- 
tice in the interests of the whole Community. The object of the 
new association is to advocate a State medical service on the follow- 
ing basis : (i) the whole profession to be organized on the lines of 
the other State Services now in existence; (2) entry to the profes- 
sion to be by one state examination; (3) each member of the serv- 
ice to be paid an adequate salary, increasing gradually according 
to the length of service and position in the service, and to be entitled 
to a Pension after a specified number of years or in case of perma- 
nent disablement; (4) members of the public to have, as far as 
possible, free choice of physicians, but no physician to be called on 
to have charge of more than a specified number of patients; (5) one 
of the primary objects of the State service to be to unite preventive 
and curative medicine ; all hospitals to be nationalized and used for 
the purpose of consultative, operative and therapeutic work at the 
request of and in conjunction with the patient's own physician; (6) 
the Services of the state physicians to be open to €very one, rieh 
or poor; (7) the state medical service to be administered by a 
board of health under a minister of public health with cabinet rank, 
assisted by expert medical advisers. This movement was started 
before the insurance act was passed and is quite independent of the 
present impasse. It is intended that the work of the association 
shall form a brauch of sociologic science, and membership will 
be open to all prominent sociologists, whether lay or medical. 



198 Biochemical News, Notes and Comnient [Sept. 

(London correspondent, Journal of the American Medical Associa- 
tion, 19 12, lix, p. 663 : August 10). 

Detection of formaldehyde in foods. In view of the introduc- 
tion of a mixture of nitrite and formaldehyde with the object of 
masking the reactions of the latter when used as a food preserva- 
tive, the following experiments may be of interest. A sample of 
f resh mixture was divided into four portions and treated as follovvs : 
(i) A small amount of commercial formaldehyde Solution was 
added; (2) small amounts of formaldehyde and sodium nitrite were 
added; (3) a small amount of sodium nitrite was added; (4) no 
addition was made. Portions of each of these were tested with 
Rimini's test (Phenylhydrazin hydrochlorid, sodium nitroprussid 
and sodium hydroxid). Prompt reactions for formaldehyde were 
obtained in i and 2; negative results in 3 and 4. Other portions 
of the samples were tested with the well-known test for nitrite (sul- 
fanilic acid and alphanaphthylamin) . The responses of 2 and 3 
were prompt and distinct. No color was produced in i and 4. The 
original mixtures were allowed to stand 24 hours at room tempera- 
ture and the tests repeated with the same results as obtained at first. 
It seems easy, therefore, to unmask nitrite and formaldehyde in the 
presence of each other. Henry Leffmann. {Journal of Industriell 
and Engineering Chemistry, 1912, iv, p. 626: August.) 

Joiirnalistic. With the September number Prof. A. R. Cushny, 
of University College, London, becomes Joint editor with Prof. 
John J. Abel, of Johns Hopkins University, Baltimore, of the Jour- 
nal of Pharmacology and Experimental Therapeutics. At the same 
time, Sir T. Lauder Brunton, of London, Professors J. T. Cash, of 
Aberdeen, W. E. Dixon, of Cambridge, J. A. Gunn, of Oxford, Sir 
Thomas R. Fräser, of Edinburgh, J. N. Langley, of Cambridge, 
C. R. Marshall, of the University of St. Andrews, R. Stockman, of 
Glasgow, F. Ransom, of London and Dr. H. H. Dale, of London, 
join the board of associate editors. By this arrangement the ablest 
representatives of phannacology in Great Britain unite with the 
American and Canadian colleagues in the conduct of the Journal and 
the publishers feel confident that it will henceforth serve as the 
medium of publication for the best pharmacological researches of the 



I9I2] General 1 99 

english-speaking countries. (Publisher's announcement, Septem- 
ber number, vol. iv, no. i.) 

Visiting agriadturalists. Mr. Paul Korchoof, agricultural ex- 
pert, department of the Russian ministry of agriculture, and Mr. 
Vaseelie Yurieff, assistant director, Kharkow Central Agricultural 
Experiment Station, have been visiting the agricultural Colleges and 
stations in this country. — Dr. E. B. Copeland, dean of the College of 
Agriculture, Los Bafios, P. I., who has been visiting the United 
States, recently returned to the Philippines. 

Parsons in Washington. Dr. Charles L. Parsons, secretary of the 
American Chemical Society, moved from Durham, N. H., to Wash- 
ington on September i. The headquarters of the American Chem- 
ical Society may now be addressed, Box 505, Washington, D. C, 

Remsen to remain at Hopkins. Owing to the difficulty of find- 
ing a suitable occupant for the post, Dr. Ira Remsen will remain at 
the head of Johns Hopkins University for the ensuing session, or 
part of it at least. 

Petroleum production in the United States, in 191 1, surpassed its 
own record (made in 1910) by an increase of nearly 11,000,000 
barreis. In 1910 the Output was 209,557,248 barreis. The total 
production of the world also surpassed all previous records, amount- 
ing to over 345,000,000 barreis. 

Johns Hopkins limits enrolment. The dean of Johns Hopkins 
Medical School announces that it has become necessary to limit the 
number of students owing to the restricted space and facilities in the 
various laboratories. The present enrolment is 355, the largest in 
the history of the school, and fifty other students were refused ad- 
mission prior to the beginning of the session. 

Standard rations for nutrition experiments. A Conference was 
held at the Graduate School of Agriculture, Lansing, Mich., on 
July 24, to discuss the formulation of Standard rations for experi- 
mental work in determining the comparative value of feed stuffs. 
Mr. B. H. Rawl, chief of the dairy division, U. S. Department of 
Agriculture, President H. J. Waters, of Kansas Agricultural Col- 
lege, Prof. C. H. Eckles, of Missouri Experiment Station, and other 
leading workers in this field were present and led the discussion. 



200 Biochemical News, Notes and Comment [Sept. 

COLUMBIA UNIVERSITY BIOCHEMICAL ASSOCIATION 

I. General notes 

Miscellaneous items. Dr. Carl L. Alsherg was one of the dis- 
tinguished non-resident scientists to participate, by invitation, in a 
series of lectures, during the late summer at Fordham University 
Medical College, in New York, on nervous and mental diseases. — 
The following members of the Association conducted investigations 
at Woods Hole, Mass., during the summer: Cora J. Beckwith, H. 
B. Goodrich, Louise H. Gregory, Charles Packard, Alwin W. Pap- 
penheimer, Henry J. Spencer, Charles R. Stockard, Isabel Wheeler, 
and L. L. Woodruff. — Dr. 'A. Richard Bliss is editor-in-chief of 
TheMask, the official national organ of the Kappa Psi Fraternity. — 
Prof. R. Burton-Opitz is now in Europe, where he is spending a 
half-year leave of absence. 

Officers of societies. Section (V) on Control of Infectious 
Diseases of the I5th International Congress on Hygiene and Demog- 
raphy (page 194) : Dr. Charles F. Boldiian, secretary. — New York 
Post-Graduate Medical School and Hospital : Dr. Arthur F. Chace, 
secretary (reelected). — Section (IV) on Organic Chemistry of the 
8th International Congress of Applied Chemistry (page 194) Dr. 
Harry L. Bisher, secretary. — N. Y. Entomological Society: Prof. 
Raymond C. Oshiirn, president. — American Association for the 
Study and Prevention of Infant Mortality: Dr. Philip Van Ingen, 
secretary. 

Appointments. Jefferson Medical College (Philadelphia) : Dr. 
Philip B. Hawk (professor of physiological chemistry, University 
of Illinois), professor of physiological chemistry and toxicology. — 
Rockefeiler Institute for Medical Research: Dr. Michael Heidel- 
berger (recently returned from Zürich), fellow in chemistry. — 
Johns Hopkins University: Dr. John Howland (professor of 
pediatrics in Washington University, St. Louis), director of the 
Harriet Lane Home for Invalid Children, professor of pediatrics, 
and physician in charge of the pediatric department of Johns 
Hopkins Hospital. — ^Cornell University Medical College, Loomis 
Laboratory : Miss Jessie A. Moore (assistant at the Rockefeiler Insti- 
tute for Medical Research), chemical assistant. — N. J. Agricultural 
Experiment Station: Mr. Carl A. Schwarze, assistant plant pathol- 




^, 



Ol*jJJ_ 



"X-^Wv^n^, 



1912] Columbia University Biochemical Association 201 

ogist. — Long Island Medical College: Dr. Matthew Steel (assist- 
ant Professor of physiological chemistry, University of Missouri), as- 
sistant professor of physiological chemistry and pharmacology. — At 
a recent annual meeting of the Imperial Cancer Research Fund, in 
London, Dr. William H. Woglom was appointed first assistant in 
New York, a position maintained under the auspices of the Crocker 
Fund for the investigation of Cancer. Dr. Woglom has returned 
from London, where he had been pursuing a course of study under 
Dr. Bashford, director of the Imperial Cancer Research Fund. 

2. Proceedings of the Association. 

Abstracts of the scientific proceedings of the third annual meet- 
ing (June) are pubHshed on pages 156-187 of this issue. 

3. Columbia Biochemical Department. 
The new Assistant Professor, Dr. Paul E. Howe, B.S., A.M., 
PhD.^ Memorandum which was presented to the Faculty 
OF Medicine with Dr. Howe's nomination to the Assistant 

PROFESSORSHIP IN BIOLOGICAL CHEMISTRY. 

Paul Edward Howe was born in Chicago, Illinois, on July 29, 
1885. His early education was received in the public schools of 
Chicago, Champaign and Urbana, Illinois (1890-1901). He at- 
tended the Urbana High School (1899-1901) and spent a year 
(i90i-'02) in the Preparatory School of the University of Illinois. 
At the end of a four-year course at the University of Illinois he 
received the degree of B.S. in Chemistry in 1906. 

Since 1906 he has been a graduate Student and officer at the Uni- 
versity of Illinois, passing by promotion through the grades of 
Scholar in chemistry in the graduate school (i9o6-'o7), assistant 
chemist in the laboratory of physiological chemistry (1907-08), 
assistant in physiological chemistry (1908-10), and instructor in 
physiological chemistry (i9io-'i2). 

In 1907 he received the degree of M.A. ; in 1910, the degree of 
Ph.D. His major subject for the Ph.D. degree was physiological 
chemistry, with Professor P. B. Hawk; his minor subjects were 
physical chemistry, physiology and histology. 

* Biochemical Bulletin: 1911-12, i, pp. 136, 570, 573 and 574. 



202 Biochcmical N'czvs, Notes and Comment [Sept. 

Dr. Howe is a member of the American Society of Biological 
Chemists, American Chemical Society, American Society of Animal 
Nutrition, American Association for the Advancement of Science, 
Illinois Academy of Science, Sigma Xi, Phi Lambda Upsilon, and 
the Gamma Alpha Graduate Scientific Fraternity. 

Dr. Howe's publications. 1907. The electrolytic corrosion of 
brasses (with A. T. Lincoln and David Klein) ; Journal of Physical 
Chemistry, 11, 501. 

1908. Comparative tests of Spiro's and Folin's methods for the 
determination of ammonia and urea (with P. B. Hawk) ; Proceedings 
of the American Society of Biological Chemists, i, 104; Journal of 
Biological Chemistry, 4, p. x. 

1909. Comparative tests of Spiro's and Folin's methods for the 
determination of ammonia and urea (with P. B. Hawk) ; Journal of 
Biological Chemistry, 5, 477. — On the preservation of feces (with T. A. 
Rutherford and P. B. Hawk) ; Proceedings of the American Society 
of Biological Chemists, i, 196; Journal of Biological Chemistry, 6, 
p. xlix, 

1910. On the preservation of feces (with T.A. Rutherford and P. 
B. Hawk) ; Journal of the American Chemical Society, 32, 1683. — A 
study in repeated fasting (with P. B. Hawk) ; Proceedings of the 
American Society of Biological Chemists, i, 259; Journal of Biological 
Chemistry, 7, p. xlvi. — Fasting studies on men and dogs (with H. A. 
Mattill and P. B. Hawk) ; Proceedings of the American Society of 
Biological Chemists, i, 260; Journal of Biological Chemistry, 7, p. xlvii. 
— Nitrogen partition in repeated fasting; Dissertation (pp. 42), pre- 
sented in partial fulfilment of the requirements for the degree of Doctor 
of Philosophy (University of Illinois). 

191 1. On the differential leucocyte count during prolonged fasting 
(with P. B. Hawk) ; Proceedings of the American Society of Biological 
Chemists, 2, 15; Journal of Biological Chemistry, 9, p. xxi. — Fasting 
studies : I. Nitrogen partition and physiological resistance as influenced 
by repeated fasting (with P. B. Hawk) ; Journal of the American 
Chemical Society, 33, 215. — Fasting studies: III. Nitrogen partition of 
two men through two seven-day fasts following the prolonged Ingestion 
of a low-protein diet : Supplemented by comparative data from the sub- 
sequent feeding period (with H. A. Mattill and P. B. Hawk) ; Journal 
of the American Chemical Society, 33, 568. — Fasting studies: V 
(Studies on water drinking: XI). Influenae of an excessive water 
Ingestion of a dog after a prolonged fast (with H. A. Mattill and P. B. 
Hawk) ; Journal of Biological Chemistry, 10, 417. 



I9I2] Columbia University Biochemical Association 203 

1912. A metaboHsm study on a fasting man (with P. B. Hawk) ; 
Proceedings of the American Society of Biological Cheniists, 2, 65 ; 
Journal of Biological Chemistry, 11, p. xxxi. — Hydrogen-ion concen- 
tration of fecal extracts (with P. B. Hawk) ; Proceedings of the 
American Society of Biological Chemists, 2, 66; Journal of Biolog- 
ical Chemistry, 11, p. xxxii. — Studies on water drinking: XIII (Fast- 
ing studies: VIII). Hydrogen-ion concentration in feces ; Journal of 
Biological Chemistry, 11, 129. — A comparison of the data from two 
fasts each exceeding one hundred days in length and upon the same 
subject (with P. B. Hawk) ; Proceedings of the American Physiological 
Society, American Journal of Physiology, 29, p. xiv. — On the differ- 
ential leucocyte count during prolonged fasting (with P, B. Hawk) ; 
American Journal of Physiology, 30, 174. — Fasting studies: VI. Dis- 
tribution of nitrogen during a fast of 117 days (with H. A. Mattill 
and P. B. Hawk) ; Journal of Biological Chemistry, 11, 103. — The gen- 
eral aspects of fasting: Address before the Columbia University Bio- 
chemical Association, May i, 1912; Biochemical Bulletin, 2, 90. — 
The distribution of urinary nitrogen as influenced by the Ingestion of 
moderate and copious quantities of distilled water at meal time (with 
D. W. Wilson and P. B. Hawk) ; (in press) Journal of the American 
Chemical Society, 34, Proceedings, p. ^^i- — Addendum. The utiliza- 
tion of individual proteins by man as influenced by repeated fasting 
(with P. B. Hawk) ; Proceedings of the Eighth International Congress 
of Applied Chemistry (preliminary edition), 19, 145. (William J. Gies, 
Secretary of the Faculty of Medicine. ) 

Resignations and appointments. The following changes in 
the staff for the year 1912-13 were officially authorized prior to 
October i, IQ12: Dr. Paul E. Howe, assistant professor, vice Prof. 
Wm. H. Welker, resigned; Dr. Clayton S. Smith, instructor (pro- 
moted), vice Dr. Ernest D. Clark, appointed instructor in chemistry 
at the Cornell University Medical College; Dr. Frederic G. Good- 
ridge, assistant, vice Reuben Ottenberg, resigned ; Messrs. E. G. Mil- 
ler, Jr., and Arthur Knudson, assistants, vice Dr. C. S. Smith (pro- 
moted) and Mr. A. R. Rose, resigned; and Misses Ethel Wickwire 
and Tula L. Harkey, assistants (at Teachers College), vice Mr. E. 
G. Miller, Jr. (promoted) and Miss Blanche Harris, resigned. 

Summer session. Courses. Professor Gies kept the labora- 
tory open daily throughout the summer and conducted courses 
(July 5-August 15) in nutrition at the College of Physicians and 



204 Biochemical News, Notes and Comment [Sept. 

Surgeons with Dr. C. S. Smith's assistance, and at Teachers' Col- 
lege with the aid of Dr. Emily C. Seaman and Miss B. E. Shaffer. 

Investigators. The workers named below conducted research, 
in the biochemical laboratory at the College of Physicians and Sur- 
geons, during all or part of the summer vacation : Louis Berman, 
R. J. Cook, Edward Cussler, F. R. Eider, N. B. Foster, Wm. J. 
Gies, Samuel Gitlow, Isidor Greenwald, W. M. Kraus, Alfred P. 
Lothrop, H. A. Mattill, H. O. Mosenthal, Jacob Rosenbloom, Emily 
C. Seaman, C. S. Smith, William Weinberger, Charles Weisman, 
Wm. H. Welker, Harry Wessler. 

Miscellaneous notes. Professor Gies was vice president of the 
Section on Biochemistry including Pharmacology of the 8th Inter- 
national Congress of Applied Chemistry (page 150). — Dr. H. 0. 
Mosenthal recently returned from Tübingen, where he had been 
working in the medical clinic under the direction of Prof. E. von 
Romberg. — Dr. Jacob Rosenbloom has resigned the affiliated Posi- 
tion of assistant pathologist at the German Hospital. — Mr. A. R. 
Rose has lately completed the requirements for the Ph.D. degree and 
will be publicly examined in October. He has begun a special study 
of amylase with Prof. H. C. Sherman. — Mr. Joseph Hepburn- has 
begun work as " university fellow " in biological chemistry. 



EDITORIALS 

Ernst Schulze was one of the great pioneers in biological chem- 
istry. He worked in an inspired way along the zone between the 
old zoöchemistry and the ancient phytochemistry, and achieved the 

distinction of removing the barriers between 

rnst c uze these two fields and uniting them in one great 
open biochemical territory. He brought Hght and understanding 
into large domains where darkness and doubt prevailed. His ex- 
ample in industry, patience, perseverance, devotion, enthusiasm, abil- 
ity and productiveness has been an inspiration to biochemical work- 
ers the world over. Schulze's classical achievements and Service 
will be forever linked with the history of fundamental developments 
in a great formative scientific era. His name and Service will be 
justly remembered, as his memory will be venerated, for very many 
generations. 

As the methods of chemical analysis become more delicate and 

refined there appears ever increasing evidence that the maintenance 

of health and nutrition depends not alone on the caloric values of 

T . ^ ^1. u food-stuffs and the relativ^e proportions of nitro- 
Important though ^ . 

unknown factors in gen and carbon in the diet, but quite as much on 
nutrition other factors which we are beginning fully to 

appreciate. Scurvy has long been one of the indications that there 
are certain unappreciated factors in a normal diet, and the antiscor- 
butic action of vegetables and vegetable Juices is strong emphasis on 
this point. The researches of Hart, McCollum, Steenbock and 
Humphrey/ on cattle, and of Osborne and Mendel,^ on rats, are 
among the many recent studies that reveal the importance of such 
unknown though influential factors in their broad bearing. 

The disease beriberi is a concrete example of the disturbance of 
such subtle influences. For a considerable time physicians in the 

^ Hart, McCollum, Steenbock and Humphrey : University of Wisconsin 
Agricultural Experiment Station Research Bulletin, No. 17 (June, 1911). 
* Osborne and Mendel: Carnegie Institution, Puhlication 156. 

205 



2o6 Important Factors in Nutrition [Sept. 

Orient have believed that certain foods were responsible for this 
form of Polyneuritis. Miura believed the noxious agent to be 
contained in a certain fish, which is much eaten raw ; but more re- 
cently the blame has fallen on rice. It has been asserted that in the 
prisons of Java, beriberi occurs in one out of every forty prisoners 
when shelled rice is eaten; in one out of ten thousand, if the un- 
shelled grain is used. The classical studies of Schaumann were sug- 
gested by observations of this kind. Schaumann believed that since 
polished rice is poor in phosphorus, beriberi is due to a deficiency of 
certain organic phosphorus Compounds. This hypothesis had some 
Support in the fact that materials which relieve the pain of neuritis, 
such as bran, are rieh in phosphorus, but the later investigations of 
Wieland^ cast doubt on the accuracy of these deductions, since it 
could not be shown that the total body-phosphorus was much in- 
fluenced by feeding mice on polished rice. In this connection the 
researches of Fingerling,^ and of McCollum and Halpin,^ are sug- 
gestive, for they have shown a synthesis of organic phosphorus Com- 
pounds from inorganic phosphates. 

The latest contributions to the study of beriberi were made by 
Chamberlain and Vedder,^ by whom it has been shown that extracts 
of rice-bran are effective as therapeutic agents and that these ex- 
tracts contain mere traces of phosphorus. The active substance in 
the bran has not yet been identified, but the interesting feature dis- 
closed by the present evidence as to the etiology of kakki is that a 
food stuff may contain an ingredient which is essential in order to 
prevent injury to the tissues by other components of such food ma- 
terial. Rice grain is harmless when eaten with the pericarp but, 
if the latter is removed by " polishing," a malady ensues which may 
be cured by extracts made from the pericarp. These facts present 
a new face to the idea of " balanced rations " and also remind us 
of the broad biological significance of Loeb's "balanced Solutions." 

N. B. F. 

' Wieland : Archiv für experimentelle Pathologie und Pharmakologie, 1912, 
Ixix, p. 293. 

* Fingerling: Biochemische Zeitschrift, 1912, xxxviii, p. 448; xxxix, p. 239. 
"McCollum and Halpin : Journal of Biological Chemistry, 1912, xi (Pro- 

ceedings of the American Society of Biological Chemists, p. xiii). 

* Chamberlain and Vedder: Philippine Journal of Science, 1912, vi, p. 251. 



1912] Editoriais 207 

In a circular with this title, Director Russell of the Wisconsin 

A'gricultural Experiment Station^ has recently given an interesting 

summary of the perfection of the Babcock quantitative test for milk- 

rru ~- t ^ fat and the influence which it has exerted on 
The Coming 01 age 

of the dairy science and practice throughout the world. 

Babcock test Milk and its numerous products play so impor- 
tant a röle in the economy of the home and in the dietary of the sick 
that the significance of Professor Babcock's contribution cannot re- 
main unnoticed in the annals of the medical world. The simple, 
yet highly accurate Babcock method of estimating the fat content 
of milk and cream finds daily application not only in dozens of 
analytic laboratories, but likewise in hundreds of creameries, in milk 
establishments, and even in the office of the busy practitioner of 
medicine, where a few inexpensive devices enable him to gauge the 
richness of a breast-milk or a modified milk mixture with facility. 

Every pediatrist appreciates what the Babcock test means for the 
exigencies of practice and successful feeding. Today, twenty-two 
years after the introduction of this procedure which, as Ex-Gover- 
nor Hoard remarked, has made dairymen more honest than the 
Bible because it has removed all opportunity for them to profit by 
any deceit, it is interesting to note that no change has been made in 
the essential features of the test during all this period. The tech- 
nic of the Operation remains the same as when the details were pub- 
lished by Dr. Babcock in 1890. The Stimulus which it has given 
to scientific dairying, to the standardization and improvement of our 
milk-supplies, to the possibilities of rational infant-feeding, and to 
what these in turn involve in the direction of the public health, is 
scarcely appreciated by the medical profession. Director Russell 
has written that the Babcock test frees the dairy farmer from the 
fetters of past traditions, and removes him from the category of 
"mossbacks." The influences here referred to have in fact been 
even more far-reaching. 

An additional feature deserves mention: No patent was taken 
out on either the method or the apparatus required to carry out the 
Babcock test. There zuas no copyrighting of a name — no commer- 

' University of Wisconsin Agriculitiral Experiment Station, Circular of 
Information, No. 2^, 1912. 



2o8 Babcock Test [Sept. 

cialism. In accord with a code of ethics now more generally recog- 
nized than at any time, the discoverer, becaiise of his connection with 
the State experiment Station, gave his invention freely to the world. 
We may gladly join in acknowledging our Obligation to the man 
whom the grate ful State of Wisconsin has presented a medal in 
recognition of " his iinselfish dedication of these inventions to the 
public Service." (Editorial : Journal of the American Medical As- 
sociation, 191 2, lix, p, 544.) 



The discovery and investigation of the specific secretions of the 
so-called ductless glands and of other organs make one of the most 
interesting chapters in physiology. Much has been learned con- 

cerning these secretions and their röle in the 

rgano- er py j^^Q^jy These extracts, theoretically, should be of 
great value in the treatment of diseases in which a certain gland or 
glands are deficient or entirely lacking in function. But actual ex- 
perience has been disappointing as a rule, for two reasons : ( i ) The 
diagnosis of insufficiency of secretion on the part of a certain gland 
or Organ is usually most difficult; (2) and even when a correct 
diagnosis is made, it is rarely possible to administer the gland sub- 
stances in such a way as to develop their specific activity. 

A notable exception to this experience is the successful use of 
thyroid extract in thyroid insufficiency or myxedema. Suprarenal 
substance has also proved highly use ful as a circulatory stimulant 
and hemostatic, but not for the treatment of Addison's disease. It 
can safely be said that the administration of gland substance from 
the thymus, hypophysis, ovaries, pancreas, testicles, etc., for dis- 
eases of these organs, has hitherto met with failure. Only härm 
can come from their promiscuous use before careful experimentation 
fully determines their value. 

A wholesome skepticism concerning the efficiency of preparations 
of the digestive enzymes is likewise commended. After years of 
usage many of our best clinical observers believe that pepsin, " pan- 
creatin " and the amylases are of little or no value. The use of se- 
cretin more recently has been similarly disappointing. The con- 
tinued routine use of these preparations is due chiefly to the claims 
of manufacturers. 



I9I2] Editorids 209 

We congratulate our English confreres on the successful con- 
summation of their plans for the formation of a biochemical society 
and the publication of a biochemical Journal^ under their associated 
Biochemical So- control. In this coimtry we have long derived 
ciety, England great benefit from the meetings and activity of 
the American Society of Biological Chemists and are confident our 
English colleagues will have a similar experience. We felicitate the 
biochemical profession at large on this further evidence of the rapid 
growth in usefulness, and the prominent place of Service, of biochem- 
ical art and science. The Bio-Chemical Journal has been highly 
esteemed in America, and we wish it long life and distinguished 
Service under its new management. " Science is essentially mutual- 
istic and the success of one Organization is the gratification of all — 
the triumphs and discoveries of one are shared with the many, 
and the feeling of pride in the progress of the one may he shared. 
zvithout loss by sister organizations. As the discovery made in one 
brauch of science may be the necessary foundation for the Solution 
of some problem in another, so the contribution from one society 
may be the stepping stone to advancement in another. It is all hail 
then, greetings and felicitation — and Godspeed in the accomplish- 
ments of your future destiny." 



The name of the writer of this note might suggest a strong par- 
tiality on his part for the incorporation into biochemical discussions, 
in English, of such words as " Baustein." He believes, neverthe- 
" Baustein " or ^^^^' ^^^^ English phrases of equal f orce thoug'h 
" construction Unit** of more abstract significance, such as "construc- 
tion Unit" for "Baustein," are more acceptable, especially to stu- 
dents receiving their introduction, in English, to the subject of pro- 
tein synthesis and similar processes. 



The foregoing remarks recall the common use, in English, of 
" Splitting product " or " split product " as equivalents for " Spal- 
tungsprodukt," when the substance referred to is neither " Splitting 
Splitting productsor "^^ "split," but has resulted from cleavage. 
cleavage products Why not term such substances "cleavage prod- 
ucts" in conformity with good usage in analogous relationships ? 

"Halliburton: Biochemical Bulletin, 1912; ii, p. 128. 



2IO X-Rays fSept. 

We received recently, with very great pleasure, a foreign money 
Order for twelve dollars instead of tzvelve Shillings in payment of 
Volume I of the Biochemical Bulletin. In view of the fact that 

this overpayment did not excite a desire to dis- 
A rare complimen ^^ontinue the subscription, we have proceeded 
with more enthusiasm than ever with our editorial work, in the hope 
that future volumes of the Bulletin may be much more deserving 
of such a compliment. 

The doing that makes commerce is born of the thinking that 

makes scholars. — Ruskin. 

Perhaps the most valuable result of all education is the ability 

_. _ to make yourself do the thing you have to do, 

when it ought to be done, whether you like it or 
not. — Huxley. 

The fabric of medical progress — indeed, of all progress — is 
woven from legitimate dreams to a greater extent than the " practi- 
cal" man is wont to realize or willing to admit. Editorial: Journal 
of the American Medical Association, 1912, lix, p. 1195. 

Who is it that, when years are gone by, we remember with the 
purest gratitude and pleasure? Not the learned or clever. But 
those who have had the force of character to prefer the future to the 
present, the good of others to their own pleasure. — Stanley. 

A fig for yesterday's convictions ! They were the cocksure 
beliefs of children lost in the dark. This is another day, and we've 
grown overnight. Do you plead the dignity of fixed opinion? It 
is enough for us to say : " We believed it when we affirmed it ; we 
have learned and changed our minds." — Ana Phylactic. 

The successful man, whether in business, in the professions and 
trades, or in politics, enjoys the game for its own sake. He is not 
a conscript in life's battles, but a volunteer, The way interests him 
as much as the goal. Not only the result, but the exercise of powers 
necessary to achieve it, gives him satisfaction. — AI I. Phatic. 

Speaking mentalwise, overfed conceit equals the blind staggers. 
The easiest kind of intoxication is that which feeds upon the poisons 
distilled by a self-caressing imagination. Open the floodgates of 
self-approval and soon you won't know whether you are making 
good or not, for you won't be able to present an intelligent compari- 
son of your own achievements with those of others. — Jaun Dice. 



BOOKS RECEIVED 

The BiocHEMicAL Bulletin will promptly acknowledge, under this heading, 
the receipt of all publications that may be presented to it. From time to time, 
selections will be made for review on pages of the volume to be appropriately 
indicated here. Reviews will be matter-of-fact Statements of the nature and 
Contents of the publications under consideration, and will be intended solely to 
guide possihle purchasers. The wishes or expectations of publishers or donors 
of volumes will be disregarded, when they are incompatible with our convictions 
regarding the interests of our colleagues. The size of the printed pages, in 
inches, is indicated in the appended notices. 

Practical physiological chemistry. A book designed for use in courses in 
practica! physiological chemistry in schools of medicine and of science. By 
Philip B. Hawk, professor of physiological chemistry and toxicology in the 
Jefferson Medical College of Philadelphia. Fourth edition, revised and en- 
larged. Pp. 475 — 4J^X8; $2.50 net. P. Blakiston's Sons & Co.. Philadelphia, 
1912. 

The protein element in nutrition. (One of the International Medical Mono- 
graphs.) By Major D. McCay, professor of physiology, Medical College, Cal- 
cutta. Pp. 216 — 4X7, with 8 füll page portraits of human subjects; $2.00 net. 
Longmans, Green and Co., New York; Edward Arnold, London, 1912. 

Oxidations and reductions in the animal body. (One of the Monographs 
on Bio chemistry.) By H, D. Dakin, The Herter Laboratory, New York. Pp. 
135 — 41^X8; $1.40 net. Longmans, Green and Co., 1912. 

Researches on cellulose. III (1905-1910). By C. F. Gross and E. J. Bevan. 
Pp. 173 — 3><X6; $2.50 net. Longmans, Green and Co., 1912. 

An introduction to the study of the protozoa, with special reference to 
the parasitic forms. By E. A. Minchin, professor of protozoology in the Univer- 
sity of London. Pp. 517 — 4 X 75^ ; $6.00 net. Longmans, Green and Co., New 
York; Edward Arnold, London, 1912. 

Studies from the Rockefeller Institute for Medical Research. Reprints: 
Volume XV ; 1912. (48 reprints). 

CoUected reprints o£ papers. By Graham Lusk. (Researches, III; 1907- 
'11 — II reprints). 

Studies from the Department of Physiology, Cornell University Medical 
College, 1911-1912. (11 reprints). 

Studies from the Departments of Pathology, Bacteriology, Experimental 
Pathology and Experimental Therapeutics, Cornell University Medical 
College, 191 1. (12 reprints). 

Les produits biologiques medicinaux. By P. Byla and R. Delaunay. Pp. 
466 — 3}iX6yi. Societe d'editions scientifiques et medicales, F. Gittler, Directeur, 
Paris, 1912. 

E. Merck's Jahresbericht über Neuerungen auf den Gebieten der 
Pharmakotherapie und Pharmazie: 25 Jahrgang (1911). E. Merck, Chemi- 
sche Fabrik, Darmstadt, 1912. Pp. 531 — 4X7, with a general index of volumes 
1-25. 

Optica! Instruments: Adam Hilger, Ltd. 75 a, Camden Road, London, 
igi2. (Catalogue). 



OFFICERS OF THE BIOCHEMICAL DEPARTMENT OF 
COLUMBIA UNIVERSITY, 1912-1913* 

OFFICIAL REGISTER, SEPT. 30, 1912 

William J. Gies: Professor and Chairman of the Staff; Consulting chemist, 
New York Botanical Garden; Pathological chemist, Bellevue Hospital ; Mem- 
ber of the Faculties of N. Y. Teachers College and N. Y. College of 
Pharmacy. [B.S., Gettysburg College, 1893 and M.S., 1896; Ph.B., Yale 
University, 1894; Ph.D., 1897. Instructor, i89&-'02; adjunct professor, 1902- 
'05; Professor, 1905-.] 

Paul E. Howe: Assistant Professor, 1912-. [B.S., University of Illinois, 1906, 
A.M., 1907 and Ph.D., 1910.] 

Nellis B. Foster: Associate; Associate Physician, New York Hospital ; Chemist, 
St. Luke's Hospital. [B.S., Amherst College, 1898; M.D., Johns Hopkins 
University, 1902. Instructor, i9o6-'o8; associate, 1908-.] 

Walter H. Eddy: Associate and Secretary of the Staff. [B.S., Amherst Col- 
lege, 1898; A.M., Columbia, 1908 and Ph.D., 1909. Assistant, i9o8-'io; 
associate, 1910-.] 

Jacob Rosenbloom : Associate; Pathological chemist, German Hospital. [B.S., 
University of Pittsburg, 1905; M.D. and Ph.D., Columbia, 1909. Assistant, 
i909-'io; associate, 1910-.] 

Alfred P. Lothrop: Associate and Departmental Registrar. [A.B., Oberlin, 
1906 and A.M., 1907; Ph.D., Columbia, 1909. Assistant, i9o8-'o9; instructor, 
i909-'i2; associate, 1912-.] 

Herman O. Mosenthal: Instructor; Assistant Attending Physician, Presby- 
terian Hospital; Assistant Physician, Vanderbilt Clinic; Instructor- in medi- 
cine. [A.B., Columbia, 1899 and M.D., 1903. Assistant, igoS-'og; instructor, 

1909-.] 
Emily C. Seaman: Instructor. [B.S., Adelphi College, 1899; A.M., Columbia, 

1905 and Ph.D., 1912. Tutor, 190g-' 10; instructor, 1910-.] 
Clayton S. Smith: Instructor. [B.S., Rutgers College, 1910 and M.S., 1912. 

Assistant, i9io-'i2; instructor, 1912-.] 
Edg.\r G. Miller, Jr. : Assistant, 191 1- [B.S., Gettysburg College, 1911.] 
Frederic G. Goodridge: Assistant, 1912-. [A.B., Harvard University, 1897; 

M.D., Columbia, 1901.] 
Arthur Knudson : Assistant, 1912-. [A.B., University of Missouri, 1912.] 
Ethel Wickwire : Assistant, 1912-. [A.B., Tri-State College, 1909.] 
TuLA L. Harkey: Assistant, 1912-. [A.B., Colorado College, 1909.] 
Christian Seifert: Laboratory assistant, 1898-. 
Stella Waldeck : Recorder, 1908-. 

Blanche E. Shaffer: Laboratory assistant, summer session, 1912. 
Joseph S. Hepburn: University fellow, 1912-13. [A.B., Central High School, 

Philadelphia, 1903 and A.M., 1908; B. S., University of Pennsylvania, 1907 

and M.S., 1907.] 

*The work of the department was inaugurated in October, 1898, by Prof. 
R. H. Chittenden (lecturer and director), Dr. William J. Gies (instructor), 
Messrs. Alfred N. Richards and Allan C. Eustis (assistants), and Christian 
Seifert (laboratory assistant). 



COURSES OFFERED BY THE BIOCHEMICAL DEPARTMENT OF 
COLUMBIA UNIVERSITY. 1911-1912 

Courses 51 (log), 105 and 215 are given during the first half-year only. 
Course loi is given during the first half-year and is repeated (102) during the 
second half-year. Courses 104 and iio (52) are given only during the second 
half year. All other courses are conducted throughout the entire academic year. 
All courses not otherwise specified are given at the College of Physicians and 
Surgeons. 

(Abbreviations : C, Conference; D, demonstration ; L, lecture; hw, labora- 
tory work; R, recitation.) 

ORGANIC CHEMISTRY 

51 (109) Elementary ORGANIC CHEMISTRY. Introductory to courses loi, 102 
and IIO (52). (Required of first year students of medicine.) L, i hr. D, i hr. 
R, 2 hr., each section (2). Lw, 6 hr. each section (2). Profs. Gies and Howe, 
Drs. Smith and Goodridge, and Messrs. Miller and Knudson. 

NUTRITION (PHYSIOLOGICAL AND PATHOLOGICAL CHEMISTRY) 

101-102. General physiological chemistry. A course in the eletnents of 
normal nutrition. (Teachers College, School of Practical Arts.) L, 2 hr. R. i 
hr., each section (2). Lw, 5 hr., each section (2). Prof. Gies, Dr. Seaman and 
Misses Wickwire and Harkey. (This course is designated "Chemistry 51" and 
"Household Arts Education 125" in the Teachers College Announcement.) 

This course is designated " 5" — H. A. 25 " in the Teachers College Division 
of the Summer School Announcement. The course was given last summer by 
Prof. Gies, Dr. Seaman and Miss Shaffer. 

104. General pathological chemistry. Lectures on nutrition in disease. 
(Teachers College, School of Practical Arts.) L, i hr. Prof. Gies. (This 
course is designated "Chemistry 52" in the Teachers College Announcement.) 

iio (52). General physiological chemistry. A course in the elements of 
normal nutrition. (Required of first year students of medicine.) L, 2 hr. R. i 
hr., each section (2). Lw, 6 hr., each section (2). Profs. Gies and Howe, 
Dr. Smith, and Messrs. Miller and Knudson. 

This course is designated " S — 104" in the Medical Division of the Summer 
School Announcement. It was given last summer by Prof. Gies and Dr. Smith. 

209-210. Chemistry of nutrition. (School of Pharmacy. Required of 
candidates for the Degree of Doctor of Pharmacy.) L, i hr. Prof. Gies. 

211-212. General biological chemistry. Specially adapted to the needs 
of secondary school teachers of biology. L, i hr. Lw, 4 hr, Dr. Eddy, 

213-214. Advanced physiological chemistry, including methods of re- 
SEARCH in nutrition. (Teachers College, School of Practical Arts.) L, i hr. 
Lw, 5 hr. Prof. Gies and Dr. Seaman. (This course is designated " House- 
hold Arts Education 127" in the Teachers College Announcement.) 

215. General biological chemistry. A course in the elements of normal 
nutrition. L, i hr. Lw, 7 hr. Prof. Gies, Dr. Lothrop and Messrs. Miller and 
Knudson. 

217-218. BiOCHEMICAL methods of RESEARCH, INCLUDING CLINICAL METHODS 

AND URiNARY ANALYsis IN GENERAL. L, I hr. Lw, 7 hr. Profs. Gies and Howe, 
Dr. Lothrop, and Messrs. Miller and Hepburn. 

219-220. Nutrition in health. A laboratory course in advanced physio- 
logical chemistry. L, 2 hr. Lw, 14 hr. Profs. Gies and Howe, Dr. Lothrop and 
Mr. Miller. 



Courses in Nutrition (continued) 

221-222, Nutrition in Disease. A laboratory course in advanced patholog- 
ical chemistry. L, 2 hr. Lw, 14 lir. Prof. Gies. 

223-224. Nutrition in Disease. L, i hr. Profs. Gics and Howe, and Drs. 
Fostcr, Moscnthal and Goodridge. 

225-226. Advanced physiological and pathological chemistry, including 
ALL PHASEs OF NUTRITION. Research. C, I hr. (individual students). Lw, 16 hr. 
Profs. Gies and Howe, and Dr. Lothrop. 

TOXICOLOGY 
231-232. Effects and detection of poisons, including food preservatives 
AND adulterants. Lw, 6 hr. Prof. Gies and Mr. Miller. 

BOTANY 

235-236. Chemical PHYSiOLOGY OF PLANTS. (New York Botanical Garden.) 
L, I hr. Lw, 5 hr. Prof. Gies. 

BACTERIOLOGY 

241-242. Chemistry of microorganisms : fermentations, putrefactions 
AND tue behavior of enzymes. An introduction to sanitary chemistry. L, i hr. 
Lw, 7 hr. Prof. Gies. 

SANITATION 
105. Sanitary chemistry. (Teachers College, School of Practica! Arts). 
L, i hr. Lw, 3 hr. Dr. Seaman and Miss Harkey. (This course is designated 
" Chemistry 57 " and " Household Arts Education 129 " in the Teachers College 
Announcement.) 

BIOCHEMICAL SEMINAR 
301-302. Biochemical Seminar, i hr. Prof. Gies. 

RESEARCH IN BIOLOGICAL CHEMISTRY 
Bloch emical research may be conducted, by advanced workers, independently 
or under guidance, in any of the departmcntal laboratories. 

LABORATORIES FOR ADVANCED WORK IN BIOCHEMISTRY 
The laboratories in which the advanced work of the biochemical department 
is conducted are situated at the College of Physicians and Surgeons, Teachers 
College, New York Botanical Garden and Bellevue Hospital. Each laboratory 
is well equipped for research in nutrition and all other phases of biological 
chemistry. 

BIOCHEMICAL LIBRARY 
Prof. Gies' library occupies a room adjoining the main biochemical labora- 
tory at the College of Physicians and Surgeons and is accessible, by appoint- 
ment, to all past and prosent workers in the Department. 

COLUMBIA UNIVERSITY BIOCHEMICAL ASSOCIATION 

The Biochemical Association holds scientific meetings regularly on the first 
Fridays in December, February and April, and on the first Monday in June. 
These meetings are open to all students in the University. 

SUMMER SCHOOL COURSES 
Summer Session courses are mentioned in the foregoing rcferences to 
Courses 101-102 and 110 (52). Prof. Gies will have charge of both courses next 
Summer. He will also conduct a special kcture course in nutrition. The labora- 
tories will be open for research throughout the summer. 



ANNOUNCEMENTS 

Professional Assistance Offered to Biological Chemists 

The Columbia University Biochemical Association will be glad to 
cooperate confidentially with all who desire the Services of biological 
chemists and with all who seek positions in biological chemistry. 

Address inquiries to William J. Gies, 437 West 5Qth St., New York. 



Seventh Annual Meeting of the American Biochemical Society. 

The seventh annual meeting of the American Society of Biological 
Chemists will be held in the buildings of the Medical Department of the 
Western Reserve University, Cleveland, Ohio, on Monday, Tuesday 
and Wednesday, December 30, 31, 1912, and January i, 1913. The 
American Physiological Society and the American Society for Pharma- 
cologv' and Experimental Therapeutics meet in Cleveland at the same 
time, and Joint sessions will be held. The headquarters of the three 
societies will be at the Hotel Colonial. For particulars, address the 
Secretary, Prof. Alfred N. Richards, Department of Pharmacology, 
School of Medicine, University of Pennsylvania, Philadelphia. 



THE BIOCHEMICAL BULLETIN 

The BiocHEMiCAL Bulletin is a quarterly biochemical review. 
It publishes results of original investigations in biological chemistry 
and presents miscellaneous items of personal and professional in- 
terest to chemical biologists. 

Biological chemists everywhere are cordially invited to forward 
contributions of any character whatever that will increase the v^lue 
and add to the interest of the Bulletin. Original contributions 
to research, preliminary reports of investigations, abstracts of 
papers, addresses, reviews, descriptions of new methods and appa- 
ratus, practical suggestions to teachers, biographical notes, histori- 
cal summaries, bibliographies; quotations, newS items, personalia, 
views on current events in chemical biolog}^ efc./are solicited.' 

The Bulletin will present as much biochemical substance öf as 
great variety and value, in,as Uttle space and for as little.money, as 
possible. Contributors afe accordingly reqiiested to keep their 
papers within the bounds of 15 printed pages and, if possible, to 
restrict them to^ lo^pages or less. Recrystaljize.all^the produots, 
reject the " mother liquors " and send the Biochemtcal Bulletin 
" preparations of tested piirity"! 

Each volume of the Biochemical Bulletin will contäin about 
600 pages, The price of Volume I is $6.00. Sirigle numbers may 
be purchased from a diminishing reserve supply at the following 
prices: No. i, $1.50; No. 2, $2.50; No. 3, $2.00; No. 4, $1.50. 

The reserve supply of copies of each number of Volume II df the 
Biochemical Bulletin will be closely restricted to the if}äicated 
desires of subscribers and the estimated needs of future members of 
the Association. Subscriptions for Volume II are accordingly so- 
licited at the following rates, payahle in advance: 

Before October i, 1912: domestic, $2.75; foreign, $3.00. 

Between October i and December i : $3.00; foreign, $3.25. 

Between Dec. i, 1912, and March i, 1913 : $3.50; foreign, $3.75. 

Between March i and June i, 1913: $4.25; foreign, $4.50. 

After June i, 1913 : $6.00 — to be sold only to subscribers. 

Subscriptions may be renewed at any time'on original terms. 

Remittances, manuscripts and correspondence should be addressed 
to the Biochemical Bulletin, 437 West 59th St., New York City. 



Vol. II January, 1913 No. 6 

Biochemical Bulletin 

Edited, for the Columbia University Biochemical Association, by the 

EDITORIAL COMMITTEE: 

ALFRED P. LOTHROP, Chairman, 

PAUL E. HOWE, Secretary, WILLIAM J. GIES, Treasurer, 

WALTER H. EDDY, JOSEPH S. HEPBURN, H. O. MOSENTHAL, 

NELLIS B. FOSTER, MAX KAHN, EMILY C. SEAMAN, 

F. G. GOODRIDGE, ARTHUR KNUDSON, ETHEL WICKWIRE, 

TULA L. HARKEY, EDGAR G. MILLER, JR., LOUIS E. WISE, 

all of the Staff of the Biochemical Department of Columbia University. 



CONTENTS 

PAGB 

Carl L. Alsberg. Biography and bibliography (with portrait). H. M. A 211 

A Differential Chemical Study of Glucoses from a Case of Pancreatic 

Diabetes. Frederic Landolph 217 

The Detection of Aceto-acetic Acid by Sodiu.m Nitroprussid and Ammonia. 

V. J. Harding and R. F. Ritt tan. 223 
Ortho-tolidin AS an Indicator for Occult Blood. 

R. F. Ruttan and R. H. M. Hardisty. 225 

Synthetical Properties of Emulsin. Vernon K. Krieble 227 

On the Occurrence of Nicotinic Acid in Rice Bran . U. Suztcki and S. Matsunaga 228 
A Study of the Influence of Cancer Extracts on the Groavth of Lupin 

Seedlings. Jacob Rosenbloom 229 

The Biochemistry of the Female Genitalia : 

3. A quantitative study of certain enzymes of the ovary, utenis, and bladder, 

of pregnant and non-pregnant sheep. 

Thuisco A. Erpf-Lefkovics and Jacob Rosenbloom 233 

4. On the absence of certain enzymes from the human chorion. 

Jacob Rosenbloom 236 
A Department of Biochemical Research at Vineland, New Jersey. 

Arnos W. Peters 238 

Biochemistry in New YorkTwenty Years Ago. E. E. Smith 243 

Immunity IN SOME OF ITS BIOCHEMICAL AsPECTS. C/iarles Frederick Bolduan 247 

A Plan for the Organization of the American Biological Society. 

Albert P. Mathews 261 
Organization of the Federation of American Societies for Experimental 
BiOLOGY, comprising THE American Phvsiological Society, American So- 
ciety of Biological Chemists, and American Society for Pharmacology 

and Experimental Therapeutics. John Auer. 269 

Annual Meetings of the Organizations comprisikgthe Federation of Ameri- 
can Societies for Experimental Biology: 
i. The American Physiological Society. Joseph Erlanger, Acting Secretary 271 

2. The American Society of Biological Chemists. Alfred A\ Richards, Secretary 275 

3. The American Society for Pharmacology and Experimental Therapeutics 

John Auer, Secretary 279 
Meeting of the American Society of Animal Nutrition (American Society of 

Animal Production). Lewis IV. Fetser 282 

Proceedings of the Eighth Scientific Meeting of the Columbia'University 

Biochemical Association. Alfred P. Lothrop, Secretary 284 

Folio Microbiologica. C. A. Pekelharing 297 

Biochemical Bibliography and Index. William J. des 298 

Biochemical News, Notes and Comment ; 307 

Editorials 329 



NEW YORK 

Columbia University Biochemical Association. 

Entered as second-clasB matter in the Post Office at Lancaster, Fa. 



Honorary Members of the Columbia Biochemical Association 

PROF. R. H. CHITTENDEN, First Director of the Columbia University De- 
partment of Biological (Physiological) Chemistry; Director of the Shef- 
field Scientific School of Yale University 

PROF. SAMUEL W. LAMBERT, Dean of the Columbia University School of 
Medicine 

DR. JACQUES LOEB, Memher of the Rockefeiler Institute for Medical Re- 
search; Head of the Department of Experimental Diology 

PROF. ALEXANDER SMITH, Head of the Department of Chemistry, Co- 
lumbia University 

Corresponding Members 

PROF. LEON ASHER, University of Bern. Siuitserland 
PROF. FILIPPO BOTTAZZI, University of Naples, Italy 
PROF. VLADIMIR S. GULEVIC, University of Moscow, Rtissia ' 
PROF. W. D. HALLIBURTON, King's College, London 
PROF. S. G. HEDIN, University of Upsala, Sweden 
PROF. FREDERICO LANDOLPH, University of La Plata, Argentina 
PROF. A. B. MACALLUM, University of Toronto, Canada 
PROF. C. A. PEKELHARING, University of Utrecht, Holland 
PROF. S. P. L. SÖRENSEN, Carlsberg Laboratory, C'openhagcn, Denmark 

k 

Editors of the Biochemical Bulletin 

The editorial committee 

The honorary members 

The corresponding members 

SPECIAL CONTRIBUTORS 

DR. JOHN AUER, Rockefeller Institute for Medical Research 

PROF. WILDER D. BANCROFT, Cornell University, Ithaca 

DR. CHARLES A. DOREMUS, 55 W. 52d St., New York City 

PROF. JOSEPH ERLANGER, Washington Univ. Medical School, St. Louis 

DR. LEWIS W. FETZER, U. S. Dep't of Agriculture, Washington, D. C. 

PROF. MARTIN H. FISCHER, University of Cincinnati 

DR. MARY LOUISE FOSTER, Smith College, Northampton, Mass. 

DR. V. J. HARDING, McGill University, Montreal, Canada 

DR. R. H. M. HARDISTY, McGill University, Montreal, Canada 

DR. VERNON K. KRIEBLE, McGill University, Montreal, Canada 

PROF. FRANCIS E. LLOYD, McGill University, Montreal, Canada 

PROF. JOHN A. MANDEL, A^. Y. Univ. and Bellevue Hospital Med. College 

PROF. ALBERT P. MATHEWS, University of Chicago 

PROF. SHINNOSUKE MATSUNAGA, University of Tokyo, Japan 

PROF. LAFAYETTE B. MENDEL, Yale University 

PROF. VICTOR C. MYERS, N. Y. Post-Graduate Med. School and Hospital 

DR. AMOS W. PETERS, The Training School, Vineland, N. J. 

PROF. R. F. RUTTAN, McGill University, Montreal, Canada 

DR. E. E. SMITH, 50 East 4ist St., New York City 

DR. A. E. SPAAR,_Cjfy Hospital, Trincomalee, Ceylon 

PROF. UMETARO SUZUKI, University of Tokyo, Japan 

MISS ANNA W. WILLIAMS, University of Illinois, Urbana, III. 

PROF. E. WINTERSTEIN, Polytechnic Institute, Zürich, Switserland 

DR. JULES WOLFF, 26 Rue Dutot. Paris 



'■■^ 



ASSOCIATE EDITORS 

DAVID ALPERIN, Eclectic Medical College 

EDGAR ALTENBURG, Department of Botany, Columbia University 

HUGH AUCHINCLOSS, Department of Surgery, Columbia University 

GEORGE BAEHR, Mount Sinai Hospital 

ELMER W. BAKER, Flushing Hospital 

CHARLES W. BALLARD, College of Pharmacy, Columbia University 

HANS G. BAUMGARD, German Hospital Dispensary 

CORA J. BECKWITH, Department of Zoology, Columbia University 

STANLEY R. BENEDICT, Cornell University Medical College 

LOUIS E. BISCH, N. Y. P osi-Graduate Medical School and Hospital 

HELENE M. BOAS, Barnard College, Columbia University 

CHARLES F. BOLDUAN, Health Department of New York City 

SAMUEL BOOKMAN, Mount Sinai Hospital 

SIDNEY BORN, Department of Chemistry, Columbia University 

WILLIAM BALLANTINE BOYD, College of the City of New York 

EDWARD- C BRENNER, Bellevue Hospital 

JACOB J. BRONFENBRENNER, Rockef eller Institute for Medical Research 

ALFRED J. BROWN, Department of Anatomy, Columbia University 

LEO BUERGER, Mt. Sinai Hospital 

JESSE G. M. BULLOWA, New York Polyclinic Medical School 

GERTRUDE S. BURLINGHAM, Eastern District High School, Brooklyn 

RUSSELL BURTON-OPITZ, Department of Physiology, Columbia University 

A. M. BUSWELL, Department of Chemistry,' Columbia University 

R. F. CALVERT, Department of Chemistry, Columbia University 

HERBERT S. CARTER, Presbyterian Hospital 

RUSSELL L. CECIL, Presbyterian Hospital 

ARTHUR F. CHACE, New York Post-Graduate Medical School 

ELLA H. CLARK, Barnard College, Columbia University 

ERNEST D. CLARK, Cornell University Medical College 

F. MORRIS CLASS, Vanderbilt Clinic, Columbia University 

HARVEY B. CLOUGH, High School of Commerce 

ALFRED E, COHN, Rockef eller Institute for Medical Research 

EDWARD M. COLIE, Jr., Bellevue Hospital 

BURRILL B. CROHN, Mt. Sinai Hospital 

LOUIS J. CURTMAN, College of the City of New York 

EDWARD CUSSLER, Department of Clinical Pathology, Columbia University 

CHESTER A. DARLING, Department of Botany, Columbia University 

WILLIAM DARR ACH, Department of Surgery, Columbia University 

NORMAN E. DITMAN, St. Luke's Hospital 

WALTER J. DONVAN, Commercial High School, Brooklyn 

GEORGE DRAPER, Hospital of the Rockef eller Institute 

JAMES G. DWYER, Department of Bacteriology, Columbia University 

GUSTAVE EGLOFF, Department of Chemistry, Columbia University 

FRANK R. ELDER, Department of Chemistry, Columbia University 

LEOPOLD L. FALKE, 5316 Thirteenth Ave'., Brooklyn 

BENJAMIN G. FEINBERG, College of the City of New York 

RUTH S. FINCH, Barnard College, Columbia University 

HARRY L. FISHER, Department of Chemistrv, Columbia Univer.Hty 

SIMON S. FRIEDMAN, Mt. Sinai Hospital 

C. STUART GAGER, Brooklyn Botanic Garden 

HELEN GAVIN, Wadleigh High School 

SAMUEL GITLOW, Lebanon Hospital Dispensary 

A. J. GOLDFARB, College of the City of New York 

DONALD GORDON, Department of Physiology, Columbia University 

MARK J. GOTTLIEB, Lebanon Hospital 



Associate editors (continued) 

ISIDOR GREENWALD, Montefiore Home Laboratory 

JAMES C. GREEN WA Y, New York Hospital 

LOUISE HOYT GREGORY, Barnard College, Columbia Uiiiversity 

ABRAHAM GROSS, Arbuckle Sugar Co., Brooklyn 

BEATRIX H. GROSS, N. Y. City Normal College 

BENJAMIN C. GRUENBERG, Brooklyn Commercial High School 

MARSTON L. HAMLIN, Harriman Research Laboratory, Roosevelt Hospital 

FREDERIC M. HANES, Rockefeller Institute for Medical Research 

FRED W. HARTWELL, High School of Commerce 

JOHN D. HASEMAN, Department of Zoology, Columbia University 

HAROLD M. HAYS, New York Eye and Ear Infirmary 

MICHAEL HEIDELBERGER, Rockefeller Institute for Medical Research 

ALFRED M. HELLMAN, German Hospital 

MELVIN G. HERZFELD, German Hospital 

ALFRED F. HESS, Health Department of New York City 

NELLIE P. HEWINS, Newtown High School, L. I. 

ELLA A. HOLMES, Jamaica High School, L. I. 

FRANK T. HUGHES, Boys High School, Brooklyn 

FREDERICK B. HUMPHRIES, German Hospital 

LOUIS HUSSAKOF, American Museum of Natural History 

PETER IRVING, Department of Clinical Pathology, Columbia University 

HENRY H. JANEWAY, City Hospital, Nezv York 

DAVID J. KALISKI, Mt. Sinai ^Hospital 

JOHN L. KANTOR, Mt. Sinai Hospital 

EDWARD C. KENDALL, St. Luke's Hospital 

LEO KESSEL, Mt. Sinai Hospital 

ROLFE KINGSLEY, Department of Surgery, Columbia University 

ISRAEL J. KLIGLER, American Museum of Natural History 

CLINTON B. KNAPP, General Memorial Hospital 

ALFRED H. KRÖPFE, Hoffman and Kropff Chemical Co., Brooklyn 

ELSIE A. KUPFER, Wadleigh High School 

ADRIAN VAN S. LAMBERT, Department of Surgery, Columbia University 

MARGUERITE T. LEE, Girls High School, Brooklyn 

CHARLES C. LIEB, Department of Pharmacology, Columbia University 

MABEL C. LITTLE, New York Polyclinic Hospital 

RALPH W. LOBENSTINE, Bellevue Hospital 

DANIEL R. LUCAS, St. Joseph's Hospital 

CHESTER A. MATHEWSON, Brooklyn Training School for Teachers 

LAURA I. MATTOON, Vettin School, i6o W. 74th Street 

WILLIAM H. McCASTLINE, University Physician, Columbia University 

MARY G. McCORMICK, Teachers College, Columbia University 

MRS. ELLEN BEERS McGOWAN, Teachers College, Columbia University 

GUSTAVE M. MEYER, Rockefeller Institute for Medical Research 

JESSIE A. MOORE, Loomis Laboratory, Cornell University Medical College 

HERMANN J. MULLER, Cornell University Medical College 

B. S. OPPENHEIMER, Department of Pathology, Columbia University 

RAYMOND C. OSBURN, New York Aquarium 

REUBEN OTTENBERG, Mt. Sinai Hospital 

CHARLES PACKARD, Department of Zoology, Columbia University 

ALWIN M. PAPPENHEIMER, Department of Pathology, Columbia University 

LOUIS PI NE. City Hospital. Blackwell's Island 

P. W. PUNNETT, Dep't of Chemistry, N. Y. Univ. and Bell. Hosp. Med. Coli. 



Associate editors (continued) 
ABRAHAM RAVICH, Jewish Hospital, Brooklyn 
ANTON R. ROSE, Tiirck Institute, 428 Lafayette Street 
HELEN G. RUSSELL, IVadleigh High School 
CHARLES H. SANFORD, German Hospital 
WINFIELD S. SCHLEY, St. Luke's Hospital 
OSCAR M. SCHLOSS, New York Nursery and Child's Hospital 
MAX SCHULMAN, Department of Applied Therapeutics, Columbia University 
H. VON W. SCHULTE, Department of Anatomy, Columbia University 
FRED J. SEAVER, New York Botanical Garden 

LEANDER H. SHEARER, Department of Physiology, Columbia University 
JAMES B. SIDBURY, Roosevelt Hospital 
CHARLES HENDEE SMITH, St. Luke's Hospital 
MORRIS STARK, Babies Hospital 
MATTHEW STEEL, Long Island Medical College 
RALPH G. STILLMAN, New York Hospital 
CHARLES R. STOCKARD, Cornell University Medical College 
EDWARD C. STONE, Department of Chemistry, Columbia University 
ARTHUR W. SWANN, Presbyterian Hospital 

WM. K. TERRIBERRY, Department of Physiology, Columbia University 
A. W. THOMAS, Department of Chemistry, Columbia University 
F. T. VAN BEUREN, Jr., Department of Surgery, Columbia University 
GEORGE W. VANDEGRIFT, Cornell University Medical College 
SADIE B. VANDERBILT, Teachers College, Columbia University 
PHILIP VAN INGEN, Medical Director, N. Y. Milk Committee 
CHARLES H. VOSBURGH, Jamaica High School 
WILBUR WARD, Department of Gynecology, Columbia University 
HELEN S. WATT, Wadleigh High School 
WILLIAM WEINBERGER, Lebanon Hospital 
FRED S. WEINGARTEN, German Hospital 

JULIUS W. WEINSTEIN, Vanderbilt Clinic, Columbia University 
HARRY WESSLER, Mt. Sinai Hospital 

H. B. WILCOX, Department of Diseases of Children, Columbia University 
WILLIAM H. WOGLOM, Dep't of Cancer Research, Columbia University 
I. OGDEN WOODRUFF, Department of Mediane, Columbia University 
(Local members of the Columbia University Biochemical Association) 

ASSISTANT EDITORS 

HERMAN M. ADLER, Psychopathie Hospital, Boston, Mass. 

JOHN S. ADRIANCE, Williams College. Williamstoivn, Mass. 

CARL L. ALSBERG, Bureau of Chemistry, U. S. Dep't of Agricultnre 

D. B. ARMSTRONG, Massachusetts Institute of Technology, Boston 

LOUIS BAUMANN, University Hospital, Iowa City, Iowa 

GEORGE D. BEAL, University of Illinois, Urbana, III. 

WILLIAM N. BERG, Bureau of Animal Industry, U. S. Dep't of Agriculture 

JOSEPHINE T. BERRY, State College, Pullman, Washington 

ISABEL BEVIER, University of Illinois, Urbana, III. 

A. RICHARD BLISS, Birmingham Medical College, Birmingham, Ala. 

JEAN BROADHURST, Cornell University, Ithaca, N. Y. 

MARY L. CHASE, Ingleside School, New Milford, Conn. 

WILLIAM D. CUTTER, Medical College of Georgia, Augusta, Ca. 

HAZEL DONHAM, High School, Passaic, N. J. 

A. D. EMMETT, University of Illinois, Urbana, III. 

ALLAN C. EUSTIS, Tulane University, New Orleans, La. 



Assistant editors (continued) 

KATHARINE A. FISHER, MacDonald College, Quebec, Canada 

MARY E. GEARING, üniversity of Texas, Austin, Texas 

GEORGE A. GEIGER. Marcus Hook, Pa. 

H. D. GOODALE, Mass. Agrictiltural College, Amhcrst, Mass. 

R. A. GORTNER, Carnegie Sta'n for Exp. Evolu'tn, Cold Spring Harbor, L. I. 

R. F. HARE, New Mex. Coli, of Agric. and Mech. Arts, Agric. College, N. M. 

BLANCHE R. HARRIS, State Normal School, Trtiro, Nova Scotia 

CONSTANCE C. HART, Nezv Bedford Indnstrial School, Nexv Bedford, Mass. 

E. NEWTON HARVEY, Princeton Üniversity, Princeton, N. J. 

P. B. HAWK, Jefferson Medical College, Philadelphia 

WILLIAM T. HÖRNE, Üniversity of California. Berkeley. Cal. 

HOMER D. HOUSE, Forest School, Biltmore, N. C. 

ROSCOE R. HYDE, State Normal School, Indiana 

CA VALIER H. JOÜET, Roselle, N. J. 

J. E. KIRKWOOD, üniversity of Montana, Missoula, Mont. 

MATHILDE KOCH, Üniversity of Chicago, Chicago, III. 

W. M. KRAUS, Johns Hopkins Medical School, Baltimore, Md. 

SIDNEY LIEBOVITZ, Hermon High School, Hermon, N. Y. 

BURTON E. LIVINGSTON, Johns Hopkins üniversity. Baltimore. Md. 

LOUISE McDANELL, State College, Pnllman, IVash. 

J. P. McKELVY, Allegheny General Hospital, Pittsburgh. Pa. 

H. A. MATTILL, üniversity of Utah, Salt Lake City, Utah 

CLÄREN CE E. MAY, Indiana üniversity, Bloomington, Ind. 

L. D. MEAD, Isolation Hospital. San Francisco, Cal. 

CLARA G. MILLER, Knox School, Tarrytown, N. Y. 

MAX W. MORSE, Trinity College, Hartford, Conn. 

EDWARDS A. PARK, Johns Hopkins Medical School 

OLIVE G. PATTERSON, Toronto üniversity, Toronto. Canada 

W. H. PETERSON. üniversity of Wisconsin, Madison, Wis. 

HELENE M. POPE, High School, Passaic, N. J. 

E. R. POSNER, Brake üniversity Medical School, Des Maines, la. 

DAVID F. RENSHAW, M'est High School, Kochester, N. Y. 

ALFRED N. RICHARDS, üniversity of Pennsylvania, Philadelphia 

ANNA E. RICHARDSON, üniversity of Texas, Austin 

L. A. ROBINSON, üniversity of Porto Rico, Las Pietras 

WINIFRED J. ROBINSON, Vassar College, Poughkeepsie, N. Y. 

JACOB ROSENBLOOM, üniversity of Pittsburgh 

WILLIAM SALANT, Bureau of Chemistry, U. S. Department of Agriculture 

CARL A. SCHWARZE, A^. /. Agric. Experiment Station, New Brunswick, N. J. 

FREDERICK W. SCHWARTZ, Rensselaer Polytechnic Institute, Troy, N. Y. 

A. D. SELBY, Ohio Agricultural Experiment Station, Wooster, Ohio 

BLANCHE E. SHAFFER, North Te.vas State Normal School, Benton, Te.ras 

A. FRANKLIN SHULL, üniversity of Michigan, Ann Arbor, Mich. 

CLAYTON S. SMITH, Bureau of Chemistry, ü. S. Department of Agriculture 

EDWARD A. SPITZKA, Jefferson Medical College. Philadelphia 

MARY E. SWEENY, üniversity of Kentucky, Lexington, Ky. 

WILLIAM A. TALTAVALL, Redlands, Cal. 

DAVID C. TWICHELL, Saranac Lake, N. Y. 

IDA C. WADSWORTH, Brockport State Normal School, Brockport, N. Y. 

EDWIN D. WATKINS, Medical School, üniv. of Tenn., Memphis, Tenn. 

WILLIAM H. WELKER, Red Hill, Pa. 

DAVID D. WHITNEY, Wesleyan üniversity, Middletown, Conn. 

LORANDE LOSS WOODRUFF, Yale üniversity, New Haven, Conn. 

HAROLD E. WOODWARD, ü. S. Food and Drug Inspection Lab'y, Philadelphia 

R. M. YERGASON, Trinity College, Hartford, Conn. 

HANS ZINSSER, Leland Stanford üniversity, Palo AUo, Cal. 

(Non-resident members of the Columbia üniversity Biochemical Association^ 




CjOaL 



U. %^^ 



BiocHEMiCAL Bulletin 



Volume II JANUARY, 1913 No. 6 

CARL L. ALSBERG 

Chief of the Bureau of Chemistry of the U. S. Department of 

Agriculture 

Carl Lucas Aisberg was born April 2, 1877, in New York City, 
son of Bertha (Baruch) and Meinhard Aisberg. From early child- 
hood Aisberg evinced an interest in natural science, especially 
biology. This was partly due to traiining and environment, and 
partly to a natural inclination. 

His father, a chemist of distinction and one of the first to manu- 
facture organic dye stuffs in this country, was graduated (Ph.D.) 
from the University of Jena, and trained under Wöhler (Uni- 
versity of Göttingen), Bunsen (University of Marburg), and 
Geuther (University of Jena). He was assistant to Professor 
Geuther ät Jena and Privatdozent in chemistry at that University. 
About 1865 he became assistant to Prof. Chandler in the School of 
Mines at Columbia College. Later he was chemist to the New 
York City Board of Health under Prof. Chandler. Subsequently 
he was occupied in chemical manufacture and chemical engineering. 
He died in 1897. He was one of the founders and first Secretary 
of the New York Chemical Society, from which the American 
Chemical Society developed. He had given up the academic career, 
in which he had unusual prospects, in order to support his wife and 
parents. But the spirit of research continued to influence the 
father, and he never ceased to have an active interest in the purely 
scientific side of chemistry This point of view was maintained in 
spite of a very active and rapidly growing business, and at a time 
r when research in theoretical science did not receive the recognition 

o in this country that later was conceded to it. It had an effect in 

211 



Q;: 
■er: 



212 Carl L. Aisberg [Jan. 

molding the growing mind of his son, which never could have been 
obtained otherwise. 

With a true appreciation, however, of the values o£ a liberal 
education, this scientific interest was not allowed to exert a narrow- 
ing effect.^ It was not until Aisberg entered the College of Physi- 
cians and Snrgeons of Columbia University, however, that he 
allowed himself to devote his entire time to scientific work He 
graduated with the degree of M.D. in 1900, and received at the 
same time the degree of A.M. for special research in physiology.^ 
During the summers and vacations of the Medical School, Aisberg 
devoted his time to research in physiology with Professor F. S. Lee, 
and in biological chemistry with Dr. P. A. Levene, now of the 
Rockefeller Institute of Medical Research, then associate in chem- 
istry at the Pathological Institute of the New York State Hospitals. 

In July, 1900, after graduating from the Columbia Medical 
School, Aisberg went to Strassburg, Germany, for post-graduate 
w^ork, where he studied pharmacology under Schmiedeberg, physio- 
logical chemistry under Hofmeister, and clinical medicine under 
Naunyn. Here he also was associated with E. S. Faust, then 
Privatdozent at the Pharmacological Institute, now professor of 
pharmacology at Würzburg; also with Wolfgang Heubner, and 
others. During the succeeding two years his time was devoted 
almost exclusively to biological chemistry, notably pharmacology, 
but he continued his interest in clinical and general biological 
matters, for the study of which there were unusually good oppor- 
tunities at the University of Strassburg. During this time he con- 
ducted special investigations into the structure and biological signifi- 

^ C. L. Aisberg was prepared for College by tutors and at the Mt. Morris 
Latin School, entering Columbia College in 1892. He received the degree of 
A.B. in 1896. 

^The Department of Physiological Chemistry in the Columbia Medical 
School was founded in 1898-99, during Alsberg's third year there. At that 
time only one course in physiological chemistry was offered and that was 
required of " second year men " in medicine. Alsberg's early interest in physio- 
logical chemistry was shown by the fact that, while a " third year man " in good 
Standing at the Medical School, he took, as an elective, the newly established 
course in that subject for second year men — something no other third year 
medical Student attempted, then or since. The records show that in spite of this 
heavy addition to his regulär work, Aisberg stood among the very highest in 
physiological chemistry and in the entire medical course. [Ed.] 



I9I3] H. M. A. 213 

cance of the nucleic acids. He went to Berlin in 1901 where he 
spent a year in chemistry with Emil Fischer and in plant physiology 
with Kny. One vacation was spent at Frankfurt a/M with Ehrlich, 
Weigert, Edinger and C. von Noorden, in studies especially of the 
side-chain theory and other conceptions of immunity ; another vaca- 
tion was devoted to clinical medicine with Kuttner, Piorkowski and 
others. 

In the fall of 1902 Aisberg returned to this country, to accept 
the Position of assistant in physiological chemistry at the Harvard 
Aiedical School. In 1905 he was advanced to instructor in bio- 
logical chemistry and put in charge of the Organization of the 
Department of Biological Chemistry at the new Harvard Medical 
School. From 1906 to 1908 he was in charge there (jointly with 
L. J. Henderson) of the teaching and research in biological chem- 
istry. From 1907 to 1908 he conducted, in addition, special in- 
vestigations for the U. S. Bureau of- Fisheries, at Woods Hole, 
Mass. 

While at Harvard, Aisberg not only organized and developed an 
efficient and unusual department for undergraduate teaching, but 
also, as head of the department, put on a firm basis, for the first 
time in that institution, a System of graduate instruction and re- 
search in biological chemistry. 

Alsberg's success as a teacher, both of undergraduates and 
graduates, has been appreciated by all who have come in contact 
with him. In fact, it has been recognized by miany that his gift in 
this direction is so pronounced that they have repeatedly urged him 
to devote himself exclusively to teaching. But the strong spirit of 
research, coupled with his broad biological interests, would not 
permit him to confine himself to teaching, and when, in 1908, the 
Position of chemical biologist, in charge of the Poisonous-Plant 
Laboratory of the Bureau of Plant Industry in the U. S. Depart- 
ment of Agriculture, at Washington, was offered to him, he accepted 
it with the belief that, by freeing himself from the enticing but 
time-consuming occupation of teaching, he might accomplish more 
in research. This conclusion has been amply justified by the results 
of his investigation of poisonous plants, notably the loco weed, and 
the biochemistry of various moulds. In this connection, it may be 



214 Carl L. Aisher g [Jan. 

recalled that the investigations of spoilt corn by Aisberg and his 
co-workers have revolutionized the metbods of testing corn for its 
fitness as food. 

Aisberg was secretary of the Section on Physiological Chemistry 
of the International Congress of Arts and Sciences, St. Louis Ex- 
position (1905); also secretary and member of the Council of 
the Boston Society of Medical Sciences. He is Chairman of the 
Division of Biological Chemistry of the American Chemical Society, 
Fellovv of the American Association for the Advancement of Sci- 
ence, and member of the following societies : American Chemical 
Society, American Society of Biological Chemists, American Phys- 
iological Society, Society for Experimental Biology and Medicine, 
Society for Pharmacology and Experimental Therapeutics, Amer- 
ican Pharmaceutical Association, Washington Academy of Science, 
American Medical Association, American Association for Cancer 
Research, Corporation of the Marine Biological Laboratory at 
Wood's Hole, Massachusetts. He is one of the assistant editors of 
Chemical Abstracts. 

Alsberg's appointment, by President Taft, as chief of the Bureau 
of Chemistry to succeed Dr. Harvey W. Wiley, has received the 
endorsement of all who know him. With his training and natural 
equipments, with his record of achievements in research and in 
practical chemistry, and with his professional Standing as a scientist, 
it seems assured that the Bureau of Chemistry will continue to 
develop along the best and most approved lines of modern chemical 
science. 

A list of Alsberg's most important papers is appended : 

1901. P. A. Levene and C. L. Alsberg: Zur Chemie der Paranuclein- 
säure; Zeitschrift für physiologische Chemie, 31, 543. 

1904. C. L. Alsberg: Beiträge zur Kenntnis der Nucleinsäure; Archiv 

für experimentelle Pathologie und Pharmakologie, 51, 239. — 
C. L. Alsberg: The influence of cholic acid upon the excre- 
tion of sulphur in the urine; Journal of Medical Research, 

13, 105. 

1905. C. L. Alsberg and Otto Polin : Protein metabolism in cystin- 

uria; American Journal of Physiology, 14, 54. 

1906. P. A. Levene and C. L. Alsberg: The cleavage products of 

vitellin ; Journal of Biological Chemistry, 2, 127. 



I9I3] H. M. A. 215 

1907. C. L. Alsberg: On the occurrence of oxidative ferments in a 

melanotic tumor of the liver; Journal of Medical Research, 
16, 117. — R. Fitz, C. L. Alsberg and L. J. Henderson : Con- 
cerning the excretion of phosphoric acid during experimental 
acidosis in rabbits; American Journal of Physiology, 18, 
113. — P. A. Levene and C. L. Alsberg: Über die Hydrolyse 
der Proteine mittels verdünnter Schwefelsäure ; Biochemische 
Zeitschrift, ^,312. 

1908. C. L. Alsberg: Beiträge zur Kenntniss der Guajak-Reaktion; 

Archiv für experimentelle Pathologie und Pharmakologie, 
Supplement-Band ("Schmiedeberg-Festschrift"), p. 39. — C. 
L, Alsberg and E. D. Clark: On a globulin from the tgg- 
yolk of the spiny dog-fish, Squalus acanthias L. ; Journal of 
Biological Chemistry, 5, 243. — C. L. Alsberg and E. D. Clark : 
The blood clot of Limulus polyphemus ; Ibid., 5, 323. 

1909. C. L, Alsberg: Agricultural aspects of the pellagra problem in 

the United States; New York Medical Journal, July 10. — C. 

* 

L. Alsberg: The formation of gluconic acid by the olive- 
tubercle organism and the function of oxidation in some micro- 
organisms ; Proceedings of the Society for Experimental Biol- 
ogy and Medicine, 6, 83. — C. L. Alsberg: The globulins of 
the egg-yolk of Selachians ; Proceedings of the American 
Society of Biological Chemists, i, 160, and Journal of Biolog- 
ical Chemistry, 6, p. xiii. — C. L, Alsberg and C. Hedblom : 
Soluble chitin; Proceedings of the American Society of Bio- 
logical Chemists, i, 192, and Journal of Biological Chemistry, 
6, p. xlv. — C. L. Alsberg and C. A. Hedblom : Soluble chitin 
from Limulus polyphemus and its peculiar osmotic behavior ; 
Journal of Biological Chemistry, 6, 483. 

1910. C. L. Alsberg: Recent work in biological chemistry; Journal of 

the American Chemical Society, 32, 704. — O. F, Black and 
C. L. Alsberg: The determination of the deterioration of 
maize with incidental reference to pellagra. Bulletin ipp, 
Bureau of Plant Industry, U. S. Department of Agriculture. 
— C. L. Alsberg: Note on the use of chitin in dialysis; Pro- 
ceedings of the American Society of Biological Chemists, i, 
225, and Journal of Biological Chemistry, 7, p. xii. — C. L. 
Alsberg and E. D. Clark : The hemocyanin of Limulus poly- 
phemus; Journal of Biological Chemistry, 8, i. 

191 1. C. L. Alsberg: The toxic action of Amianthium muscaetoxicum; 



2i6 Carl L. Alsberg [Jan. 

Proceedings of the Society for Pharmacology and Experi- 
mental Therapeutics, Journal of Pharmacology and Experi- 
mental Therapeutics, 3, 473. — C. L. Alsberg: Mechanisms of 
cell activity ; Science, 34 (n. s.), 97. — C. L. Alsberg and O. F. 
Black : Biological and toxicological studies upon Penicillium 
puberulum, Bainier; Proceedings of the Society for Experi- 
mental Biology and Mediane, 9, 6, and Proceedings of the 
American Chemical Society, Biochemical Bulletin, i, 103. 
— C. L. Alsberg: The formation of (i-gluconic acid by Bac- 
terium savastoni, Smith; Journal of Biological Chemistry, 9, 
I. — C. L. Alsberg: Proceedings of the meeting of the section 
of biological chemistry of the American Chemical Society 
(Chairman's report) ; Biochemical Bulletin, i, 94. — O. F. 
Black and C. L. Alsberg: Observations on the deterioration 
of maize; Ibid., i, 130. 
1912. C. L. Alsberg and O. F. Black: Studies on barium feeding; 
Proceedings of the Society for Experimental Biology and 
Medicine, 9, 37. — C. L. Alsberg and O. F. Black ; Laboratory 
studies on the relation of barium to the loco-weed disease. 
Bulletin 246, Bureau of Plant Industry, U. S. Dept. of Agri- 
culture. — C. L. Alsberg and O. F. Black: Biochemical and 
toxicological studies on Penicillium stoloniferum, Thom ; Pro- 
ceedings of the Eighth International Congress of Applied 
Chemistry, 19, 15. 

H. M. A. 



A DIFFERENTIAL CHEMICAL STUDY OF GLUCOSES 
FROM A CASE OF PANCREATIC DIABETES^ 

FREDERIC LANDOLPH 

(Laboratory of Organic Chemistry of the University of La Plata, and the 
National Hospital of Buenos Aires, Argentina) 

My new chemical method for the differential or f ractional study 
of carbohydrates has been successfully applied to the study of the 
lactoses of milk,^ and also of the glucoses in the urine of the last 
period of cachexia in a case of diabetes.^ I have lately applied 
this method to the sugar in the urine of Louis Dufaut, a hospital 
patient for a year in ward IV of the National Hospital of Buenos 
Aires, where he was under the immediate treatment of my illustrious 
teacher and friend, Dr. Abel Ayerza.'* 

For nearly three years I have been engaged in the laborious task 
of endeavoring to fractionate the glucoses^ in the urines of this 
patient, as I have already fractionated the lactoses of milk. I frac- 
tionated the products of condensation (or perhaps alsoof decomposi- 
tion) of glucoses in urine that yielded a residue, after evaporation, 
of 83.76 grams per thousand parts of urine, a polaristrobometric 
deviation of 54° 06' per thousand, a reduction corresponding to 75 
grams of sugar per thousand, and a fermentation representing about 
60.8 grams of sugar per thousand, but obtained only a single osa- 

^Translated (and in part abstracted) by Dr. Max Kahn from the introduc- 
tion to the author's paper, in French, in the Revista de la Universidad de Buenos 
Aires, 1912, xvii, pp. 108-221, a copy of which was forwarded by Professor 
Landolph for this purpose. 

* Landolph, Argentina Medica, July 27 and August 3, 1907, and March 28. 
1908. 

^ Landolph, Revista de la Universidad de Buenos Aires, 1906, xi, pp. lOi, 
153 and 232. 

* The clinical history of this patient was published by the author in the 
Revista de la Universidad de Buenos Aires, 1912, xvii, p. 108. 

° Professor Landolph believes that diabetic urine contains a number of 
GLUCOSES differing in their fermentability, optical properties, reducing powers 
and ability to yield osazones. [Trans.] 

217 



21 8 Differential Chemical Study of Glucoses [Jan. 

Zone with a melting point of 189-190° C, equivalent to 41 grams 
of sugar per thousand. After hydrolysis^ of a large quantity of 
urinary residue, the values for polariscopic deviation, reduction, and 
fermentation were about ten tinits less than the corresponding 
figures for non-hydrolyzed urine ; and the quantity of sugar in the 
hydrolyzed urine, as represented by osazone, did not equal half 
the amount of sugar obtained from the original urine. These dif- 
ferences can be explained in several ways ; but since " diabetic 
sugar," as I have already demonstrated, is a collection of several 
distinct chemical substances, it is highly probable that in such treat- 
ment some of these components are so modified that they no longer 
form osazones, although they retain their fermentability. It is tnie 
that this explatiation does not accord zvith conventional views, but, 
nowadays, the phenoniena in chemistry and physics zvhich do not 
agree zvith current theories are the ones that shoidd be probed and 
investigated in the interest of^ triith. 

When a portion of evaporated, syrupy residue from my pa- 
tient's urine was allowed to age, there was a marked change in the 
residue, due either to the nature of the syrup or to chemical decom- 
position in it. This was not surprising in view of the complex 
composition of diabetic urine, and the further fact that the original 
sugar may undergo a process of slow hydrolysis, or rather conden- 
sation, to form higher Compounds like dextrins and analogous sub- 
stances, which then yield, with Phenylhydrazin, resinous pseudoösa- 
zones having very low melting points, such as I have isolated from 
gastric Juices.'^ 

Discussing, now, some of the details of my analytic data, I find 
that in the second treatment^ down to the fourth extraction, the 

' Hydrolysis was performed in the following way : 100 c.c. of urine, or 
urinary extract, were heated for seven hours on a water bath with 5 c.c. of 
hydrochloric acid Solution (strength not stated). The liquid was then evapo- 
rated to a volume of 60 c.c. 

^ Landolph, Revista de la Universidad de Buenos Aires, October, November 
and December, 1910. 

* Professor Landolph examined a number of urines from the same patient 
by several processes for the determination of the content of sugar. The urines 
were evaporated over a water bath to a syrupy consistency and the examina- 
tions of the separate urinary residues are called "Treatment" I, II, etc. After 
completion of the alcoholic extractions, a portion of each extract was hydrolyzed 



1913] Frederic Landolph 219 

residue,^ though always quite abundant, decreased from 68.17 grams 
per thousand parts of urine in the first to 36.26 grams in the third. 
On comparing the figures obtained from the original urine with the 
figures for the extracts of its residues, I observed polariscopic devia- 
tions which were only one-fourth, one-fifth and one-tenth as great, 
respectively, as those for the untreated urine. The disappearance of 
the aldehydic function led me to suppose that there was condensa- 
tion and not oxidation, since fermentation and reduction tests were 
still very marked and showed the presence of half or even more of 
the total dry residue. As regards the characteristic osazone, it was 
obtained only in small amounts with nearly, but not quite, the same 
melting point, i. e., instead of melting at 189-190° C. it melted at 
185-186° C. ; whereas the pseudoösazone (with a melting point of 
yS~7^° C.), which was resinous and alcohol-soluble, corresponded 
to 66.72 grams of sugar per thousand (33.36 grams of polymer- 
ized sugar). , 

Hydrolysis produced analogous changes in the urine, but here 
the triic osazone, with a melting point of 174-175° C, was repre- 
sented by the small quantity of 1.75 grams per thousand of urine 
(0.87 gram of glucosej. This, there fore, was different from the 
osazone obtained from the non-hydrolyzed material. The pseudo- 
ösazone from the hydrolyzed Solution amounted to 44.94 grams of 
sugar per thousand, or 22 units less than the pseudoösasone from 
the non-hydrolyzed Solution (melting point of 95° C. instead of 
75° C). For the fourth, fifth and sixth alcoholic extracts of the 
first treatment, analogous figures were obtained. 

In the first extract^^ of the second treatment, the degree of the 
reduction was maintained for each Observation, while the fermenta- 
bility of the non-hydrolyzed Solution became very feeble. Here the 
principal osazone was pseiidoösazone (with a melting point of 
112° C.) amountingto 38 grams per thousand of urine (17.5 grams 

with dilute hydrochloric acid Solution and the " sugar s" in the hydrolyzed and 
non-hydrolyzed portions fractionated with basic lead acetate and mercuric 
nitrate. [Trans.] 

' After the urine was evaporated over a water bath to a syrupy consistency, 
the residues were successively extracted with absolute alcohol. These " treat- 
ments " are called " extraction " i, 2, etc. 

^" After extraction with alcohol, 24.167 grams of extract were dissolved in 
500 c.c. of water. This was divided into two parts. The first part was pre- 
cipitated with basic lead acetate, the second with mercuric nitrate. 



220 Differential Chemical Study of Glucoses [Jan. 

of glucose). The tnic osazone, with a melting point o£ 177- 
178° C, amounted to only 1.74 grams per thousand of iirine (0.87 
gram of glncose). After hydrolysis^^ we obtained in three differ- 
ent fractions (12.20, 10.62 and 2.96) a total of 25.78 grams of 
osazones per thousand of urine (14.39 gi'ams of glucose). The 
melting points were respectively 170°, 177° and 157-159° C. It 
is Singular that here no pseudoösazoncs were obtained. 

For the second extract^^ of the second treatment, we obtained, 
from the non-hydrolyzed Solution, osazone with a melting point 
of 95~97° C., amounting to 11 grams per thousand of urine (5.5 
grams of sugar). Here too, after hydrolysis, we obtained two 
fractions of tnie osazone (melting points of 172-173 and 180- 
181° C.) amounting to 19.75 grams per thousand of urine (9.87 
grams of sugar) but again no pseudoösazone. 

For the third alcoholic extract of the second treatment, we ob- 
tained practically no fermentation, and only psctidoösasones. In 
the hydrolyzed Solution, however, fermentation was pronounced; 
and we obtained a mixture of true osa:^ones (melting points of 180- 
190° and 187-188° C.) amounting to 10.06 grams per thousand 
of urine (5.03 grams of sugar — 5.40 grams of fermentable sugar). 
The pseudoösazone with a melting point of 100° C. and amounting 
to 6.83 grams per thousand of urine (3.42 grams of sugar), 
corresponded, perhaps, to the difference between 7.42 grams of 
sugar per thousand of urine (polariscopic determination) and 
5.40 grams per thousand of urine (estimated by fermentation), i. e., 
2.02 grams. 

For the fourth alcoholic extract of the second treatment, we 
obtained from both the non-hydrolyzed and hydrolyzed Solutions, 
mixtures of true and pscudoösazones in approximately equal pro- 
portions. 

In the alcoholic extracts of the third treatment, fermentation in 
general was always pronounced, corresponding to an increase in the 
amounts of true osazones and pseudoösazones; but, with this 

" Hydrolysis was conducted by heating the extract with hydrochloric acid 
Solution over a water bath. See footnote 6. 

*' The residue remaining after the first alcoholic extract was again ex- 
tracted with alcohol, and also treated with mercuric nitrate and basic lead 
acetate. See footnotes 8 and 9. 



1913] Frederic Landolph 221 

anomaly, that in the sixth and seventh extracts (third treatment) 
the vveights of the sugars, as represented by the amounts of pseiido- 
ösa^ones, were three or four times greater than the weights of the 
total dry residues. 

As regards the fourth, fifth and sixth treatments/^ we obtained 
neither polariscopic deviation, reduction nor fermentation, but we 
did obtain pseudoösasones. 

We also noticed that for the fifth alcoholic extract of the second 
treatment the total dry residue obtained upon hydrolysis was always 
greater than the total dry residue of the non-hydrolyzed portion, 
evidently due to the absorption of oxygen from the air (which per- 
haps also provoked a condensation or resinification of Phenylhy- 
drazine). 

In Order to obtain an approximately correct idea of the action of 
basic lead acetate or mercuric nitrate upon the urinary glucoses, I 
treated each of the first and second extracts of the second treatment 
with basic lead acetate in one portion and mercuric nitrate in an- 
other. The lead sali precipitated nearly all of the optically active 
siigar from the non-hydrolyzed Solution, white the mercuric nitrate 
did not have any effect in this regard, but both reagents diminished 
the quantity of the reducing sugars. 

From the first fraction of the second treatment, basic lead 
acetate removed all the sugar that yielded pseudoösazones, and left 
only the sugar which formed true osazones — the latter in dimin- 
ished quantities, at least in the first and second crystallizations. 
Treatment with mercuric nitrate yielded only traces of true osasones 
but, in the second crystallization, a marked amount of pseudoösa- 
zoncs was obtained. 

From another portion of the second alcoholic extract of the 
second treatment, basic lead acetate again precipitated all the active 
sugars, while mercuric nitrate did not affect them. With mercuric 
nitrate, only pseudoösazones were obtained. 

*' Procedures similar to those previously indicated were adopted in the study 
of " treatments " IV, V and VI : The urine was evaporated in each case, the 
residue extracted with alcohol several times, and each of the extracts divided 
into two portions. One portion was hydrolyzed, the other was not. The 
hydrolyzed and non-hydrolyzed portions were individually treated in different 
portions with basic lead acetate and mercuric nitrate. See footnote 8. 



222 Differential Chemical Study of Glucoses [Jan. 

These observations show that the Isolation and identification of 
urinary sugars is a very complicated process. One must work with 
large quantities of material in order to be able to differentiate, re- 
crystallize and purify all the fractions. I am now engaged in an 
extension of this work. 



THE DETECTION OF ACETO-ACETIC ACID BY 
SODIUM NITROPRUSSID AND AMMONIA^ 

V. J. HARDING AND R. F. RUTTAN 
(Chemical Laboratory, McGill University, Montreal, Canada) 

The use o£ sodium nitroprussid and ammonia, followed by the 
addition of an acid insufficient in amount to completely neutralize 
the ammonia, was first suggested by Le Nobel as a method of de- 
tecting small quantities of acetone. This test is based on the orig- 
inal test of Legal but, as the two tests differ in result, it is proposed 
to call the test depending on the use of ammonia, the Le Nobel test, 
and to reserve the term Legal test exclusively for the action of 
sodium nitroprussid and potassium (or sodium) hydroxid followed 
by acidification. 

The two tests differ in the f ollowing points : ( i ) The Le Nobel 
test gives with acetone a much bluer shade of purple and (2) is an 
extremely delicate test for aceto-acetic acid. 

The usual way, in clinical work, of applying the Le Nobel test is 
to first acidify the urine with acetic acid, add a few drops of a dilute 
Solution of sodium nitroprussid and then overlay the Solution with 
concentrated aqueous ammonium hydroxid Solution. On applying 
this test, the authors discovered several anomalies which can be sum- 
marized as follows : 

(A) Acetone in water and when added to urine, making concen- 
trations similar to those occurring in cases of acetonuria, gives the 
test very faintly and only after long standing — ^by no means as dis- 
tinctly as the natural cases. 

(B) If some samples of urine which give a marked response to 
the Le Nobel test be distilled with acids, the test given by the dis- 
tillate (where the acetone is presumably ten to twenty times more 
concentrated than In the original urine) is very much less marked. 

* Abstract of paper published in the Bio-Chemical Journal, 1912, vi, p. 445 
(Oct). 

223 



224 Dctcction of Aceto-Acetic Acid [Jan. 

If such a urine (B) first be boiled iinder a reflux condenser in 
presence of a little acetic, oxalic or sulfuric acid, and the Le Nobel 
test aiDplied, the test gives either a negative result or the response 
is much diminished in intensity. As these urines contained aceto- 
acetic acid, which would be destroyed by heat, it was evident that the 
previous presence of this acid in the urine could account for the 
anomahes observed. That this was so was estabhshed in the fol- 
lowing way. 

The urine was acidified with oxalic acid, saturated with sodium 
Chlorid and rendered acetone-free by aspiration for an hour with a 
current of air, as in the Folin method of estimating acetone. At 
the end of that time a determination of free acetone, by the Folin 
method, showed that none was present, although the residual urine 
responded to the Le Nobel test with undiminished intensity, and 
the test became negative when the liquid was boiled for fifteen 
minutes under a reflux cond.enser. 

Aceto-acetic acid, however, is stated in the literature to give a 
faint reddish-brown or orange-red coloration with sodium nitroprus- 
sid and ammonia — unchanged by the addition of acid. To deter- 
mine this point, asolutionof aceto-acetic acid was madebyhydrolyz- 
ing ethyl aceto-acetate with the theoretical quantity of potassium 
hydroxid in the cold for twenty-four hours. This hydrolyzed Solu- 
tion was found to respond to the Le Nobel test exactly as the urine 
of an acidosis patient. The test became negative on boiling the 
Solution under a reflux condenser, and was unaff ected by the removal 
of the free acetone. 

In consequence of these facts the authors have no hesitation in 
saying that aceto-acetic acid of itself responds to the Le Nobel test 
and that, in the great majority of cases, a positive result when the 
Le Nobel test is applied to a urine indicates aceto-acetic acid and 
not acetone. On comparing the delicacy of the Le Nobel test for 
aceto-acetic acid in urine, with the ferric chlorid test, the authors 
found that the former will just detect about one part of aceto- 
acetic acid in 30,000 parts of urine, while the latter fails at i part 
in 7,000. The limit of detection of aceto-acetic acid in water by 
the Le Nobel test is over i part in 80,000. 



ORTHO-TOLIDIN AS AN INDICATOR FOR OCCULT 

BLOOD 

R. F. RUTTAN and R. H. M. HARDISTY 
(Chemical Lahoratory, McGill University, Montreal, Canada) 

The authors have lately called attention to the advantages of 
o-tolidin over benzidin and phenolphthalin as a clinical reagent for 
the detection of occult blood.^ 

The properties and derivatives of o-tolidin, 

(4) NH (I) (I) NH, (4) 
\c H r H / 

(2) CH/ \CH3 (2) 

were first described by one of the writers in 1886.2 This substance 
was compared with guaiacum, benzidin and phenolphthalin in aque- 
ous Solutions of blood and in Solutions containing urine, stomach 
Contents, and feces. It was found to be a very delicate reagent for 
the detection of blood in aqueous Solution, and to have some impor- 
tant advantages over the other clinical reagents when used in the 
detection of blood in excretions and secretions. The reagents were 
made up as follows: Guaiacum, i in 25 methylated spirits; benzidin 
and o-tolidin, in Solutions of similar strength, in glacial acetic acid ; 
phenolphthalin, prepared as recommended by Kastle.^ 

The Solutions to be tested were made up from a o.i per cent. 
Solution of cry stalline hemoglobin in water. The hydrogen per- 
oxide Solution employed was made up to approximately 3 per cent. 
from Merck's perhydrol. In testing, i c.c. of the reagent, i c.c. of 
the Solution to be tested and i c.c. of diluted perhydrol were em- 
ployed. 

In aqueous Solution, as the average of ten tests, it was found 
that guaiacum detected blood, i in 50,000 ; benzidin detected blood, i 
in 700,000; o-tolidin detected blood, i in 7,000,000; phenolphthalin 
detected blood, i in 10,000,000, or even in greater dilutions. 

^ Ruttan and Hardisty : Canadian Medical Association Journal, Nov., 1912. 
^ Ruttan : Proccedings of the British Association for the Advancement of 
Science, 1886. 

^ Kastle : Bulletin 51, Hygienic Laboratory, Washington, D. C. 

225 



226 Ortho-Tolidin as Indicator for Occidt Blood [Jan. 

Guaiaciim and benzidin, when positive, gave prompt reactions 
but in very dilute Solutions the color faded quickly. o-Tolidin de- 
veloped the greenish-blue, or deep blue, more slowly but the color 
persisted for some time, even several hours. 

The results of the comparative tests are briefly summarized 
below. 

{i) In urine: Guaiacum and benzidin detected blood, i in 6000; 
benzidin was slightly the more sensitive reagent; o-tolidin detected 
I in 24,000; phenolphthalin, less than i in 2,000. 

(2) In feces: Feces of patients on a meat-free diet for seven 
to ten days vvere used and a 2 per cent. emulsion prepared. Guaia- 
cum detected blood, i in 10,000; benzidin and o-tolidin, i in 100,- 
000, the tolidin reaction being slightly slower but persisting — the 
benzidin color fading quickly in very dilute Solutions; phenol- 
phthalin gave reactions only when dilutions did not exceed i in 
2,000. 

(3) In stomach contents: Stomach contents after ordinary test- 
meals were employed. One c.c. of stomach contents was added to 
the reagent before the diluted blood Solution was introduced. 
Guaiacum detected i in 5,000; benzidin and o-tolidin, i in 30,000; 
phenolphthalin, even after the acidity of the stomach contents had 
been neutralized before applying the Solution, was less delicate than 
guaiacum. 

Experiments were conducted to determine the keeping properties 
of the reagents. Although benzidin and o-tolidin are about equal 
in delicacy for blood in feces and stomach contents, the delicacy of 
the benzidin reagent diminishes 50 per cent. in 24-36 hours, while 
o-tolidin will remain unchanged in delicacy for from three to four 
weeks. 

o-Tolidin is as sensitive a reagent for occult blood in stomach 
contents and feces as benzidin. Its action is less inhibited by urine 
than any of the other reagents. Its Solution in acetic acid can be 
kept for one month without its delicacy being reduced. After that 
its value decreases slowly. Benzidin'* Solutions in acetic acid cannot 
be kept twenty-four hours without very serious deterioration in deli- 
cacy; some preparations decreasing over 50 per cent. 

* Three different products were compared. 



SYNTHETICAL PROPERTIES OF EMULSIN 

VERNON K. KRIEBLE 
(Chemical Laboratory, McGill University, Montreal, Canada) 

In a recent communication^ the writer described an emulsin 
which produced Isevo-mandelonitrile when allowed to act for three 
and one-half hours on an amygdalin Solution. Those experiments 
were conducted during the spring of 1910. Much to our surprise 
when the research was continued in October, 1912, it was found 
that, under the conditions previously described, the nitrile produced 
was dextro active. This seems to explain the fact that the author's 
results differed from those of Feist, Rosenthaler, and Auld, who 
found dextro-nitrile. Their samples "of emulsin were evidently 
much older than the one used by the author for his first determina- 
tions. 

It seems very probable that there are two synthetic enzymes in 
a fresh sample of emulsin, one of which synthesizes dextro-mandelo- 
nitrile from benzaldehyde and hydrocyanic acid, while the other 
synthesizes a Isevo-nitrile. The one synthesizing the dextro-nitrile 
is evidently more stable. 

Fresh emulsin was extracted from bitter and from sweet 
almonds. It was found that the sample from sweet almonds, 
when allowed to act on amygdalin for three and one-half hours, 
produced Isevo-nitrile while the one from the bitter almonds was 
dextro active. 

The detailed experimental results will appear very shortly in 
one of the chemical Journals. 

^Krieble: Journal of the American Chemical Society, 1912, xxxiv, p. 716. 



227 



ON THE OCCURRENCE OF NICOTINIC ACID IN 

RICE BRAN 

U. SUZUKI AND S. MATSUNAGA 
(Agricultural College, Imperial University of Tokyo, Japan) 

One kilo of fat-free rice bran was extracted with hot alcohol 
(80-85 per Cent.). The alcoholic extract was greatly concentrated 
by evaporation, diluted with water, and shaken with ether for the 
removal of fat, etc. The residual aqueous liquid, after evaporation 
of the ether, was treated with sulfuric acid (total, 3 per cent.) and 
precipitated with phosphotungstic acid. After barium decomposi- 
tion of the precipitate, in the customary manner, about i gram of 
nicotinic acid (picrate) was isolated. The free acid, and the copper 
as well as the platinic-chlorid double salts, were also prepared and 
identified. Analytic data are appended. 

A SUMMARY OF THE ANALYTIC PERCENTAGE DATA 

Picrate, CeHsNOz • CeHsNaOi 



Calculated . 
Found . . . . 



40.91 

40.45 
40.68 



H 



2.27 
2.41 
2.51 



N 



15-91 
16.50 
16.19 



Picric acid 



65.06 
65-50 



Cu 



Pt 



Free acid, CsHsNO- 


Calculated 


58.54 
58.36 


4.07 
4.32 


11.38 
11.80 


.... 




Found 


.... 


Copper Salt, (C6H4N02)2Cu 


Calculated 







9-13 
9.06 





20.68 1 


Found 


20.94 







Platinum-chlorid 


double Salt, (GH=N0.-HCl)2PtCU 




Calculated 
















29.72 


Found 


30.00 



This appears to be the first time that nicotinic acid has been 
detected in vegetable matter, although Jahns,^ and Schulze and 
Frankfurt^ have found trigonellin (the methyl-betain Compound of 
nicotinic acid) in plants, and Schreiner and Shorey^ have identified, 
in humus soils, picolin carboxylic acid (a homolog of nicotinic 
acid). 

'Jahns: Ber. d. deut. ehem. Gesell, 1885, xviii, p. 2518; 1887, xx, p. 2840. 

^Schulze and Frankfurt: Ibid., 1894, xxvii, p. 769; Biochemical Bulletin, 
1912, ii, p. 18. 

'Schreiner and Shorey: Bull. 53, U. S. Dept. of Agric, p. 28 (1909). 

228 



A STUDY OF THE INFLUENCE OF CANCER 
EXTRACTS ON THE GROWTH OF LUPIN 

SEEDLINGS^ 

JACOB ROSENBLOOM 

{Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

Introduction. One of the peculiar effects of Cancer is the 
resultant cachexia. There have been many efforts to find, in cancer 
tissue, a poison that might account for the characteristic cachexia. 
It has been claimed that the cachexia is due to pressure by the grow- 
ing tumor on the blood vessels and consequent interference with 
adjacent circulation, with development öf areas of necrosis, autol- 
ysis, and production of hemolytic and toxic substances. 

Rülf^ considers that proteases are important factors in the cau- 
sation of cancer cachexia. Bard^ found that blood is rapidly hemo- 
lyzed in hemorrhagic carcinomatous exudates in serous cavities, 
which is not the case in exudates under other conditions. KuHmann* 
observed that extracts of Carcinoma contain hemolytic substances 
that are active in vivo and in vitro, soluble in alcohol and water, and 
toxic for all varieties of corpuscles. Micheli and Donati^ also 
found hemolytic substances in eight of sixteen tumors, of which five 
hemolyzed all varieties of corpuscles and three acted on some varie- 
ties only. They thought the hemolytic substances result f rom autol- 
ysis of the tumors, as it is well known that certain hemolytic sub- 
stances occur among the products of autolysis of normal tissues. 

^This paper presents the results of a preliminary study that was begun, at 
Dr. Gies' Suggestion, as a part of the plan of biochemical research described 
in Sttidies in cancer and allied subjects, conducted under the auspices of the 
George Crocker Special Research Fund, 1912, iii, p. 153 (in press). The plan 
includes a study of the effect of cancer extracts on cells of all kinds, including 
Cancer cells. 

-Rülf: Zeit. f. Krebsforsch., 1906, iv, p. 417. 

*Bard: La semaine med., 1901, xxi, p. 201. 

*Kullmann: Zeit. f. klin. Med., 1904, liii, p. 293. 

" Micheli and Donati : Riforma med., 1903, xix, p. 1037. 

229 



230 Influence of Cancer Extracts on Lupin Seedlings [Jan. 

Müller'' claimed, from the results of a study of nitrogenous metabo- 
lism in Cancer patients, that in cachexia of Cancer there is toxogenic 
destruction of protoplasm independent of nutrition; i. e., a specific 
toxic efifect of cancerous tissue. Müller's results have been con- 
firmed by other workers'^ but cumulative research has shown that 
the cases with normal protein catabolism exceed in number those 
with increased protein catabolism.^ 

According to the prevailing opinion Cancer cachexia is not spe- 
cific, but is the same as the cachexia of other conditions. It has 
been impossible to show the occurrence in Cancer tissue of any sub- 
stance that would account for the cachexia of this disease.^ 

In the study described below, we ascertained some of the effects 
of extracts of cancer tissue on the growth of lupin seedlings, in the 
hope that this procedure for the detection of toxic substances might 
yield significant results. 

Experimental. Preparation of lupin seedlings. Lupin seeds 
were soaked in water overnight. Seeds of the same size were then 
selected and planted in wet moss. After three or four days the 
seedlings were taken from the moss, the coat of each removed, and 
the sprout rinsed with distilled water. The root was carefully 
measured on a millimeter scale. The seedlings were then fastened 
on glass rods drawn out at one end to form a sharp pointed L and 
suspended in perforated cork covers over 400 c.c. Jena beakers, each 
containing 200 c.c. of water and 5 c.c. of boiled or unboiled cancer 
extract prepared as described below. " Control " seedlings were 
suspended in distilled water. The glass rods were so adjusted that 
the roots were immersed in the liquid but the cotyledons were not 
in contact with it. Four seedlings were suspended in each beaker. 
At intervals of 20 hours all the seedling roots were measured.^^ 

' Müller : Zeit. f. klin. Med., 1889, xvi, p, 496. 

'' Char. Annal, 1891, xvi, p. 138; Arch. prov. de Med., 1899, March; Arch. f. 
Verdauungskr., 1899, v, p. 540; Riv. ven. d. sei. Med., 1899, xvi, p. 31; Zeit, f 
klin. Med., 1897, xxxiii, p. 385. 

^ Zeit. f. Krebsforsch., 1904, i, p. 199: Salkowski Festschrift, Berlin, 1904, 
P- 75; Fifth Ann. Rep't Cancer Lab., New York State Dep't of Health, 1903- 
1904. 

' Blumenthal : Salkowski Festschrift, Berlin, 1904. 

^"True and Gies : Bulletin of the Torrey Botanical Club, 1903, xxx, p. 390; 
Rose: Biochemical Bulletin, 191 i, i, p. 428. 



I9I3] 



Jacob Rosenbloom 



231 



DATA SHOWING EFFECTS OF EXTRACTS OF CANCEROUS AND NORMAL TISSUES ON THE 

GKOWTH OF LUPIN SEEDLINGS 

I. Extract of Bone Sarcoma 





Rate of growth per plant, in millimeters 


Lupin seedlings 


Unboiled extract 


Boiled extract 




ist 20 hr. 


zd 20 hr. 


ist 20 hr. 


2d 20 hr. 


A 


14 
12 

20 

17 
16 

5 


19 
28 
22 
40 

27 

8 


18 
13 
15 
16 

16 

6 


30 
38 
24 
30 

^8 


B 


C. 


£> 


Average 


Control ( average ) 











II. 


Extract of Fibroma of Uterus 






A 


8 
6 
8 
6 

7 

8 


12 
12 

10 

8 

10.5 

10 


6 
8 
8 
8 

7-5 
8 


10 


B. 


10 


C. 


12 


£> 


16 


Average 


12 


Control (average) 


II 











III (0). 


Extract of a 


Carcinoma of 


the Breast 




A 


18 
12 
15 

15 
17 


9 
7 
8 
8 
8 
7 


12 
12 

9 
15 
12 
16 


8 


B 


6 


c. 


8 


D 


8 


Average 


7.S 

8 


Control (average) 







III (b). 


Extract of Normal Breast Tissue Near the Cancer^^ 


A 


20 
18 
24 
22 
21 


8 
6 
8 
6 
7 


26 

17 
24 

25 
23 


6 


B 


8 


C 


8- 


D 


7 


Average 


7.3 





III. (c). 


Extract 


f Pectoral M 


uscle Removed 


at Operation^^ 




A 


14 

26 

14 

24 

19.5 


7 
6 
II 
8 
8 


26 
18 
26 
24 
23-S 




9 

9 

12 


B 


C 


D 


10 


Averaee 


10 







Preparation of Cancer extracts. Fresh cancerous tissue, direct 
f rom the operating room, was minced, then triturated with saiid and 
water, and the thin mixture frequently shaken for about an hour. 

""Control" figures are given in section III (a). 



232 Infliicnce of Cancer Extracts on Lupin Seedlings [Jan. 

The liquid was strained through gauze, then filtered. Portions of 
this filtered extract (boiled or unboiled) were used in the manner 
indicated above. 

Data pertaining to grozvth. The summary on page 231 pre- 
sents the results of this study. 

General conclusion. The extracts failed to inhibit growth of 
the seedlings. The observed acceleration of growth was probably 
due to inorganic salts in the extracts. It is possible, of course, that 
deleterious action by Cancer toxins was neutralized or overcome by 
the stimulating power of associated nutrient substances. This par- 
ticular poinit requires special investigation. 



THE BIOCHEMISTRY OF THE FEMALE GENITALIA^ 

3. A quantitative study of certain enzymes of the ovary, Uterus, 
and bladder, of pregnant and non-pregnant sheep 

THUISCO A. ERPF-LEFKOVICS' and JACOB ROSENBLOOM 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

Introduction. In this study we used the pregnant and non- 
pregnant ovaries and uteri, and also the bladder, in order to com- 
pare our genital results with those for an organ with a supposedly 
non-dynamic function. We desire to express our thanks to Dr. 
Robert T. Frank for his interest, and for his kindness in placing at 
our disposal the genital material employed. 

Methods. i. Preparation of extracts. {A) Five grams 
of finely divided fresh material, washed free from blood and thor- 
oughly triturated with sand, were treated with 100 c.c. of water 
and allowed to stand for 24 hours, under toluene, with frequent 
shakings. At the end of this time the extract was filtered through 
muslin, made up to 100 c.c, and aliquot portions used for the en- 
zyme tests. {B) Glycerol extracts were made in the same manner. 

2. EsTiMATiON OF ENZYMES. In cach casc, a control test was 
made with boiled extract. Lipase. A mixture of 10 c.c. of the 
extract, 0.5 c.c. of neutral ethyl butyrate and i c.c. of toluene was 
placed in a bottle and allowed to digest at 40° C. After 24 hours 
n/20 sodium hydroxide Solution was used to determine the acidity, 
with Phenolphthalein as the indicator. From this amount was 
subtracted the "control" acidity (10 c.c. of extract and i c.c. of 
toluene). 

Amylase. To 10 c.c. of i per cent. freshly prepared starch 

*The first paper in this series (a general review of the subject) has been 
accepted for publication in a later issue of the Biochemical Bulletin. The 
second paper appeared in the January issue of the Journal of Biological Chetn- 
istry, 1913, xiii, p. 511. See also Biochemical Bulletin, 1911, i, p. 115. 

^ Mr. Lefkovics died shortly after the completion of this work. See Bio- 
chemical Bulletin, 1912, i, p. 573. 

233 



234 



Biochcmistry of the Female Genitalia 



[Jan. 



paste were added lo c.c. of extract and i c.c. of toluene, and the 
mixture allowed to digest at 40° C, until duplicates no longer be- 
came blue with iodin Solution. At this point the bottles contain- 
ing the digestive mixtures were placed in boiling water to stop the 
digestions simultaneously. The contents of each bottle were then 
made up to 50 c.c. and run from a burette into boiling Fehling Solu- 
tion, acetic acid and potassium ferrocyanid being used to determine 
the end point. When the amount of sugar in the digestive mix- 
ture was less than that required completely to reduce the copper, a 
Standard glucose Solution was employed for that purpose. 

Acid- and alkali-proteases. Ten grams of gelatin were dis- 
solved in 100 c.c. of warm i per cent. Solution of sodium fluorid col- 
ored with methyl violet. This Solution was drawn into glass tubes 
I mm. in diameter and the filled tubes quickly placed in cold water 
to congeal the gelatin. The tubes were then cut into lengths of 
2-3 cm. Ten c.c. of extract were placed in a small bottle closed 
with a perforated cork through which the gelatin tubes could be 
inserted; i c.c. of toluene was added and the digestions kept at room 
temperature for 48 hours. In the estimation of acid-protease (pep- 
sin) the mixture was made acid with 0.2 per cent. hydrochloric acid 
Solution and for alkali-protease (trypsin) they were rendered alka- 
line with 0.5 per cent. sodium carbonate Solution. 

Table showing enzyme values of pregnant and non-pregnant ovary, uterus 

and bladder of sheep 

A. Non-pregnant condition 





Aqueous extract 


Glycerol extract 


Organ 


Lipase, 
c.c. 


Amy- 
lase, 
mg. 


Acid- 
pro- 
tease, 
mm. 


Alkali- 
pro- 
tease, 
mm. 


Lipase, 
c.c. 


Amy- 
lase, 
mg. 


Acid- 
pro- 
tease, 
mm. 


Alkali- 
pro- 
tease, 
mm. 


Ovary 


0.65 
305 
1-3 


5 
10 
10 


2 

I 

1-5 


2 

I 



0.65 
1-95 
1-35 


6 

5 

IS 


6 
3 
3 


0.25 

4 



Uterina mucosa 


Bladder mucosa 





B. Pregnant condition 



Ovary 

Uterina mucosa . 
Bladder mucosa. 



1-3 


10 


2 


7 


2.0 


7 


7 


8.35 


25-5 


I 


7 


8.3 


12 


3 


i.i 


7.5 


1-5 





1-35 


6 


4 



0.5 

5 
o 



The accompanying table presents the results obtained in this 
Study. The lipase values are given in terms of the amount of ji/20 



1913] Thuisco A. Erpf-Lefkomcs and Jacob Roscnhloom 235 

sodium hydroxid Solution necessary to neutralize the acidity devel- 
oped by i gram of tissue. The amylase values are given in terms of 
the amount of maitose in mg. formed per gram of tissue. The 
acid- and alkali-protease values are given in terms of the number of 
millimeters of gelatin digested in a tube by i gram of tissue. 

The results show that lipase and amylase were most abundant in 
both the ovaries and uterine mucosae of pregnant animals. Preg- 
nancy had no quantitative effect on the acid-protease (pepsin), but 
alkali-protease was increased in both the ovary and uterine mucosa. 
The bladder extracts contained lipase, amylase, and acid-protease 
(pepsin), but no alkali-protease (trypsin). 



THE BIOCHEMISTRY OF THE FEMALE GENITALIA^ 

4. On the absence of certain enzymes from the 
human chorion^ 

JACOB ROSENBLOOM 
(Laboratory of Biochemistry, University of Pittshurgh, Pittsburgh, Pa.) 

During pregnancy the chorion frondosum unites with the de- 
cidua serotina to form the placenta. The enzymes of the placenta^ 
have often been studied, but I am unable to find any record of a 
study of the enzymes of a human chorion. Through the kindness 
of Dr. Robert T. Frank, of New York, the vvriter received a fresh 
human chorion for such an investigation. 

The available chorion, which weighed 10 grams after it had been 
washed free from all blood by means of a small amount of water, 
was finely minced and two portions, 5 grams each, were taken for 
the preparation of extracts, which were made as follows : ( i ) The 
material was triturated with sand, 200 c.c. of water added, and 
the mixture allowed to stand with frequent shakings for 24 hours 
under toluene. At the end of that time, the extract was filtered 
through muslin, and the filtrate used in the tests for various en- 
zymes. (2) A glycerol extract was made in the same way. In 
testing for enzymes, control portions were always taken, which 
were boiled before their addition to the Solutions or suspensions of 
Substrate.'' 

The accompanying table presents the data obtained in this study. 
The data show that both glycerol and aqueous extracts of a human 

* See the first footnote of the preceding paper in this issue of the Biochem- 
ICAL Bulletin. 

" The analytic work was done in the Biochemical Laboratory of Columbia 
University, at the College of Physicians and Surgeons, New York. 

^ Frank: Surgery, Gynecology and Obstetrics, 1912, xv, p. 561. 

' See the description of methods in the paper preceding this one. 

236 



I9I3] 



Jacob Rosenhloom 



237 



chorion were free from amylase, sucrase, maltase, lactase, lipase. 
peptidase, ereptase, acid-protease and alkali-protease. 

Table showing results of ensyme tests in glycerol and aqueous extracts of a 

human chorion 



Enzyme 


Substrate 


Glycerol extract 


Aqueous extract 


Amylase 

Sucrase 


Starch 


Absent 


Absent 


Sucrose 


Absent 


Absent 


Lactase 


Lactose 


Absent 


Absent 


A'Ialtasp 


Maltose 


Absent 


Absent 


Lipase 


Ethyl butyrate 

Glycyltryptophan 

Witte Peptone 


Absent 


Absent 


Ppntida'sp 


Absent 


Absent 




Absent 


Absent 


Acid-protease 

Alkali-protease 


Gelatin and fibrin^ 

Gelatin and fibrin^ 


Absent 


Absent 


Absent 


Absent 







It would seem, from the results of this study, either that the 
enzymes of the placenta are formed at a comparatively late period 
or that the decidua serotina furnishes the enzymes which subse- 
quently appear in the placenta. Possibly the presence of blood in 
the placenta accounts for the occurrence of certain enzymes that 
have been detected in the placenta. 

° Extract rendered acid with 0.2 per cent. hydrochloric acid Solution. 
* Extract rendered alkaline with 0.5 per cent. sodium carbonate Solution. 



A DEPARTMENT OF BIOCHEMICAL RESEARCH AT 
VINELAND, NEW JERSEY^ 

AMOS W. PETERS 

To the Training School at Vineland, N. J., belongs the credit for 
the first establishment anywhere in the world of a biochemical 
laboratory as one means of investigation of the problem of feeble- 
mindedness in children. To the writer of this article has fallen the 
honor as well as the heavy duty of testing what are the possibilities 
of biochemical research in the field of feeble-mindedness. The 
large problem which this unfortunate affliction of a considerable 
portion of humanity presents to organized society is becoming daily 
more evident, as its economic bürden and its social consequences 
force themselves on public attention. Research on the problem is a 
crying need not simply f rom the humanitarian Standpoint, but also as 
an economic necessity. The care and treatment of these cases and 
the governmental management of this problem, including its amelior- 
ment and prevention, will in the future rest on the basis of data ob- 
tained by scientific research. At present we are proceeding on a 
very small amount of such data and we are just discovering, after 
some preliminary efforts made in the psychological direction, how 
extensive and manysided this problem is. What assurance have we 
that our present method of dealing with the problem is in rational 
accord with the nature and origin of the condition? Our proce- 
dures are in the stage of costly empiricism and in the very infancy 
of scientific investigation. It is therefore an important step for- 
ward when this Institution ventures to add to its present psycho- 
logical method of investigation that of the rapidly growing and 
fundamental science of biochemistry. The need for this additional 
method of attack, and the tendency of expert thought toward it, is 
well illustrated by the following quotation from the words of a 
leader in the study of problems of psychopathology, Dr. Southard : 

^Reprinted from The Training School, 1912, ix, p. 70 (Sep.). 

238 



1913] Arnos W. Peters 239 

The majority of cases of mental diseases are, I am convinced by 
special studies, characterized by the occurrence of obvious brain lesions, 
i. e., even in the present stage of science they possess a structural pa- 
thology. Do they therefore possess no functional pathology? Their 
possession of the two aspects is a truism. Should we not study both 
aspects ? 

Furthermore, suppose we learn that, whereas three quarters of our 
cases of mental disease exhibit obvious irrecoverable brain lesions, 
another quarter falls to shovv these. Suppose the methods of micro- 
scopic research should still fall to show in many cases essential or 
irreversible brain lesions, should we not stultify ourselves if we did 
not abandon jor the research campaign both that psychopathology 
which has taught us the main course of our disease and the neuro- 
pathology which has proved usefully negative? Should we not repair 
at once to the chemistry of metabolism, the physiology of internal 
secretions, and the entire point of view of psychopathology? Dis- 
coveries in the latter fields, concrete and pertinent facts, would carry 
US back to the tissues and back to the processes of the nervous System, 
to neuropathology, structural and functional and to psychopathology, 
and enlighten many dark corners therein. He who adheres to the clas- 
sical Problems as they He within the teaching divisions of any science is 
not apt to change the face of that science.^ 

It is the method of science to develop the ultimate truth with 
its numerous and involved qualifications, which are due to the infi- 
nite complexity of natura itself, by means of hypotheses. These 
are repeatedly set up and repeatedly confirmed or refuted and re- 
placed by others of better construction in view of previous expe- 
rience. Whether the hypothesis was exactly correct or not — ulti- 
mately tenable or untenable — ^becomes a matter of no practical sig- 
nificance. The testing of hypotheses develops facts, and facts, dem- 
onstrated and adequately qualified truths, are the precious heritage 
of the race f rom previous human endeavor. Now, then, the hypoth- 
esis which underlies the use of the biochemical method in this 
problem is that which postulates, simply, a relation between patho- 

^ Southard, E. E. : " Psychopathology and Neuropathology : The Problems 
of Teaching and Research Contrasted," Amer. Jour. of Psychol., 23 : 230-235, 
1912. Read by invitation in a Symposium at a meeting of the American Psycho- 
logical Association, December 28, 191 1, at the Hospital for the Insane, Wash- 
ington, D. C. 



240 Biochcmical Research at Vineland, N. J . [Jan. 

logical mental action on the one band, and the physical condition 
of the brain and body on the other. We will not discuss this propo- 
sition — no, this hypothesis — with our readers. It is not worth while. 
We only wish gently to call their attention to it and to prevent 
them from shying at this subject on theoretical grounds. This, 
then, is our generalized hypothesis, and it is clear that finally our 
logical efforts will be directed toward the correlation of data, psy- 
chological and biological, taken in their widest sense. This part of 
our effort will be small, however, compared with the requirement for 
painstaking and persistent experimental determination of facts 
which are the real values we are seeking. In this connection it 
should be noticed that the present literature of chemical biology 
contains numerous concrete examples of investigations which have 
an evident relation to the problems of psychopathology viewed from 
the broad Standpoint of Southard, as above quoted. In future 
numbers of the Biochemical Bulletin we shall, from time to 
time, present our readers with notes and criticisms on this literature. 

It is important that the general aim of this biochemical effort 
should not be misunderstood, nor its results misinterpreted. The 
primary and only initial object is to contribute toward the elncida- 
tion of the conditions of psychopathological action by means of the 
biochemical method. The curing of tuberculosis was an entirely 
premature and abortive expenditure of effort before the elucidation 
of the cause and conditions of that disease. When once these con- 
ditions have been adequately determined, valuable applications of 
the new knowledge always follow, and sometimes with astonish- 
ing results. But now we are only in the beginning of the period of 
strenuous seeking after much needed information. We wish also 
to emphasize that we regard the biochemical as only one, but after 
the psychological the next in importance, of the methods that are 
available for determining conditions of abnormal mental action. 
We picture our final understanding of these conditions to be a com- 
posite and correlated result obtained by different methods, none of 
which alone would have ever yielded adequate knowledge. 

Now we are asked just what, concretely, is the field of application 
of biochemistry to the problems of feeble-mindedness. 

This question could be best answered by illustrations from the 



1913] Arnos W. Peters 241 

literature of investigation along biochemical lines; but, as above 
stated, this we shall continue to present in future numbers of this 
Bulletin. At present, before we have actually begun oiir own 
experimental work, we can give only an outline of the topics we 
plan to pursue to such extent as workers and material resources per- 
mit. The field is so rieh as to tax the judgment in the selection of 
the first attacks, and we are well aware that we are outlining more 
than our present resources permit to be done in the near future. 
Publicity and hearty Cooperation with other individuals and in- 
stitutions is, of course, our policy. In the present article, however, 
and at the very beginning of our work, we are describing only the 
nature of the work to be done without specific detail regarding par- 
ticular problems or methods. 

Our primary line of effort to which the others are logically re- 
lated is the study of the conditions of metabolism presented by the 
feeble minded of this institution. Very few studies of this nature 
have been made, and the material for them is here presented under 
favorable conditions for investigation. Promiscuous examinations 
or experiments will not be made. But at first typical and psycho- 
logically well-known and defined cases will be selected. For orien- 
tation they will at first be studied in their undisturbed condition 
before the experimental factor is introduced. By metabolism we 
understand, of course, the sum total of the chemical changes which 
a living organism continually performs within its tissues and upon 
the substances which it utilizes. The progress of biological science 
has made the term practically synonymous with the processes of life 
in so far as they are non-psychical. Under this head we intend to 
subject the idea of intoxication, whether endogenous (autointoxi- 
cation) or exogenous, to a rather thorough testing, especially in its 
relation to psychopathological phenomena. Two other related 
topics with which we will be compelled to deal in this connection 
pertain to the subject of glandulär secretions and that of lipoid or 
phosphorus metabolism. It is well known that the method of glan- 
dulär feeding is extensively practiced in psychopathological cases 
and institutions. It appears that this is usually done in a promiscu- 
ous way with but little of the Clements of control experiments 
or of adequate therapeutic indications. In our future notes and 



242 Biochemical Research at Vineland, N. J. [Jan. 

criticisms on the literature, we shall treat this subject more fully. 
It seems a pity, f rom both the scientific and the humanitarian stand- 
point, that such potentially valuable experiments on human subjects 
should pass without an examination of their most important factor 
— that of the metaboHsm of the physiologically much afifected 
subject. 

Our second line of effort will be that of lipoid and brain chem- 
istry. It will not be pursued extensively until we have obtained, 
from the observations of metabolism and the third line of effort 
described below, some indications of the directions in this large and 
inherently difficult field that it would be best to pursue. Contrary 
to the common Impression, the present literature already shows the 
important and practical bearing of this little developed field of chem- 
istry on the psychopathological problem. 

A third kind of work which in the near future will become 
practically inevitable is the study of heredity, growth and develop- 
ment from the particitlar angle of view of the psychopathologist. 
It is well known how strongly the scientific and the public attention 
is now fixed upon the hereditary and congenital (if not hereditary) 
factors involved in the conditions of abnormal mental action. With- 
out going into detail, we wish to emphasize the fact that the hered- 
itary factor in this problem by no means removes it from the field 
of biochemical study, nor makes the pathological conditions any less 
amenable to elucidation by that method. In fact, the only real hope 
for the elucidation of the processes of reproduction and heredity 
seems, in the light of experiments already made, to lie in the direc- 
tion of an intimate knowledge of the chemistry and physics of the 
protoplasmic basis of Hfe. 

Biochemical Laboratory, Training School, 
Vineland, New Jersey. 



BIOCHEMISTRY IN NEW YORK TWENTY 

YEARS AGO^ 

E. E. SMITH 

The Status of biochemistry in New York City, in 1891, is well 
depicted by an incident that occurred in that year at the library of 
the New York Academy of Medicine. The writer had just come to 
the city and was seeking, for reference, a copy of Maly's Jahres- 
bericht für Thier-Chemie. In reply to an inquiry, he was informed 
that such a work was not in the Academy library and would most 
likely be found at the Veterinary College. Chemistry was, indeed, 
well established at that time in the curriculum of the medical schools 
of the city, but it consisted largely of descriptive organic and in- 
organic chemistry, and found relatively scant application in physiol- 
ogy and pathology.^ 

To the younger graduates of Yale, the pioneer work of Chitten- 
den was known, but it had not been implanted here. It did lead, 
however, to the Inspiration of Dr. C. A. Herter, then beginning to 
specialize in neurology ; and when he came to realize, as he soon did, 
how closely related was this field to the pathology of nutrition and 
determined to establish a laboratory for the investigation of this sub- 
ject, he naturally turned to Prof. Chittenden for someone with tech- 
nical training to undertake this work. 

^For previous special contributions to the history of biological chemistry in 
New York see the Biochemical Bulletin, 191 i, i, p. 245, and 1912, i, p. 377. 

*"To appreciate the significance of all this, it should be remembered that, 
with the exception of the work in the pathological laboratories of the Colleges, 
the work of the Board of Health, and the work done by Dr. S. J. Meltzer, there 
was practically no scientific investigation in medicine worthy of the name in 
New York City at that time (when the 'Laboratory of C. A. Herter' was 
created). What was true of New York was essentially true of the country at 
large. . . . Dr. Herter found the study of the nervous System so abounding in 
confusion that he soon turned his attention to chemical problems, especially those 
connected with pathological conditions. Among those intimately associated with 
him in this work have been E. E. Smith, A. J. Wakeman and, of late, H. D. 
Dakin." Lusk: Science, 191 1, xxxiii, p. 846. [Ed.] 

243 



244 Biochemistry in New York Twcnty Ycars Ago [Jan. 

How little Dr. Herter appreciated the equipment that would be 
required is indicated by bis Suggestion that the work be conducted in 
the art studio of his brother, then absent in Europe. It did not re- 
quire many months, however, to reveal to him something of the 
technical scope of the field to which he was to devote the two decades 
permitted him for the completion of his hfe work, At the outset, 
he desired adequate equipment ; and when he returned f rom his sum- 
mer rest, in 1891, he was enthusiastically appreciative of the well- 
equipped laboratory awaiting him in the basement of his residence. 
This, three years later, was transferred to his newly built home 
where the entire upper floor, 50 X 100 feet, was devoted to this 
special work. It was no unusual, though an unique, experience to 
house in his animal room, rabbits, dogs, monkeys, füll grown hogs, 
and other animals in an array that would have astounded the unin- 
formed passerby in this district of elegant homes. 

It was not, however, the equipment that invites attention to 
Herter's early work nor was it the display he made of his devotion 
to this new field. Both were modest. What has lingered and al- 
ways will remain in my memory of twenty years ago is the serious- 
ness with which the work was undertaken. In later years, when 
his life was so filled with the success and magnitude of his work, it 
was to be expected that he would throw all that was in him into it ; 
but that he should have devoted himself so largely to it when its 
value was uncertain, or at least not demonstrated, indicates the 
profound purpose that was leading him to undertake it. Not in- 
frequently, when night had come and the day's work was done, we 
forgot ourselves in both discussing what we had attempted and 
planning what we hoped to do; finally awakening to a realization 
that we were neglecting the proper demands of our respective family 
circles. 

Only a fraction of the work done at that period was ever pub- 
lished. The first paper, " Uric acid elimination in health and dis- 
ease," was a record of investigations inspired by the extravagant 
Claims of the English physician, Haig. We differed with him in 
many important conclusions. We did not find that uric acid forma- 
tion was always constant and that elimination was determined by the 
degree of alkalinity of the blood, but found it to vary with the diet 



1913] -E- E. Smith 245 

in health and with conditions unknown to us in disease. Moreover, 
we did not recognize it as a causative factor in the many diseases to 
which this role was assigned by Haig, but rather regarded its in- 
creased elimination as a result of the morbid condition. Horbac- 
zewsky's work did not come to our attention tili after the publication 
of this first paper. 

The study of epileptics led to the conclusion that, in some cases 
of so-called idiopathic epilepsy, the onset of the seizures was deter- 
mined not by a uric acid accumulation, as claimed by Haig, but by 
a toxemia of gastro-intestinal origin. Indican, which had received 
scant attention from clinicians up to this time, was found to be a 
valuable index to the condition; as was also the elimination of phenol 
and ethereal sulphates. The occurrence of these products in undue 
quantity seemed to bear a direct relation to the onset of the seizures. 

As was natural, there followed an elaborate study of the gastro- 
intestinal conditions in other diseases, especially those with marked 
neurotic manifestations ; and the conclusion was reached that the neu- 
rotic exacerbations in many conditions were due to a gastro-intes- 
tinal toxemia. An entirely different line of study was the presence 
of lead and its distribution in cases of chronic lead poisoning. The 
results of these analyses were never published. 

Investigations to which was devoted a very great amount of 
work and which covered a very wide scope, as well, were the studies 
of the causes of uremic intoxication. The many theories which had 
been elaborated to explain this condition were each in turn subjected 
to investigation, involving extensive animal experimentation as well 
as intricate chemical research. The work covered several years and 
the results were of very great interest to us, and certainly influenced 
Herter's later work, but they were never published. 

During this period, there was a striking lack of activity in re- 
search in chemical pathology in New York; indeed, this was only 
the time of the awakening of general interest in the most active Cen- 
ters of medical science. Aside from Herter's work, only a single 
paper presented at the Academy of Medicine in that period comes to 
my mind ; and that was so glaringly f aulty that one hesitates to con- 
sider its sincerity. 

Von Noorden's Pathologie des Stoffwechsels, which appeared 



246 Biochemistry in New York Twenty Years Ago [Jan. 

at this time, was as a beacon light on a dark night. I received my 
copy before Dr. Herter's attention was called to the work. He saw 
it on my table and, borrowing it, informed me shortly afterwards 
that if I really wanted a copy I had better send for another. True 
to bis word, I never saw this first copy again and I doubt not that it 
rests now in his library well worn with eager study, which it received 
at that time. My second copy served a similar purpose in my own 
hands. 

My personal relation with Dr. Herter was interrupted by the 
decision to study medicine. The modest beginnings of his work, 
which hardly interested more than a narrow circle of personal 
friends and admirers, grew to a proportion that brought him into 
national and, indeed, international prominence. A man of unusual 
personal charm and sincere purpose, he demonstrated how personal 
opportunity could find unselfish application to the benefit of his 
fellowmen in the field of applied medical science. 

Laboratory, 50 East Forty-first Street, 
New York City. 



IMMUNITY IN SOME OF ITS BIOCHEMICAL 

ASPECTS^ 

CHARLES FREDERICK BOLDUAN 
(Department of Health, New York City) 

(WITH PLATE 2) 

Contents. — Infection, 247, Immunity: natural, 248; acquired, 248; speci- 
ficity, 249; additional defenses, 249; Behring's discovery of antitoxin, 250; bac- 
teriolysins, hemolysins (cytolysins), 250; complement and immune body, 251; 
agglutinins, 251; Opsonins, 252; precipitins, 252; anti-antibodies, 252. Immunity 
from the Standpoint of cell niitrition, 253; Ehrlich's " side chain " theory, 253; 
receptors, 254; Weigert's " overproduction " theory, 254; natural immunity, 256; 
anaphylaxis, 256; results of enteral and parenteral introduction of protein, 257; 
significance of period of incubation, 258, and bearing on intoxication by infection 
(endotoxins), 258. Modern chemotherapy according to Ehrlich, 259. Chemical 
nature of antibodies, 260. 

Infection. One of the most interesting problems to all of us 
is that presented by disease, especially by what we call " infectious " 
disease. Under this term we mean disease produced by living organ- 
isms or their products. Among the organisms producing disease 
in man are bacteria, molds, yeasts, and protozoa, and we may con- 
veniently speak of these collectively as germs. 

The manner in which the various germs produce disease in man, 

their mode of entrance into the body, the part of the body attacked 

— all these differ considerably with the different germs. Some 

like the bacillus of diphtheria and the bacillus of tetanus (lockjaw) 

secrete very powerful poisons, and while the germs themselves do 

not penetrate deeply into the body tissues, their poison is absorbed 

and gives rise to severe Symptoms. In the case of other germs, for 

example the tubercle bacillus, the organisms penetrate deeply into 

the body tissues and there multiply. In their growth they destroy 

the cells in which they lodge and, by their poisons, affect the 

entire body. 

^ Lecture delivered, by invitation, under the auspices of the Columbia Univer- 
sity Biochemical Association, at the College of Physicians and Surgeons, Novem- 
ber 16, 1912. 

247 



248 Immnnity in Some of its Biochemical Aspects [Jan. 

Most germs, for some obscure reason, affect by preference cer- 
taiii parts of the body. The typhoid bacillus usually lodges in the 
wall of the small intestine; the meningococcus prefers the lining 
membranes of the brain and spinal cord; the gonococcus is very 
prone to attack the mucons membrane of the genital organs and of 
the eye; the pneumococcus affects chiefly the respiratory organs; 
the diphtheria bacillus lodges in the throat and nasal passages ; the 
malaria parasite lodges only in the red blood cells; and certain 
molds affect only the skin. 

Immunity. Natural immunity. It is very well known, 
however, that certain infectious diseases occur naturally only among 
some of the lower animals and do not affect man, while conversely, 
others appear to attack only man. Among the latter may be men- 
tioned typhoid fever, syphilis, gonorrhea. In speaking of the re- 
sistance evidently possessed by certain species we make use of the 
term natural immunity. Thus chickens and frogs possess a natural 
immunity against tetanus (lockjaw) ; dogs, a natural immunity 
against anthrax; goats, a natural immunity against tuberculosis ; 
and man, a natural immunity against certain diseases of cattle. 
This natural immunity, however, is not always absolute. Chickens, 
for example, can be infected with tetanus if their bodies are chilled, 
and frogs can be made susceptible to tetanus by keeping them un- 
duly warm. 

Acquired immunity. Another form of immunity is that ob- 
served in individuals who have had one attack of a particular in- 
fection; thereafter they are practically safe from a second attack. 
These individuals are said to possess an acquired immunity. This 
form of immunity is well illustrated in scarlet fever, measles, small- 
pox, yellow fever. Often this immunity lasts throughout the life- 
time of the individual though there are exceptions. 

In studying this form of immunity, Pasteur conceived the idea 
of artificially producing an attack of a given infection in order to 
Protect the individual against another attack. He realized that it 
was necessary, however, to so control matters that the original 
attack should run a very mild course and not endanger the life of 
the individual. After considerable experimental work, Pasteur 
found that this could be accomplished by artificially weakening the 



1913] Charles Frederick Boldnan 249 

bacteria with which the original attack was produced. Subse- 
quently Salmon and Smith, in this country, showed that it was not 
necessary to produce even a mild attack of the disease by injecting 
living germs, but that the injection of dead germs would produce an 
immunity against that particiliar infection. 

Specificity of acquired immunity. Acquired immunity, whether 
caused by a previous natural attack of the disease, or artificially by 
the inoculation of living or dead germs, is always strictly specific; 
that is, the protection extends only to the particular disease which 
has previously occurred or against germs of the kind previously 
injected. An attack of scarlet fever protects only against scarlet 
fever but not against measles. Inoculating an individual with ty- 
phoid bacilli protects him only against typhoid fever, but not against 
dysentery, plague or cholera. This acquired immunity is often 
transmitted from mother to offspring, transmission being effected 
mainly, according to Famulener, through the Colostrum. 

Additional natural defenses against DISEASE. Beforc ex- 
amining into the nature of specific acquired immunity, let me call 
attention to certain important means by which the body is protected 
against infectious diseases in general. Many of these means are 
so commonplace that their significance is often overlooked. 

The protection afforded by the unbroken skin is undoubtedly one 
of the most important means of defense. A similar protection, 
though less effective, is afforded by intact and healthy mucous mem- 
branes. The acid gastric juice undoubtedly destroys large numbers 
of swallowed germs. It has been found that fresh blood serum is 
able to kill a considerable number of germs, and this is therefore 
another mode of defense. The white blood cells (leucocytes) 
appear to be designed especially to destroy invading micröorganisms. 
These cells take hold of, or rather engulf, the germs and digest 
them. Still another mode of defense is seen in what takes place in 
abscesses. When these are examined, it is found that the body has 
built a wall of cells around the infected area, thus shutting off the 
germs and their poisonous products from the rest of the body. 
Finally, mention may be made of the collection of fluid, i. e., of 
serum, as perhaps a means designed to dilute irritant poisons (pleu- 
risy, Peritonitis). 



250 hnmunity in Some of its Biochemicol Aspects [Jan. 

The means of protection we have just recited are all general in 
their action, that is, not directed specifically against only one partic- 
ular infection, Let us now return to a consideration of the specific 
acquired immunity already mentioned. 

Behring's DISCOVERY OF ANTITOXIN, Most of our knowlcdgc 
concerning specific acquired immunity dates from Behring's discov- 
ery of the antitoxins of diphtheria and tetanus, in 1890. 

Behring found that when an animal is injected with gradually 
increasing doses of toxin, e. g., with diphtheria toxin, it is able, after 
a time, to withstand doses of the poison sufficient to kill hundreds 
of animals not so treated. He found that the blood serum of the 
treated animals contained something which neutralized the diph- 
theria poison, and rendered it harmless. This something he called 
an antitoxin. Investigation showed that the antitoxin was strictly 
specific, the antitoxin for diphtheria neutralized only the toxin of 
diphtheria, the antitoxin for tetanus, only that of tetanus. 

Bacteriolysins and hemolysins (cytolysins). Another 
important advance was made in 1894 when Pfeiffer showed that, 
just as an animal injected with gradually increasing doses of toxin 
produces an antitoxin in its blood, so also, when injected with bac- 
teria (cholera bacilli), it produces substances which kill and dissolve 
the injected micröorganisms. We have already said that fresh 
blood serum is able to kill a considerable number of bacteria, and 
that this probably constitutes one of the defenses of the body against 
bacterial Invasion. When the animal is injected with gradually in- 
creasing amounts of bacteria, however, this destructive power in- 
creases very greatly, but only for the particular kind of bacterium 
used for injection. In other words, the action is strictly specific. 
If an animal is injected with cholera bacilli, the serum will, after a 
time, even in very small doses kill enormous numbers of cholera 
bacilli ; tested against typhoid bacilli, or on other bacteria, its de- 
structive effect is merely that of normal serum from an untreated 
animal. When the action of the serum is studied under the micro- 
scope, it is seen that the bacteria are actually broken up and dis- 
solved. Hence such a serum is spoken of as a " bactcriolysin." 
Since the bacteria are also killed by this action, we also use the term 
" bactericidal " in speaking of such a serum. 



1913] Charles Frederick Bolduan 251 

It has been found that this action may be developed against cells 
other than bacteria. When red blood cells are used for the injec- 
tions, the serum acquires dissolving properties for these ; and here 
again the action is strictly specific, so that when blood cells from a 
chicken are injected into an animal, the serum of the injected animal 
acquires increased solvent powers only for chicken blood cells, not 
for blood cells of other animals. Sera directed against blood cells 
are usually spoken of as hemolysins. The term cytolysin is used to 
embrace all these cell-dissolving sera. 

Complement and immune body. Investigation has shown that 
the mode of action of these dissolving sera is somewhat complex, 
and consists of the Joint action of two substances. It may be re- 
called that this dissolving action was observed in fresli serum. 
Serum which had stood for several days no longer possessed this 
property. The researches of Metchnikoff and Bordet showed that 
the füll solvent power could be restored by the addition of a little 
fresh serum, even from a normal, untreated animal. Evidently, 
then, of the two substances concerned in this dissolving action, one 
is quite stable, and the other highly labile. The labile substance, 
derived from a normal untreated animal, is spoken of as the com- 
plement; it is not specific. The stable substance, present only in 
the serum of the treated animal, is called the immune body; it is 
highly specific. When an animal is repeatedly injected with grad- 
ually increasing doses of bacteria, or other cells, it responds by man- 
ufacturing large quantities of this "immune body" directed spe- 
cifically against the injected cells. The complement is not increased 
in the process. 

Agglutinins. When the serum of an animal which has been 
repeatedly injected with gradually increasing doses of bacteria is 
brought into contact with some of the bacteria, careful Observation 
under the microscope reveals a very interesting series of changes. 
Thus, if typhoid bacilli are mixed with a specific antityphoid serum 
(obtained, let us say, from a rabbit previously injected with typhoid 
bacilli), one notices, first, that the motility of the bacilli becomes 
markedly diminished. This is followed by the gradual collection 
of the bacilli into clumps. At the end of an hour or two, in place 
of countless bacteria moving quickly through the field, one sees 



252 Iinmiinity in Some of its Biochemical Aspccts [Jan. 

merely several groups of absolutely immobile bacilli. If the reac- 
tion is feeble, the clumps are small, and one finds comparatively 
many isolated and, perhaps, also moving bacteria. This phenome- 
non is spoken of as aggliitination, and the substance in the serum 
which brings this about is called agglutinin. The clumping thus 
broiight about does not kill the bacteria ; moreover, it makes no dif- 
ference whether the serum is freshly drawn or has been kept for 
some time — it will agglutinate equally well ; and it does not require 
the addition of fresh serum as do the bacteriolysins. Like the an- 
titoxins and the bacteriolysins, the agglutinins are strictly specific, so 
that serum from an animal previously injected with typhoid bacilli 
will agglutinate only typhoid bacilli; one from an animal injected 
with dysentery bacilli, only such bacilli, etc. 

Opsonins. We have already said that the white blood corpus- 
cles (leucocytes) take up bacteria and destroy them. Wright, of 
England, showed that certain substances in blood serum have the 
power of increasing the appetite, as it were, of the leucocytes, and 
furthermore, that the amount of these substances can be increased 
by properly graduated injections of the appropriate bacteria. These 
substances he called Opsonins. They are specific, just as are the 
antitoxins, the bacteriolysins, and the agglutinins; that is to say, 
when typhoid bacilli are injected into the body, only the Opsonin 
for typhoid bacilli is affected; when staphylococci are employed, 
only the Opsonin for such organisms is affected, etc. 

Precipitins. If, instead of injecting bacteria or other cells, we 
inject an animal with Solutions of albuminous material ; for example, 
if we inject a rabbit with chicken-egg albumin, we find that the 
rabbit serum acquires the power to produce a precipitate when mixed 
with chicken-egg albumin. This action, too, is highly specific, so 
that if the serum is tested against the albumin from any other animal, 
e. g., from a duck tgg, no precipitate will be produced. If a rabbit 
is treated with human blood, the rabbit serum will produce a pre- 
cipitate when mixed with human blood, but not when mixed with 
any other blood. The substance in the treated animal's serum is 
spoken of as a precipitin. This test, as you probably know, is used 
in criminal cases to determine whether or not certain stains are 
those of human blood or otherwise. 



1913] Charles Frederick Bolduan 253 

Anti-antibodies. But even this list does not exhaust the list 
of "antibodies" which it is possible to produce. When enzymes 
are injected into an animal, the latter responds by producing anti- 
enzymes, and when certain " antibodies " are injected, anti-anti- 
bodies are produced. 

Immunity regarded from the Standpoint o£ cell nutrition. 
The whole subject of infection and immunity, and particularly the 
production of the antibodies just discussed, is best appreciated when 
regarded from the Standpoint of nutrition; for what, after all, is 
this apparent conflict between bacteria and the animal body but 
the mutual attempt of each to use the other for food. Let it be 
noted that production of the various antibodies takes place only 
when the bacteria or other allen cells are introduced parenterally, 
i. e., by ways other than the gastrointestinal tract. We may ex- 
plain this by saying that when introduced by the gastrointestinal tract 
the molecules of the food stufifs (organic) are split up and rebuilt 
in such a way that the material requires no further extensive altera- 
tion in order to serve as food for the various cells of the body. In 
the animal body this breaking down and building up is delegated to 
certain specialized cells ; in the primitive organisms, however, we 
must believe that each cell was required to break down and build up 
its own food. When parenterally situated cells of the higher animal 
are thus presented with the unprepared food which the parenteral 
introduction brings them, it may be assumed that they behave as 
does the primitive cell, and proceed to lay hold of and attempt to 
assimilate the injected material. With this introduction, we may 
pass at once to a consideration of Ehrlich's "side chain theory," 
which still offers the best explanation for the formation of the 
various antibodies. It is essentially a theory of cell nutrition. 

Ehrliches "side chain" theory. According to Ehrlich's 
conception, every cell is armed with a large number of chemical 
groups whose function is to lay hold of nutriment and anchor this 
in the cell. These groups he calls receptors or side chains. Only 
such substances can serve as nutriment which can thus be bound 
chemically to the cell protoplasm. He believes that the receptors are 
of at least three different kinds, and speaks of receptors of the " first 
Order," "second order" and " third order." These are best de- 
scribed with the aid of a diagram such as the accompanying one. 



254 Immunity in Some of its Biochemical Aspccts [Jan. 

Rcceptors. In view of what has been said it is obvious that 
the simplest mechanism by which the cell can lay hold on food par- 
ticles is a receptor which merely anchors food, leaving the digestion 
entirely to the cell proper. It may be assumed that this type of re- 
ceptor suffices for comparatively small food molecules. When a 
larger and more complex food molecule presents itself, it may be as- 
sumed that a receptor would be reqiiired which not merely anchors 
but also acts on the food molecule to make it more readily assimi- 
lable. These two types are shown in A and B respectively (Plate 2). 
It will be noted that the receptor in B possesses an anchoring group 
{h) and an active group (Z) which acts on the food molecule. It 
is conceivable that an economy in structure could be effected in B, 
if, in place of the active group (Z), there were merely provision for 
the anchoring of an enzyme. The active group (Z) could then be 
dispensed with and the enzyme called upon only when a food mole- 
cule had been anchored by the receptor. Such an arrangement is 
shown in C (Plate 2). 

Weigert's " over prodtiction " theory. At this point you may 
very properly inquire why we assume the existence of receptors of 
these types. To explain this, let us go back to the productiön of an- 
titoxin in response to injections of toxin. It will be recalled that 
the toxin can be neutralized by the antitoxin. Moreover, and this 
is the important point, this action is strictly specific, so that, for 
example, diphtheria antitoxin neutralizes only diphtheria toxin; 
against any other toxin it is absolutely without effect. Since it 
can be satisfactorily shown that the antitoxin is not altered toxin, 
it is necessary to explain the productiön of antitoxin by the body 
cells. We have said above that only such substances can serve as 
nutriment for the cell which can be tied chemically to the cell pro- 
toplasm. Expressing this in terms of receptors, we would say that 
only such substances as possess groups fitting the receptors of the 
cell can be anchored to the cell. In thinking of these groups and the 
way in which they fit together, we must have stereochemical rela- 
tions in mind. Ehrlich cites with approval a simile used by Emil 
Fischer, saying that the relation of the two groups must be that of 
lock and key. Granted, now, that certain food molecules have been 
anchored by fitting cell receptors, what follows? To explain this. 



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1913] Charles Frederick Bolduan 255 

Ehrlich makes use of an hypothesis advanced by Weigert in con- 
nection with regeneration. According to this, physiological func- 
tion and structure depend upon an equilibrium of the tissues that is 
maintained by virtue of mutual restraint between their component 
cells. Destruction of a single integer or group of integers of a 
tissue or a cell removes a corresponding amount of restraint at the 
point injured, and therefore destroys equilibrium. This permits 
of the abnormal exhibition of bioplastic energies on the part of the 
remaining uninjured components, which activity may be viewed as a 
compensating hyperplasia. When such bioplastic activity is called 
into play there is always hypercompensation ; that is, there is always 
more plastic material generated than is necessary to compensate for 
the loss. Thus far Weigert. 

Ehrlich, in line with Weigert's over production theory, points 
out that, owing to the combination of toxin with receptors of the 
cell, the receptors are practically lost (at least temporarily) to the 
cell; that the cell or its fellows now produces new receptors to re- 
place this loss ; but that this production always goes so far as to make 
a surplus of receptors ; that these receptors are thrown off by the cell, 
as unnecessary bailast so to speak, and then circulate in the blood as 
antitoxin. The same substance therefore, which, when part of the 
cell, combines with the anchoring group of the toxin, enabling this 
to act on the cell, when circulating free in the blood combines with 
and satisfies this anchoring group of the toxin, and prevents the 
poison from combining with and damaging the cells of the organism. 
It is obvious that this affords a complete explanation of specificity. 

If we now go back to our diagrams (plate 2) we shall see that 
all of the antibodies discussed above fit readily into this scheme. So 
far as the antitoxins are concerned, these would be merely receptors 
of the first order, thrust off from the cell and circulating in the 
serum. Agglutinins and precipitins would belong to the second 
Order; they have the active group as an integral part of the receptor. 
The hemolysins and bacteriolysins would be in the third order; 
fresh serum is active because it contains complement, but since 
the complement is very labile, the serum after a while contains only 
the immune body, i. e., that part of the receptor which anchors the 
food molecule on the one band and the ferment substance on the 
other. 



256 Imniunity in Some of its Biochemical Aspccts [Jan. 

Explanat'wn of natural immimity. Ehrlich's views concerning 
the necessity for fitting receptors in order that a microörganism may 
attack the body cells afford a satisfactory explanation of the im- 
munity possessed by certain animals against particular infections. 
Thus, it is obvious that the entire absence of receptors fitting a cer- 
tain microörganism renders the body immune against infection by 
that microörganism. Moreover, the location of the receptors may 
be responsible for the relative immunity of an animal under natural 
conditions and its susceptibility when these conditions are changed. 
Thus, if receptors for a particular poison are present both in a vital 
tissue, like the brain, and in an indifferent tissue, like the muscles, 
it is clear that while an intracerebral injection of the poison might 
prove fatal, an intramuscular one might be almost without effect. 

Anaphylaxis. Most of you are probably familiär with the 
work of Vaughan and Wheeler concerning the cleavage products of 
proteins, and recall that some of their products were highly poison- 
ous. Certain observations of the past few years indicate that, in the 
parenteral digestion of proteins, similar cleavage products are pro- 
duced. Historically this aspect of immunity may be said to date 
from 1906, from the studies undertaken by Ehrlich's pupil, Otto, 
and from experiments made about the same time in the U. S. Hy- 
gienic Laboratory by Rosenau and Anderson. In the course of the 
standardization of diphtheria antitoxin, it had been noted that 
guinea pigs, which had previously been injected with toxin-antitoxin 
mixtures, were often killed by a subsequent injection of horse 
serum. When the subject was studied it was found that, when 
an animal is injected with an allen protein, there develops after a 
time a specific hypersusceptibility for this protein. After a definite 
interval, if the animal is given a second injection of the same pro- 
tein, violent Symptoms occur, which may end fatally. The reac- 
tion is specific, so that animals sensitized, for example, to horse 
serum, manifest little or no hypersusceptibility to other sera. It is 
possible, however, to sensitize an animal to several proteins simul- 
taneously. The sensitizing dose may be very small — even as little 
as one-millionth of a cubic centimeter of horse serum has sufficed 
to render a guinea-pig sensitive. A varying length of time must 
elapse after the sensitizing injection before the animal becomes fully 



1913] Charles Frederick Bolduan 257 

sensitive. In guinea-pigs treated with small doses of horse serum, 
from 12 to 14 days suffice; with large doses, the time required is 
longer, and may extend over weeks or even months. In any case, 
in Order to produce severe Symptoms, it is important that the second 
injection be large, say 5 to 10 c.c. in a guinea-pig. This phenom- 
enon, spoken of as anaphylaxis, has come to occupy an important 
place in the theory of infection and immunity. What is the ex- 
planation of the phenomenon? 

Enteral and parenteral introdiictions of protein contrasted. We 
know that the subcutaneous, intraperitoneal, or intravenous intro- 
duction of alien protein is followed by the formation of antibodies ; 
at the same time it can readily be shown that no antibodies develop 
after the oral introduction of milk, eggs, or even of raw meat. In 
other words there is a marked contrast in the behavior of the body 
toward enteral and parenteral introductions of protein. In the 
former the protein is acted on by specialized cells, which, through 
their pepsin, trypsin and enterokinase, and erepsin, break down the 
protein molecule so that it loses its species identity. After this, 
absorption takes place, and with it there is a synthesis or rearrange- 
ment of the molecule whereby it is built up into protein specific to 
the body. Under normal conditions it is impossible to produce 
antibodies by feeding alien protein, though precipitins have been 
produced by overfeeding animals with large amounts of alien pro- 
tein. When protein is introduced parenterally it gives rise to the 
formation of specific antibodies. In the case of the sensitized ani- 
mals described above, the first injection causes the production of 
specific antibodies, among them specific cytolysins acting on the alien 
protein molecule. When the second injection comes, the alien pro- 
tein is at once laid hold of by this antibody, protein cleavage results, 
and with it the liberation of poisonous cleavage products. These 
cleavage products cause the severe Symptoms and even death that 
characterize anaphylaxis in guinea-pigs. That similar Symptoms do 
not arise in the enteral digestion of protein would then be explained 
by saying that, in the specialized digestive apparatus, provision has 
been made either for preventing the formation of such poisonous 
cleavage products or for neutralizing them (conjugations?) before 
they can cause injury. 



2S8 Imnmnity in Some of its Biochemical Aspects [Jan. 

Significance of the pcriod of inciibation in anaphylaxis. An in- 
teresting result of these studies on anaphylaxis is the light they shed 
on the significance of the period of incubation, and also on the poi- 
sonous Symptoms produced by bacteria f rom which no very poisonous 
siibstance can be extracted. Taking up first the latter point : It has 
been held that the various pathogenic bacteria, like the diphtheria 
and tetanus bacilli, either secrete toxins or, at least, contain such 
toxins bound np in their protoplasm. In the latter case, it was 
believed that these endotoxins, as they were termed, were set free 
during the destruction of bacteria in the body. From his studies 
on anaphylaxis, Friedberger concludes that it is entirely unneces- 
sary to assume the existence of specific endotoxins in bacteria to 
account for the various Symptoms seen in bacterial infections. By 
repeatedly injecting sensitized animals with minute doses of sheep or 
horse serum, he found it possible to produce all manner of fever 
curves at will, merely by varying the size of the dose and the inter- 
val between the injections. From this he concludes that the diver- 
sity of clinical Symptoms of various infectious diseases can readily 
be explained, even on the assumption of but a single poison. He 
speaks of this as anaphylatoxin, and regards it as a cleavage product 
of protein of whatever origin introduced parenterally. Just as in 
enteral digestion, uniform cleavage products are formed from most 
diverse proteins, so, he believes, that in the parenteral protein de- 
composition leading to the formation of anaphylatoxin, a certain 
poison is uniformly produced. Whether or not, in addition to 
anaphylatoxin, there are other specific poisons for the various in- 
fectious diseases is immaterial; their existence (except in certain 
diseases) has not been proved, and the assumption of their existence 
is unnecessary. According to Friedberger, the assumption of a 
common anaphylatoxin is only apparently in contradiction to the 
well known law of specificity of infectious diseases. In the infec- 
tious diseases it is not the poison which is specific, but only the mode 
of its production. The production of anaphylatoxin requires the 
action of antibodies; the mere Solution or disintegration of bacteria 
by other means does not suffice. In other words, a particular 
cleavage of the protein molecule is necessary. 

Bearing of the period of incubation on intoxication by bacterial 



1913] Charles Frederick Boldiian 259 

infection in general. With this conception of the effects of paren- 
teral protein cleavage, it is a simple matter to explain the significance 
of the period of incubation. For those of you who are not medical 
students, I will say that every infectious disease manifests itself only 
in a certain period of time after infection has taken place. More- 
over, this interval is fairly constant. Thus, after a person has con- 
tracted typhoid fever, some ten to fifteen days elapse before Symp- 
toms develop. In measles, the incubation is regularly fifteen to 
eighteen days; in scarlet fever, regularly from three to five days, 
etc. Formerly this period was explained as the time necessary for 
the development of germs in sufficient number to produce Symptoms. 
This explanation was unsatisfactory, because, in artificial in- 
fections, no matter how large the dose, it was never possible 
to shorten the incubation period below a certain minimum, and 
this minimum could not be explained. If, however, we regard in- 
fecting bacteria as protein introduced parenterally, we shall have no 
difficulty in explaining the incubation period as the time necessary 
for the body to develop antibodies which shall act on the bacteria 
and produce poisonous cleavage products. Even if we do not accept 
Friedberger's assumption of but a single anaphylatoxin, the same 
explanation holds for the liberation of endotoxins. In this connec- 
tion, I ought to say that bacteria invading the body through the in- 
testinal tract, e. g., the typhoid bacilli, may still be regarded as intro- 
duced parenterally, because they pass the intestinal barrier and gain 
access to the other tissues of the body. 

Modern chemotherapy according to Ehrlich. Before leaving 
the subject of infection and immunity, I should like to say a few 
words about the chemistry of the cell in relation to chemotherapy. 
I have already pointed out that Ehrlich holds that the action of a 
chemical substance on a given cell denotes the existence of definite 
chemical affinities between the substance and the cell. Applying 
this conception to the germicidal action of chemicals, he maintains 
that the latter must have a certain chemical affinity for the parasites 
in Order to kill them. Substances having such affinities he terms 
parasitotropic. It is clear, however, that substances which can de- 
stroy parasites will also be poisonous for the animal body, i. e., they 
will have chemical affinity for the tissues of the host. They are 



200 Immunity in So ine of iis Biochemical As pect s [Jan. 

therefore also Organotropie. In the employment of chemical sub- 
stances to combat infectious diseases, it follows that success can only 
be attained if the affinity of the chemical substances for the infecting 
parasite bears certain relations to their affinity for the infected body. 
Ehrlich's studies in this direction have, therefore, aimed to find 
poisonous substances whose parasitotropic affinity should be great 
in comparison to their Organotropie affinity. In his studies on Syph- 
ilis, he tested a very large number of substances, many of them com- 
binations of arsenic. As each substance was tested it received a 
serial laboratory number, and finally, in " 606," a substance was 
found which fitted the requirements to a high degree. This sub- 
stance, salvarsan, has produced really marvelous results in the treat- 
ment of syphilis. This line of work appears very promising. 

Chemical natura of antibodies. A closing word concerning 
the chemical nature of antibodies. Most of the studies have been 
made on diphtheria antitoxin, and although little is known concern- 
ing the Constitution of this substance, it seems proable that it is 
protein in character. Certain it is that the antitoxin is associated 
with the globulins of the serum, and highly concentrated Solutions 
of antitoxin have been prepared, by Gibson, by precipitating and then 
redissolving these globulins. Moreover, as Atkinson showed, the 
globulins increase markedly in the serum of immunized horses as 
the antitoxic strength of the serum increases. 

The influence which the development of the field of immunity 
has had on biochemistry has been tremendous, for it has contributed 
not only new view points, but also entirely novel methods. Much 
has been learned about substances which no one had ever yet seen 
and which we know only through their action. That all this has 
been achieved is due mostly to one master mind, Paul Ehrlich. May 
he long continue to lead us ! 



A PLAN FOR THE ORGANIZATION OF THE AMER- 
ICAN BIOLOGICAL SOCIETY^ 

ALBERT P. MATHEWS 

The present condition of the biological interests of the country 
may be called chaotic. There is no general Organization and little 
Cooperation between various subdivisions of the science; there are 
a multitude of small societies and a large number of Journals, few 
with any permanent support. This condition renders the science as 
a whole less effective in the Community than it ought to be, and is 
expensive both of time and money. The time has come to effect 
some kind of Cooperation of all biologists to secure the advantages 
which come f rom Cooperation. These advantages could be obtained 
by the formation of a general society, to be called the American 
Biological Society, along the lines of the American Chemical 
Society. (This Society might act as the Biological Section of the 
American Association for the Advancement of Science.) 

Objects of the society: (i) To unite the biological interests of 
the country for purposes of education; mutual support; increased 
Cooperation, defense and encouragement of scientific investigation; 
and to increase the influence of biological knowledge in the country; 
(2) to Start and support a Biological Abstract Journal; (3) to pro- 
vide for the permanent support of the biological Journals of the 
country and to provide for new ones as necessity arises ; (4) to 

^ This plan was proposed by Professor Mathews in 1908, in multigraphed 
circular form, to the members of the American Physiological Society. The plan 
was formally laid before the Council of the Physiological Society in December, 
1908, in the hope that the Physiological Society would endorse the essential fea- 
tures of the Suggestion. The author was appointed a committee of one to 
agitate the matter. Nothing further was done, however. Theunsuccessful eflfort 
in December, 191 1, to bring about an Organization of a greater American Physio- 
logical Society, and the recent formation of the Federation of American Societies 
for Experimental Biology, give new interest to Professor Mathews' plan, which 
is published here in its original form, at our reqiiest, and with the permission of 
the author. See pages 269 and 271 of this issue of the Biochemical Bulletin. 
[Ed.] 

261 



202 Organization of the American Biological Society [Jan. 

diminish the cost, to the members of the society, of dues to societies 
and siibscriptions for these Journals. 

Details of Organization. Membership. All members of the 
present biological societies should be eligible for membership with- 
out further action and should become members on payment of the 
dues. Such societies are those of Anatomy, Physiology, Zoology, 
Botany, Experimental Medicine, Pharmacology when organized, 
Psychology, Biochemistry, Bacteriology, and so on. All persons 
sufficiently interested in the progress of biology to pay the dues 
of the society should be eligible for membership. 

LocAL SECTiONS. The Constitution should provide for the for- 
mation of local sections in different cities, a certain per cent. of the 
dues of the members of such a local section to be repaid to the 
section for local expenses. 

Affiliation of present societies. The present societies 
should ultimately organize as sections of the Biological Society, 
thus saving extra dues. Membership in these sections might be 
determined by the sections themselves. 

Dues. Dues should be sufficient to provide that each member 
should receive the Biological Abstract Journal, and some or all of 
the other biological Journals. How this may be arranged is shown 
beyond (page 265 ) . The cost of the Journals should be much lower 
to the members of the society than to Outsiders. 

Explanation of the proposed plan. The plan presented in 
the foregoing Statements is virtually that adopted with such great 
success by the chemists of the country. A few years ago the chem- 
ists were in the position of the biologists today. There were sev- 
eral small societies; there was nominally a general Organization 
dragging out an unprofitable existence. There were several Jour- 
nals badly supported. The American Chemical Society was orga- 
nized and ultimately the smaller societies became convinced of the 
advantage of Cooperation. Now, most of them have become sec- 
tions of the general society. The growth of this society has been 
very rapid ; and it has grown in vigor as well as in size. Two years 
ago^ the society started a chemical abstract Journal and it is not too 
much to say that this has done more for the chemical interests 

*The reader is reminded that this was written in 1908. [Ed.] 



1913] Albert P. Mathews 263 

of the country than any other step taken. Chemical Abstracts has 
welded the various divisions of the science together, and so great 
has its value proved to be, that the membership in the society has 
almost doubled since it was started.^ The society publishes three 
Journals, Chemical Abstracts (the abstract Journal), the Journal of 
the American Chemical Society, and the Journal of Industrial 
Chemistry, which are distributed to all members of the society for 
the dues, $10 a year. Chemical Abstracts appears every two weeks ; 
the other two are monthly Journals. 

Relation of the society to the naturalists. Two possibilities 
are open to us in forming the Biological Society: we could make 
use of the American Society of Naturalists, reorganize that and 
change it into a new society with new aims; or we might Start a 
new society, leaving the " Naturalists " to f ulfill some other usef ul 
function (such as that adopted in their recent reorganization). The 
name of the " Naturalists " is badly chosen for a general biological 
society, such as that proposed; and, since its partial resuscitation 
along its old lines might weaken our efforts (if the two societies 
should Cover in any way the same field), it appears wiser to me to 
organize a new society, and to allow the " Naturalists " to have its 
aim changed to the one sketched in the plan of reorganization. 

Discussion of the objects of the proposed biological society. 
The importance of a biological abstract journal. ( i ) The 
objects in paragraph i, page 261, are so desirable as not to need dis- 
cussion. (2) The desirability of starting a Biological Abstract 
Journal, in English, has long been apparent. Funds alone have been 
lacking in the past to accomplish this object. The Organization of 
this society would make it possible to issue such a Journal. This 
would do more to unify and stimiilate biology than any move we 
could make. (3) How the ends sought in objects 2, 3, and 4 (page 
261) could be attained, will now be shown. 

List of the present biological Journals and their esti- 
MATED cosT AND PRiCE OF SUBSCRIPTION.^ The figures submittcd 
in this list are approximate only and are based on estimates supplied 
by various firms and individuals. The subscription list is a rough 

* The reader is reminded that this was written in 1908. [Ed.] 



264 Organization of the American Biological Society [Jan. 

estimate only. The cost is estimated on an edition of 500 copies. 

Estimated cost of Journals, containing tables and cuts, and 
printed on good paper with press work included; edition of 500 
copies: 12-point (a good sized body type) $1.40 a page; lo-point 
(used for reviews) $1.80 a page; 8-point (bibliography) $2.11 a 
page; 8-point (tables) $3.40 a page. 

Blank pages, and pages made up wholly of figures, $0.90. For 
half-tones, in the text, there is an extra charge of $1.00 each for 
makeready. For process-plates on coated paper: single plates, 
$4.00 ; double plates, $8.00. 

An additional 1,000 copies would increase the cost only for 
inserts, the press work and the paper, and may be estimated at about 
$500 or $600 a year, on a Journal of say 1,000 pages. The cost 
of a Journal is thus seen to be almost wholly the cost of putting it on 
the press, or the cost of its first 500 copies. 

Name of Journal. Subscription price per year Estimated 

on basis of present issues.- cost of 500 

copies. 

American Journal of Physiology $15 $7,000 

American Journal of Anatomy 5 3,000 

Journal of Comparative Neurology 4 2,000 

Journal of Morphology 9 3,000 

Journal of Infectious Diseases 5 3,000 

Journal of Experimental Medicine 5 2,500 

Journal of Medical Research 8 4,000 

Biological Bulletin 6 2,500 

Journal of Biological Chemistry 8 3^500 

Journal of Experimental Zoology 5 2,500 

Anatomical Record 3 2,000 

PsychologicalReview{hu.\\etmand'mdQx) 5 3,000 

Botanical Gazette 7 4,000 

Total number, 13 $85 $42,000 

Biological Abstract Journal 4,000 

American Journal of Psychology 5 2,000 

There are several other Journals which might be added to this 
list and there are a few Journals on the list which might be taken 

' The reader is reminded that this was written in 1908. [Ed.] 



1913] Albert P. Mathews ^ 265 

care of by special Institutes.^ It is obvious, however, that the biol- 
ogists have before them the problem of putting on a permanent foun- 
dation Journals costing about $50,000 a year. This can best be done 
by making each Journal seif supporting, and this is only possible by 
increasing the number of subscribers. 

HOW TG INCREASE THE NUMBER OF SUBSCRIBERS FOR THE JOUR- 
NALS. At present it costs, let us say, $28 a year to subscribe for the 
Journal of Physiology, the Journal of Biologkai Chemistry, and the 
Journal of Infectious Diseases. Each of these Journals probably 
has on the average a paid subscription list of something under 400.^ 
Membership in the corresponding societies costs, in addition, about 
$2 a year for each society, or a total yearly expense of $34. 

Now, if we could make a society of 2,000 members and charge 
each member $25 a year for all dues or, to be more liberal, let us say 
$30 a year, the society would have an annual income of $60,000 ; and 
for this sum, it could publish and supply to its members not three 
but thirteen Journals without further cost. Moreover, each of these 
Journals would have a large circulation, beneficial alike to the man 
who published in it and to the Journal itself. Furthermore, the 
amount of the individual society expenses would be greatly re- 
duced since, by proper Organization, one or two paid secretaries 
would look after notices of meetings; bills for postage, announce- 
ments, programs, etc., would be less; and a saving would be effected 
all along the line, with a great gain in efficiency. 

The income of the Journals would also be augmented beyond 
the dues by the constant sale of back numbers, the sale of extra re- 
prints and, in some cases, by legitimate advertising. Moreover, 
by keeping the present prices in effect for all non-members, nearly 
everyone would hasten to join the society ; thus increasing our num- 
bers and increasing the number of those among whom the expense 
would be divided, and making it possible, from time to time, to Start 
new special Journals with little increase in expense to the members. 
Furthermore, by maintaining the present prices to libraries and for- 
eign subscribers, a considerable sum would be added to the treasury, 
For example, the present cost of these Journals to subscribers is $83 
a year. If there were a hundred subscribers at this price, it would 
add $8,300 to our income. Of course the total may easily be less 

* The reader is reminded that this was written in 1908. [Ed.] 



266 Organisation of the American Biological Society [Jan. 

than this, but it will certainly amount to half that sum, since there 
must be fifty libraries subscribing at the old rate. 

How to get the necessary 2000 members. As a nucleus of 
the Society all members of the biological societies would probably 
join. There are 1,500 names in Cattell's American Men of Science 
vvho would be eligible and upon whose support we might confidently 
count. In addition probably 300 have joined the ranks of biology 
since that publication was issued or whose names were omitted 
through oversight. Let us say, at a liberal estimate, 1,800 all told. 
There probably could be found in addition 500 intelligent and public 
spirited physicians, and others sufficiently interested in biology, to 
join such a society with such great advantages in the matter of Jour- 
nals. These figures are maximum figures but they suffice to show 
that we could count on perhaps a thousand members at the start; 
and there is no doubt that the numbers would increase rapidly, just 
as they have done in the Chemical Society. We might also soon 
Start a Journal of Biological Industries, or in other ways increase 
Cooperation between the practical applications of biology and the 
science itself. 

Other arguments might be presented, but these suffice to show 
the great advantages of Cooperation and to make it evident that, in 
this way, we could attain these desirable objects: increase the in- 
fluence of biology, increase Cooperation; knit the science together, 
strengthen its practical applications; start a Biological Abstract 
Journal; provide for the support, and enlarge the usefulness, of 
our present Journals, and provide for new ones as the need arises; 
and diminish the cost, to each one of us, of subscriptions and dues. 

We should also accomplish more than this, for, by such an Organ- 
ization, we should be providing for the future, and organizing with 
the object of attaining certain well defined ideals. Whatever Organ- 
ization is attempted at this time should have in view the practical 
attainment of these ideals and should not be a mere repetition of 
what we have, with no definite plan and without foresight. 

In view of the foregoing facts I move the adoption of the fol- 
lowing: That the American Physiological Society expresses its ap- 
proval of the objects sought in the plan presented for the forma- 
tion of the American Biological Society; and it recommends. 



1913] Albert P. Mathews 267 

further, that the Society transmit to the other societies copies of this 
plan with the request that the plan be presented to the members of 
the societies; that each society appoint one delegate to meet mem- 
bers appointed by the other societies to act as a committee of Or- 
ganization of the American Biological Society; and that such 
committee shall carefully examine into the feasibility of such an 
Organization and, if possible, draft a Constitution and report to the 
societies at their next annual meeting. 

Suggestions for carrying out, practically, the Journal part of 
the plan. ( i ) It will possibly be f ound that $30 or $20 a year is 
more than the majority of the Society feel able or willing to pay. 
Arrangements could be made whereby at a somewhat larger relative 
cost such members could subscribe to two, three, or half a dozen of 
the Journals as they desired. Arrangements could be made with the 
Journals whereby copies would be sent to the members of the Society 
at a reduced price, if a certain number of subscribers was received 
in this way. For example, the Society might offer the Biological 
Abstract Journal, and any two others, for $10 a year ; the Biological 
Abstract Journal, and five others, for $20 a year; and the whole 
number, say, for $30 a year. In this way there would always be 
an incentive for the members, who could not at the Start pay the 
füll sum, to increase their subscriptions and thereby enable everyone 
to get his subscription at a reduced cost. It would not, however, be 
possible on this basis to give so much to the members as if all 
subscribed to all the Journals, but still a great reduction of cost could 
be obtained. The object aimed at should be to increase as rapidly 
as possible the numbers of those taking the whole number of 
Journals. 

(2) The relation of the Society to the management of the Jour- 
nals would, of course, have to be worked out gradually. Several 
courses are open to the society. One is to leave the Journals as they 
are under their present control and for the Society to make such 
arrangements with the Journals as would be most advantageous to 
the members. This is the club-rate principle, the society buying so 
many copies at a reduced rate to distribute to its members. This. 
arrangement might do as a temporary makeshift, to get started, but 
would probably be unsatisfactory in the long run, since it would not 
be permanent enough. 



268 Organization of the American Biological Society [Jan. 

The Society might take over the financial responsibihty of such 
Journals as the Council of the Society deemed best; beginning, for 
example, with one or two with the largest circulation, adding the 
Biological Abstract Journal, and Publishing the three for $io or 
$12 a year, and distributing them to all its members. Then, as the 
Journals wished and the Council and the Society decided, one after 
another of the other Journals could be added until the whole list was 
included. This scheme would be feasible if we had a thousand 
members at the start. In any such arrangement the editorial boards 
of the Journals would retain entire charge of the editorial manage- 
ment, so that the independence of the Journals would be secured. 

(3) If such an arrangement could be made with the Wistar In- 
stitute of Anatomy, it might become the Publishing house for the 
Society, taking over the financial responsibihty for additional Jour- 
nals, as the Institute has already done for several, and thus greatly 
extending the usefulness of the Institute. In this way the Society 
would aid the Institute in getting the Journals on a firm basis by 
uniting in its support the biological interests of the country. At 
the Start this plan might involve an increased outlay by the Wistar 
Institute, but, in the long run, the dues of the Society should suffice 
to maintain the Journals. This plan would aid the Wistar Institute 
in doing the work it has undertaken. 

(4) Provision could be made for the starting of new Journals 

at any time, or for the support by the Society of those established by 

Outsiders. 

University of Chicago, 
Chicago, Illinois. 



ORGANIZATION OF THE FEDERATION OF AMER- 
ICAN SOCIETIES FOR EXPERIMENTAL BIOLOGY^ 

Comprising the American Physiological Society, the American 

Society of Biological Chemists, and the American Society 

for Pharmacology and Experimental Therapeutics 

JOHN AUER 

Among the most enjoyable features of the recent meetings at 
Cleveland (pages 271, 275, and 279) were the subscription dinners 
and smokers, held on the evenings of December 30 and 31, at the 
Colonial Hotel. These informal dinners were attended by the 
pharmacologists, physiologists and biochemists, and a pleasant 
flavoring of naturalists, zoologists and anatomists. 

At the last of these dinners perhaps the most important develop- 
ment of the Cleveland sessions, so far as the pharmacological, 
physiological and biochemical societies are concerned, took place. 
At this dinner, delegates f rom the three societies, empowered to act, 
met in Conference on the formation of an alliance which should more 
closely knit together the three societies zvhile yet jealoiisly preserving 
the individtiality of each compotient Organisation. The delegates 
f rom the Physiological Society were Drs. Meltzer, Lee and Cannon ; 
from the Biochemical Society, Drs. Lusk and Wells ;2 from the 
Pharmacological Society, Drs. Sollmann, Loevenhart and Auer. 

Dr. Meltzer was elected temporary chairman and Dr. Cannon 
temporary secretary. The outcome of the proceedings of this Con- 
ference committee can best be shown by a transcript of its minutes. 
The f ollowing motions were voted unanimously : 

That a Federation of the three societies be hereby established. 

^ This account was presented, originally, as a part of Dr. Auer's report of the 
proceedings of the Society for Pharmacology and Experimental Therapeutics, 
page 279. [Ed.] 

^Dr, Gies, the third delegate from the Biochemical Society (page 278), was 
unable to attend the Cleveland meetings because of the illness of his eldest son. 

269 



270 Fcderation of American Biological Societics [Jan. 

That the presidents and the secretaries of the constituent societies 
form the executive committee of the Federation. 

That the chairmanship of the executive committee be held in turn 
by the presidents of the constituent societies who shall succeed one 
another annually in the order of seniority of the constituent societies 
(Physiological, Biochemical, Pharmacological). 

That the secretary of the society whose president is chairman 
shall be the secretary of the executive committee. 

That the secretaries of the three societies shall consult in pre- 
paring the programs of the annual meeting, and that, so far as prac- 
ticable, and with the authors' consent, papers be so distributed as to 
be read to the society in which they properly belong. 

That the programs of the three societies be published by the 
secretary of the Federation under one cover and that the expense 
of publication be shared pro rata by the societies according to the 
number of members. 

That the official title of the new Organization be the "Federa- 
tion of American Societies for Experimental Biology : Comprising 
the American Physiological Society, the American Society of Bio- 
logical Chemists, and the American Society for Pharmacology and 
Experimental Therapeutics." 

That a common meeting place of the Federation with the socie- 
ties of Anatomists, Zoologists and Naturalists is desirable but not 
mandatory. 

That, in the name of the Federation, the International Physio- 
logical Congress be invited to meet in the United States in 191 6. 

That the present Conference committee delegate all its powers to 
the executive committee of the Federation. 

The first meeting of the new Federation will be held in Decem- 
ber, 191 3, in Philadelphia. 

Rockefeller Institute for Medical Research, 
New York City. 



ANNUAL MEETINGS OF THE ORGANIZATIONS COM- 

PRISING THE FEDERATION OF AMERICAN 

SOCIETIES FOR EXPERIMENTAL 

BIOLOGYi 

Proceedings reported by THE Secretaries, 

JOSEPH ERLANGER, A. N. RICHARDS, and JOHN AUER 

I. THE AMERICAN PHYSIOLOGICAL SOCIETY 

Joseph Erlanger^ 

The Society held its twenty-fifth annual meeting in the Medical 
Building of Western Reserve University, Cleveland, Ohio, Decem- 
ber 29, 1912 to January i, 1913. Sixty-nine members were in at- 
tendance. Two executive sessions and six scientific sessions were 
held, two of the latter being Joint sessions, one each with the Amer- 
ican Society of Biological Chemists and Section K of the American 
Association for the Advancement of Science. The Joint session 
with the American Society of Biological Chemists was opened with 
exercises in memory of the late Waldemar Koch. After the mem- 
bers of the Society had arisen as a token of respect to the memory 
of Doctor Koch, Prof. A. P. Mathews delivered a memorial address. 

Papers and demonstrations. The titles of the papers and 
demonstrations, fifty-two in all, which were read and discussed, with 
the names of the authors, are appended : 

S. Simpson: The rate of growth in the dog. — G. N. Stezvart: 
Further observations on the blood-flow in man. — /. A. E. Eyster 
and W. J. Meek: Experiments on the sinus region of the mammalian 
heart. — G. C. Robinson (by invitation) and /. Aner: Cardiac 
anaphylaxis as shown by the string galvanometer. — W. T. Porter: 
The functional relations of cells in nerve centers. — R. S. Lillie: 
Correlation between the anti-stimulating action and the anti-cyto- 

^ See report on the recent Organization of the Federation, page 269. 
''Acting Secretary, vice Dr. A. J. Carlson, unavoidably absent. 

271 



2/2 Anmial Meetings of Federated Socicties [Jan. 

lytic action of anesthetics. — E. B. Meigs: Studies inthe general phys- 
iology of smooth muscle. — W. P. Lombard: The tickle sense. — O. 
FoUn, W. B. Cannon, and W. Denis (by invitation) : A new colori- 
metric method for the determination of epinephrin. — /. Aiier and 
5. /. Meltser: The splanchnic as a depressor nerve. — F. R. Miller: 
The saHvary secretion centers in the medulla. — M. Dresbach (by 
invitation); A bloodless method of recording blood pressure in 
animals. — W. T. Porter: A new electrica! clock. — S. P. Beebe: A 
new form of apparatus for artificial respiration. — A. D. Hirsch- 
felder: Some new apparatus. — R. S. Hoslins: Relation of fatigue 
metabolites to epinephrin efficiency. — D. R. Hooker: Perfusion of 
the respiratory center in f rogs ; the influence of calcium and potas- 
sium on the respiratory rhythm. — A. Hunter: The nitrogen excretion 
of normal and of thyroidectomized sheep. — A. L. Tatum (by in- 
vitation) : Studies in experimental cretinism with suggestions as to 
a biological test for thyroid secretion. — R. Gesell (by invitation) : 
The relation of pulse pressure to renal secretion. — C. Brooks and 
A. B. Lnckhardt: The arterial blood pressure during vomiting. — 
T. Sollmann and /. D. Pilcher (by invitation) : The effects of aortic 
compression on the circulation. — E. G. Grey (by invitation) and A. 
D. Hirschfelder: Clinical observations upon the carbon dioxide per- 
centage of alveolar air. — C. W. Greene and W. Y. Skaer (by invi- 
tation) : On the fat contents of the mammalian gastric glands in re- 
lation to the stages of digestion. — 5". Toshiro (by invitation).- The 
chemical change in nervous tissue during excitation. — /. F. Zucker 
(by invitation) : The pressor property of shed blood. — H. Cushing, 
L. H. Weed (by invitation) and C. Jacobsen: Further studies on 
the role of the pituitary gland in carbohydrate metabolism, with 
special reference to the autonomic control of the posterior lobe secre- 
tion. — S. A. Matthews and D. D. Lewis (by invitation) : The pars 
intermedia; its place in Diabetes insipidus. — Lydia M. Degner (by 
invitation) and A. E. Livingston (by invitation) : Effects ofthyroid- 
ectomy and castration, respectively, on the pituitary in the rabbit. 
— P. W. Cobb and L. R. Geisler (by invitation) : The influence on 
foveal vision of the brightness of surroundings. — D. E. Jackson: 
Some observations on the peripheral action of certain drugs. — G. L. 
Kite (by invitation) : The relative permeability of the surface and 



1913] Joseph Erlanger 273 

the interior portions of the cytoplasm of animal and plant cells. — 
— J. D. Pilcher (by invitation) : The excretion of nitrogen subse* 
quent to ligation of successive branches of the renal arteries. — W. E. 
Biirge: The uniform rate of destruction of ptyalin and pepsin by the 
electric current. — G. H. Whipple: Hematogenous jaundice and its 
relation to the liver. — 5". /. Meltzer: Is the pulsation of the anterior 
lymph hearts responsible for the action of some drugs in cardiec- 
tomized frogs? — H. McGnigan: The Synergie action of morphin 
and strychnin. 

Joint programs. With Section K (Physiology and Experi- 
mental Medicine) of the American Association for the Advancement 
of Science : page 277 ; with the American Society of Biological 
Chemists : page 275. 

The following ten papers were read by title: — C. D. Snyder: 
The influence of temperature on the mammalian heart. — A. J. 
Carlson: Some observations on the physiology of the empty stom- 
ach and esophagus in man and dog. — H. C. Bradley: The problem 
of enzyme synthesis. — G. W. Crile: The relation between the phys- 
ical State of the brain cells and brain functions ; experimental and 
elinical. — F. Henderson and C. T. Flynn (by invitation) : Oligemia 
in acute disease. — H. McGuigan: The secondary depression by 
epinephrin; the rate of destruction of the pressor and the hypergly- 
cemic actions of epinephrin. — W. B. Wherry (by invitation) : On 
the transformation of amoebae into flagellates and vice versa. — P. 
E. Howe (by invitation) and P. B. Hazvk: The influence of fasting 
on the creatine content of muscle. — C. D. Snyder: A study of the 
electromyograms. — A. J. Carlson: The correlation of the physiolog- 
ical States of the thyroid of the fetus and of the mother. 

New members: G. C. Robinson, Rockefeiler Institute for Med- 
ical Research. — /. D. Pilcher, P. J. Hanzlik, R. S. Pearce, Western 
Reserve Medical School. — E. C. Schneider, Colorado College. — A. 
H. Ryan, University of Pittsburgh. — M. Dresbach, Cornell Uni- 
versity. — G. Bachmann, Atlanta, Ga. — H. G. Barbour, Yale Medical 
School. — W. DeB. MacNider, University of North Carolina. — A 
R. Moore, University of California. — H. B. Williams, Columbia 
University — V. H. K. Moorhonse, Washington University. 

The federation. At this meeting considerable progress was 



274 Anmial Meetings of Federated Societies [Jan. 

made toward the formation of a close federation of the American 
Physiological Society, the American Society of Biological Chem- 
ists and the American Society for Pharmacology and Experimental 
Therapeutics. The Society expressed its desire to enter into such a 
federation, and a committee was appointed to confer with similar 
committees of the sister societies with a view to bringing about such 
a federation. The committee was granted power to make the ar- 
rangements for the next annual meeting. This committee was also 
directed to confer with a similar committee of the American Society 
of Naturalists to consider the advisability of establishing closer rela- 
tions with that society (page 278). 

Future programs. With regard to the measures of remedying 
the threatening congestion of programs that were referred to the 
Council at the last annual meeting, it was decided that should the 
federation of the three societies be accomplished (page 269), the 
secretaries of the federated societies be empowered to attempt the 
equalization of the programs of the three societies by placing papers 
on the program of the society to which its subject is most closely 
related. It was also decided to place at the end of the program 
papers presented by non-members, and, in the event of congestion 
of the program, to read these by title. 

Officers-elect. The following officers were elected for the year 
1913: 

President — S. J. Meltzer; Secretary — A. J. Carlson; Treas- 
URER — Joseph Erlanger. 

Additional memeers of the COUNCIL — W. B. Camion and 
F. S. Lee. 

Editorial committee on the publication of the American 
Journal of Physiology for 1913 — W. T. Porter, A. J. Carlson, 
Joseph Erlanger, W. H. Howell, F. S. Lee, Graham Lusk, S. J. 
Meltser. (Appointed by the president.) 

Local entertainment. The local Committee on Enterta%iment, 
following the plan that was first tried last year at Baltimore by 
the members and friends of the Society, again agreed to dispense 
with all private entertainment, and to Substitute for it informal 
subscription dinners followed by smokers each evening while the 
Society was in session. These functions were open to all members 



1913] Alfred N. Richards 275 

and guests of the Societies of the Experimental Biological Sciences. 
It was again demonstrated that this method of entertainment, by 
bringing all of the members together under conditions permitting 
informal discussion and exchange of ideas, adds greatly to the 
pleasure and value of the meeting. 

Abstracts of the papers. The abstracts of the papers will be 
published in the February number of the "American Journal of 
Physiology. 

Washington University Medical School, 



IL THE AMERICAN SOCIETY OF BIOLOGICAL CHEMISTS 

Alfred N. Richards 

The sessions of the seventh annual meeting of the American 
Society of Biological Chemists were held in the medical buildings 
of Western Reserv^e University, Cleveland, Ohio, December 30, 
1912-January I, 191 3. The scientific programs, which were of ex- 
ceptional interest, are appended. 

First Session. December 30, 9 a. m. Presiding officer: 
President A. B. Macallum. 

A. B. Macallum: Presidential address, on The energy of muscu- 
lar contraction; thermodynamic or chemodynamic ? — M. H. Givens 
and A. H. Hunt er: The excretion of pure catabolites in sundry types 
of mammalia. — O. Polin and W. Denis: The occurrence of uric acid 
in blood. — L. J. Henderson and W. W. Palmer: Studies of the ex- 
cretion of acid. — A. E. Taylor and A. I. Ringer: On the utilization 
of ammonia nitrogen in the protein metabolism. — W. McK. Mar- 
riott: The determination of acetone substances in blood and tissues 
by micro methods. — *jF/. McGuigan and F. C. Becht: The com- 
pression of the lung by inert gases. — /. Rosenhloom: A new method 
for drying tissues and fluids. — W. N. Berg: Surface tension in 
muscle contraction. 

Second session. December 31, g.oo a. m. Joint Session 
wiTH THE American Physiological Society. Presiding offi- 
CERS : President A. B. Macallum and President S. J. Meltzer. 

* Read by title. 



276 Anmial Meetings of Federated Societies [Jan. 

A. P. Mathews: Memorial address on Waldemar Koch. — L. 
Loeb: The infliience of pregnancy on the cydic changes in the 
Uterus. — G. Ltisk: Metabolism of a dwarf. — H. S. Gasser (by in- 
vitation) and A. S. Loevcnhart: The mechanism of Stimulation by 
oxygen want. — T. B. Osborne and L. B. Mendel: Feeding experi- 
ments relating to the nutritive value of the proteins of maize. — 
A. I. Ringer: The fate of fatty acids in diabetic organisms. — A. B. 
Macallum and W. R. Campbell: On the secretion of pure acid by 
the kidney (with demonstration). — D. Marine: Hypertrophy and 
hyperplasia of the parathyroid in birds. — G. H. Whipple: Intestinal 
obstruction; study of a toxic substance present in the intestinal 
mucosa. — E. V. McCollum: The influence of the plane of protein 
intake on nitrogenous retention in the pig. 

Third session. December 31, 2.00 p. m. Presiding offi- 
cer: President A. B. Macallum. 

*W. Salant and /. B. Rieger: Further observations on the influ- 
ence of cafTein on creatin and Creatinin metabolism. — *//. H. Bunuel: 
Quantitative oxidase measurements. — *H. S. Reed: The regulating 
function of amylase in the fungus, Glomerella. — A. P. Mathews: 
A new method of determining valence based on molecular cohesion. 
— H. G. Wells: The entrance of chemical substances into diseased 
tissues. — H. C. Bradley: The problem of enzyme synthesis. — R. T. 
Woody att (by invitation) : Certain 3-carbon atom complexes in 
metabolism. — *£. G. Hastings and E. B. Hart: The presence of a 
lactic acid producing enzyme in Bact. lactici acidi. — E. V. Mc- 
Collum and Marguerite Davis: The influence of the composition 
and amount of the mineral content of the ration on growth. — H. C. 
Bradley: Connective tissues of Limulus. — S. Tashiro (by invita- 
tion) : A new method for the detection of minute amounts of carbon 
dioxid. — *L. H. Davis and A. D. Emmett: A study of the chemical 
changes in meats during the process of drying by the vacuum 
method. — *H. T. Leo and P. E. Howe: Muscle creatin; dialysis of 
creatin f rom dog muscle. — *N. Stadtmüller, M. Kahn and /. Rosen- 
bloom: Studies on sulfur metabolism; the urinary sulfur partition in 
various diseases. — *£. V. McCollum and H. Steenbock: The meta- 
bolic end-products of the lipoid nitrogen of tgg yolk. 

* Read by title. 



1913] Alfred N. Richards 277 

Fourth Session. January i, 2.30 p. m. Joint Session with 
Section K of THE American Association for the Advance- 
MENT OF Science, and the American Physiological Society. 
Presiding officer : Professor J. J. R. Macleod. 

Symposium. Some recent applications of physical chemistry 
in biology — {A) A. B. Macallum: Surface tension; {B) L. J. Hen- 
derson: The control of neutrality in the animal body; (C) A. S. 
Loevenhart: The physical chemistry of enzyme action. 

Memorial addresses. In the presidential address at the open- 
ing of the first Session, President Macallum made extended refer- 
ence to the life, character, and achievements of the late Waldemar 
Koch, a charter member of the Society. At the opening of the Joint 
Session with the American Physiological Society, Prof. A. P. 
Mathews delivered a memorial address on Professor Koch. 

New members: Louis Baumann, University Hospital, Iowa 
City; F, J. Birchard, U. S. Department of Agriculture; Samuel 
Bookman, Mt. Sinai Hospital, New York; E. D. Clark, Cornell 
University Medical College; H. J. Corper, University of Chicago; 
A. A. Epstein, Mt. Sinai Hospital, New York; M. S. Eine, N. Y. 
Post Graduate Medical School; Isidor Greenwald, Montefiore 
Home, New York ; W. H. Howell, Johns Hopkins Medical School ; 
N. W. Janney, The Herter Laboratory, New York; /. S. Kleiner, 
Rockefeller Institute for Medical Research ; P. A. Kober, Roosevelt 
Hospital, New York; E. C. Koch, University of Chicago; Leo 
Kristeller, Berlin, Germany ; E. B. La Borge, Rockefeller Institute 
for Medical Research; A. P. Lothrop, Columbia University; W. 
DeB. MacNider, University of North Carolina; A. L Ringer, Uni- 
versity of Pennsylvania; W. C. Rose, University of Pennsylvania; 
E. C. Schneider, Colorado College ; Harry Steenbock, University of 
Wisconsin; R. T. Woodyatt, University of Chicago. 

Officers-elect. The following officers were elected for the year 
1913: 

President — A. B. Macallum; Vice-president — Graham Lusk; 
Secretary — Philip A. Shaffer; Treasurer — Donald D. Van Slyke. 

Additional members of the COUNCIL — H. P. Armsby, Lafa- 
yette B. Mendel, H. Gideon Wells. 

Nominating committee — Carl L. Aisberg, H. D. Dakin, P. B. 



2/8 Annual Meetings of Federated Societies [Jan. 

Hau'k, Reid Hunt, Walter Jones, T. B. Oshorne, A. N. Richards, 
H. C. Shernian, F. P. Underhill. 

Special committees. President Macallum appointed William 
J. Gies, Graham Lusk and H. Gideon Wells a committee to confer 
with similar committees from the American Physiological Society 
and the American Pharmacological Society concerning the forma- 
tion of a federation of the three societies having for its object "the 
establishment of a stable connection between the three societies, for 
the purpose of fixing the time and place of the annual meetings, 
the arranging of Joint sessions whenever possible, and in general 
to establish, officially, closer scientific and social affiliations between 
the sister societies, while retaining their individual independence." 
This committee was further empowered to act upon such matters 
connected with the proposed federation as should require decision 
before the next annual meeting of the Society, and was also author- 
ized to confer with representatives of the American Society of 
Naturalists concerning closer affiliation with that Society (p. 274). 

The Committee appointed at the sixth annual meeting to prepare 
a report concerning the nomenclature of the lipoids reported prog- 
ress. Professor Leathes, a former member of the committee, was 
appointed chairman to succeed the late Professor Koch, Dr. E. K. 
Dunham was appointed to the vacancy created by Professor Koch's 
death, and Dr. P. A. Levene to the vacancy created by Dr. Jacques 
Loeb's resignation. The members of the reconstructed committee 
are J. B. Leathes, chairman, H. D. Dakin, E. K. Dunham, WiUiam 
J. Gies and P, A. Levene. 

Vote of thanks. A unanimous vote of thanks was extended by 
the Society to Professors Macleod, Sollmann, Stewart and Pearce, 
and to the members of the "Local Committee," for the hospitality 
which the Society enjoyed. 

Attendance. The following members were present at one or 
more of the sessions : J. J. Abel, Samuel Amberg, S. P. Beebe, W. 
N. Berg, W. R. Bloor, H. C. Bradley, H. J. Corper, Otto Polin, 
W. E. Garrey, H. D. Haskins, Shinkishi Hatai, R. A. Hatcher, P. 
B. Hawk, L. J. Henderson, A. H. Hunter, J. B. Leathes, A. S. 
Loevenhart, Graham Lusk, A. B. Macallum, J. J. R. Macleod, 
W. DeB. MacNider, W. McK. Marriott, A. P. Mathews, H. A. 



1913] John Aller 279 

Mattill, E. V. McCollum, F. H. McCrudden, L. B. Mendel, V. C. 
Myers, H. S. Raper, A. N. Richards, A. I. Ringer, E. W. Rock- 
wood, Jacob Rosenbloom, L. G. Rowntree, Torald Sollmann, H. C. 
Wells, R. T. Woodyatt. 

Abstracts of the papers. Abstracts of the papers will be pub- 
lished in the March number of the Journal of Biological Chemistry. 

University of Pennsylvania. 

III. THE AMERICAN SOCIETY FOR PHARMACOLOGY AND EXPERI- 

MENTAL THERAPEUTICS. 

John Auer 

The fourth annual meeting of the Society was held in the Med- 
ical Building of Western Reserve University, Cleveland, Ohio, on 
December 30 and 31, 1912. The scientific programs are appended: 

First Session. December 30, 9.00 a. m. — *W. Salant: The 
influence of temperature on the toxicity of caffein. — *fF. Salant: 
Further observations on the influence of cafTein on the circulation. 
— S. P. Beebe and Eleanor Van Alstyne: The effect of high protein 
diet on the growth of transplantable tumors of the white rat. — L. B. 
Mendel and R. L. Kahn: The physiological action of some methyl 
purins. — /. A. E. Eyster and W. J. Meek: The action of certain 
drugs on the electrocardiogram. — P. J. Handik (by invitation) : 
The intestinal absorption of alcohol. — P. J. Handik (by invitation) : 
The "toxic dose" of salicylates according to clinical statistics. — 
W. H. Brown and A. S. Loevenhart: The effect of hematin upon the 
circulation and respiration. — W. DeB. MacNider: The effect of 
anesthetics on the Output of urine in uranium nephritis. — G. B. 
Roth: The physiological assay of aconitin. 

Second session. December 30, 2.00 p. m. L. G. Rozvntree 
and R. Fitz: Renal function in experimental passive congestion. — 
R. Fits and L. G. Rowntree: The effect of temporary occlusion of 
renal circulation on renal function. — W. W. Ford: Observations on 
three poisonous fungi not previously described. — /. D. Pilcher: The 
protective action of lipoids against hemolysis. — H. G. Barboiir (by 
invitation) : The action of histamin upon surviving arteries. — G. W. 
* Read by title. 



28o Annual Meetings of Federated Societies [Jan. 

Crile and /. B. Austin: Nitrous oxide sleep compared with normal 
sleep; brain cell studies. — W. T. Porter and /. H. Pratt: The action 
of diphtheria toxin on the vasomotor centre. — H. Nogttchi and 
/. Bronfenbrenner: The effects of certain disinfectants and therapeu- 
tic preparations upon the cultivated spirochetes. — F. M. Surface 
(by invitation) : The effect of surplus cow serum on complement 
fixation with infectious abortion. — */. Adler and C. L. Aisberg: 
Studies lipon the long continued administration of adrenalin and 
nicotin. — *C L. Aisberg: The hemolytic power of various plants. 

Third session. December 31, 9.00 a. m. — *F. H ender son: 
Demonstration of a carbonator for quantitative carbon-dioxide 
therapy. — P. Lcivis: Further observations on the relations of vital 
stains to the tubercle. — T. S. Githens and 6". /. Meltser: On the 
course of the toxic effects of ether and Chloroform under intra- 
tracheal insufflation. — T. S. Githens: On the influence of decerebra- 
tion upon morphin tetanus in frogs. — /, 5*. Kleiner (by invitation) : 
On the effect of sodium bicarbonate and sodium chlorid upon the 
convulsions produced by heroin and strychnin. — /. Aiier and 5. /. 
Meltzer: The influence of pituitrin upon the depressor action of the 
vagus nerve in cats. — B. T. Terry: The influence of heat upön the 
toxicity for trypanosomes of blood containing transformed atoxyl. 
— B. T. Terry: Variations in the toxicity of transformed atoxyl for 
trypanosomes, caused by altering the number of organisms. 

Officers-elect. The following officers were elected for 1913 : 

President — Torald Sollmann; Secretary — JohnAuer; Treas- 
URER — A. S. Loevenhart. 

New members of the Council — /. 7. Abel and Wm. DeB. 
MacNider. 

Membership committee — C. W. Edmunds was reelected to 
serve three years, and the place made vacant by Dr. Sollmann's elec- 
tion to the presidency was filled by the election of Reid Hunt. 

New members. Among the candidates for membership under 
investigation by the Membership Committee, the following were 
favorably reported to the Council, recommended for election, and 
elected by the Society : H. G. Barboiir, Yale University ; Clyde 
Brooks, University of Pittsburgh ; Gary Eggleston, Cornell Uni- 

* Read by title. 



1913] John Auer 281 

versity Medical College; P. J. Hanslik, Western Reserve Univer- 
sity; D. E. Jackson, Washington University; /. 5'. Kleiner, Rocke- 
feiler Institute for Medical Research; O. H. Plant, University of 
Pennsylvania; A. H. Ryan, University of Pittsburgh; F. P. Under- 
hill, Yale University. 

Attendance. The following members were present at one or 
more sessions of this meeting: J. J. Abel, Samuel Amberg, John 
Auer, S. P. Beebe, E. D. Brown, G. W. Crile, J. A. E. Eyster, 
W. W. Ford, T. S. Githens, C. W. Greene, Worth Haie, R. A. 
Hatcher, V. E. Henderson, A. W. Hewlett, A. D. Hirschfelder, 
D. R. Hooker, D. R. Joseph, P. A. Lewis, A. S. Loevenhart, W. 
DeB. MacNider, S. J. Meltzer, L. B. Mendel, J. D. Pilcher, J. H. 
Pratt, A. N. Richards, G. B. Roth, L. G. Rowntree, Torald Soll- 
mann, G. N. Stewart, B. R. Terry. 

Vote of thanks. At the last meeting the Society passed a vote 
of thanks to the Western Reserve University for the hospitality ex- 
tended and to the " Local Committee," Drs. Macleod, Sollmann and 
Pearce, for their thorough arrangement of all the details which 
made the Cleveland meeting so pleasant. 

Abstracts of the papers. Abstracts of the papers will be pub- 
lished in the March number of the Journal of Pharmacology and 
Experimental Therapeiitics. 

[The report on the Organization of the Federation of Amer- 
ican SociETiES FOR ExPERiMENTAL BiOLOGY, which appears on 
pages 269-70, was taken bodily from Dr. Auer's account of the pro- 
ceedings of the Pharmacological Society. (Ed.)] 

Rockefeiler Institute for Medical Research. 



MEETING OF THE AMERICAN SOCIETY OF 
ANIMAL NUTRITION 

(American Society o£ Animal Production) 

PROCEEDINGS REPORTED BY 

LEWIS W. FETZER 

The annual meeting of the American Society o£ Animal Nutri- 
tion was held at Chicago, 111., on November 30, 191 2. The ad- 
dress of the President, H. J. Waters, of the Kansas State Agricul- 
tural College, at Manhattan, Kansas, dealt with a report on a 
" Study of the effects of different proteins and ash constituents on 
the growth of pigs." 

Professor E. W. Morse, of the Office of Experiment Stations, 
U. S. Department of Agriculture, read a paper entitled " Sugges- 
tions concerning the planning and reporting of feeding trials," in 
which he pointed out some needed improvements in planning feed- 
ing tests so that the results obtained could be interpreted by modern 
biometrical and Statistical methods, and that the work as a whole 
could be so systematized and coördinated that the results of each 
investigation could be compared with those obtained by others. At 
present the work in this direction is so uncorrelated that a compila- 
tion of results on any uniform basis is out of the question. 

The Standing committee on methods of reporting results of 
feeding experiments made a special report, which contained recom- 
mendations urging uniform methods of reporting data obtained in 
such experiments. The recommendations include a summary of the 
opinions expressed by a large nuniber of investigators in response 
to questions previously propounded to members of the Society. The 
recommendations were adopted. 

It was voted to enlarge the scope of the work of this society, to 
include all animal husbandry interests — problems connected with 
the breeding, judging and management of live stock. This is to be 
in addition to investigations in regard to nutritive value of feeds 

282 



1913] Lewis W. Fetzer 283 

and other problems pertaining to animal nutrition. The name of 
the Society was accordingly changed to "The American Society 
of Animal Production." 

The officers elected for the coming year are as f ollows : Pres- 
ident, C. F. Ciirtiss (Iowa State Agricultural College) ; Vice Pres- 
ident^ E. B. Forhes (Ohio State Agricultural Experiment Station) ; 
Secretary-Treasurer, D. H. Otis (University of Wisconsin) ; 
CoMMiTTEE ON Experimentation, H. J. Wüters (Kansas State 
Agricultural College) and E. B. Forhes (Ohio State Agricultural 
Experiment Station). 

A meeting of the Society will be held at the time of the Panama- 
Pacific International Exposition, at San Francisco in 19 15. 

Office of Experiment Stations, 

U. S. Department of Agriculture. 



EIGHTH SCIENTIFIC MEETING OF THE COLUMBIA 

UNIVERSITY BIOCHEMICAL ASSOCIATION AT 

THE COLLEGE OF PHYSICIANS AND SUR- 

GEONS, NEW YORK, DEC. 6, 1912* 

Proceedings reported by THE Secretary, 
ALFRED P. LOTHROP 

The eighth scientific scssion of the Columbia University Bio- 
chemical Association was held at the Columbia Medical School, at 
4 p. m., on December 6, 1912.^ Abstracts of the papers are pre- 
sented here (pages 285-96) in two groups : (I) Abstracts of papers 
on research by non-resident members^ and (II) abstracts of papers 
from the Columbia Biochemical Department and affiliated labora- 
tories. The appended summary f acilitates ref erence to the abstracts 
(45-62).^ 

A SUMMARY OF THE NAMES OF THE AUTHORS AND OF THE 
TITLES OF THE SUCCEEDING ABSTRACTS 



Allan C. Eustis. Biochemical rea- 
sons why free purgation is necessary 
in combating acidosis of diabetes ; 
results of clinico-chemical observa- 
tions. (45) 

A. J. GoLDFARB. Studies on the effects 
of salinity changes upon regenera- 
tion. (46) 

I. Greenwald. The procedure of 
Salomon and Saxl as a diagnostic 
test for Carcinoma. (47) 



J. Arthur Harris and Ross Aiken 
GoRTNER. On the relationship be- 
tvreen the weight of the sugar beet 
and the composition of its juice. 
(48) 

Paul E. Howe (with H. C. Biddle), 
Fasting Studies. XL Note on the 
composition of the muscle of fast- 
ing dogs. (49) 

Max Morse. The role of phagocytes 
in the involuting tail of amphibian 
larvae. (50) 



* Scientific meetings are held regularly on the first Fridays of December, 
February and April, and on the first Monday in June. 

^ Proceedings of the sixth meeting were published in the last number of the 
Biochemical Bulletin, 1912, ii, p. 156. Proceedings of the seyenth meeting are 
published on page 322 of this issue. 

* Members of the Association who were not officially connected with the 
Columbia Biochemical Department when the research was conducted. 

'For abstracts 1-44 see Biochemical Bulletin, 1912, ii, p. 156. 

284 



I9I3] 



Alfred P. Lothrop 



285 



Max Morse. Laboratory hints. (51) 
Jacob Rosenbloom. The diflfusion o£ 

iodo-eosin from ether through rub- 

ber into ether. (52) 



Jacob Rosenbloom. The absence of 
certain enzymes from the human 
chorion, (53) 



II 



Walter H. Eddy. The preparation of 
histon from flounder sperm. (54) 

Thuisco A. Erpf-Lefkovics and 
Jacob Rosenbloom. A quantitative 
study of certain enzymes of the 
ovary, Uterus and bladder, of preg- 
nant and non-pregnant sheep. (55) 

Nellis B. Foster. Pathological devia- 
tions in the chemistry of uremic 
blood. (56) 

Nellis B. Foster. Effect of phlor- 
hizin on a dog with Eck fistula. 

(57) 
Frederic G. Goodridge and Nellis B. 
Foster. The relation of uricolysis 
to sub-oxidation. (58) 



William J. Gies. A demonstration 
of some of the tinctorial properties 
of pigments produced from thymol 
by ammonium hydroxid. (59) 

B. Horowitz. Experiments on pig- 
ments produced from thymol by the 
action of ammonia. (60) 

W. M. Kraus. Influence of uranium 
nephritis on the excretion of Crea- 
tinin, uric acid and chlorids, and the 
effect of Creatinin injections during 
uranium nephritis. (61) 

Charles Weisman. On the question 
of protein in expired air. (62) 



I. ABSTRACTS of PAPERS on RESEARCH BY 
NON-RESIDENT MEMBERS* 

45. Biochemical reasons why free purgation is necessary in 
combating acidosis of diabetes; results of clinico-chemical ob- 
servations. Allan C. Eustis. (Laboratory of Clinical Medicine, 
Medical Department, Tulane University, New Orleans, La.) Ob- 
servations were made upon twelve patients in the von Noorden 
clinic in Vienna and lipon four patients in the private practice of 
the writer during the past year. Tests for indican were made by 
Salkowski's method, the same amount of urine being used for each 
test, and the grade of the color imparted by the indigo was recorded 
+, ++, + + +J etc. The ammonia determinations were made 
according to Folin's method. In each case there was a marked 
degree of acetonuria, also a large percentage of ammonia nitrogen 
in the urine, and a high indican index. 

According to the experiments of Dale, when />-hydroxyphenyl 

* Members of the Association who were not officially connected with the 
Columbia Biochemical Department when the research was conducted. 



286 Proceedings Columbia Biochemical Association [Jan. 

ethylamin is perfused through the liver, it yields phenyl acetic acid. 
As the amin is a product of intestinal putrefaction of tyrosin, and 
assuming that some such cleavage takes place with other aromatic 
amins of intestinal putrefaction, their formation should be markedly 
hindered by free purgation, with consequent reduction of acidemia. 
In the cases observed there was a drop in the proportions of am- 
monia, acetone, and indican when free purgation was instituted in 
conjunction with a low protein diet. 

In two cases of fatal acidemia it was impossible to obtain free 
purgation owing to intestinal paresis, while in those cases in which 
free purgation was obtained, prompt relief from acidemia was noted. 

46. Studies on the eff ects of salinity changes upon regenera- 
tion. A. J. GoLDFARB. (Marine Biological Laboratory of the 
Carnegie Institution^ Dry Tortugas, Florida, and the Biological 
Laboratories of the College of the City of New York.) This in- 
vestigation was made with the idea of ascertaining to what extent 
changes in density of sea water afifected an organism, Cassiopea 
xamanacha, normally subject to relatively great changes in density 
of the sea. It was furthermore intended to compare the results, on 
one band, with those upon the hydroid, Endendriuin, which lives in 
more dilute and more static densities, and on the other, with the 
classic results of Loeb on the Tubidaria of Messina. Much care 
was devoted to reducing the number of variable factors, or in ren- 
dering them uniform, or eliminating them altogether, such as, varia- 
tions in respiration due to varying volume, surface and depth of 
the Solutions, variations due to differences in size of the medusas, to 
level of amputation, to cyclical variations in density, to limited num- 
bers (323 arms were used), etc. 

The results are not easily summarized. Normal and super- 
normal regeneration occurred in normal sea water and in sea water 
diluted from 5 to 15 per cent. In the gradient series, regeneration 
was at first gradually, then more rapidly, reduced until a 50 per cent. 
Solution was reached, in which regeneration was inhibited altogether. 
The medusae however lived in this and in the 45 per cent. solutions. 
The other half of the curve was strikingly different, in that re- 
generation feil off very rapidly, ceasing completely in 125 or 133 
per cent. solutions. The whole curve was strikingly different in 
character from the one described by Loeb. 



I9I3] 



Alfred P. Lothrop 



287 



It is particularly noteworthy that the curve for the Eudendrium 
is intermediate in character between Tuhnlaria and Cassiopea. It 
seems altogether certain that Loeb's curve can no longer serve as a 
type, expressive of the behavior of organisms under varying condi- 
tions of density of the sea water, and it is doubtful whether the 
phenomenon can be expressed in a simple curve based on two vari- 
able factors. It appears also probable that the relatively high con- 
centration in which Cassiopea normally lives may be associated with 
the high Optimum density for regeneration in this animal. It is also 
probable that the minimal effects of increasing dilution may be an 
adaptive response to the extreme dilutions to which Cassiopea is 
normally subject. 

47. The procedure of Salomon and Saxl as a diagnostic test 
for Carcinoma. I. Green wald. (Chemical Laborafory of the 
Monte fiore Home, New York City.) The procedure of Salomon 
and Saxl,^ proposed as a test for Carcinoma, was tried in a number 
of urines. All were positive. The precipitates obtained were 
filtered off, ignited, and weighed, with the results shown in the 
appended summary. Total sulfur Was also determined. There was 
no apparent relation between the amount of sulfur precipitated by 
the Salomon-Saxl procedure, either absolute or relative to the total 
sulfur, and the presence or absence of Carcinoma. 





Number of 
urines 


Number of 
cases 


Sulfur precipitated in the test : 


Types of cases 


As BaS04 
average, mg. 


Relation to total 
sulfur, per Cent. 


Normal 


8 

5 
9 

2 


6 

5 
8 

2 


9-5 

7.2 

6.6 
7-7 


I.2I 


Pathological 


I-5S 
1.23 


Carcinoma 


Sarcoma 





48. On the relationship between the weight of the sugar 
beet and the composition of its juice. J. Arthur Harris and 
Ross AiKEN Gortner, {Carnegie Institution of Washington, Sta- 
tion for Experimental Evolution, Cold Spring Harbor, L. /.) Al- 
though the literature pertaining to work on the sugar beet is very 
voluminous, but little attention has been paid to the relationships 

"Salomon and Saxl: Wiener klinische Wochenschrift, 191 1, xxiv, p. 449; 
Deutsche medizinische Wochenschrift, 1912, xxxviii, p. 53. 



288 Procccd'mgs Columbia Biochemical Association [Jan. 

that may exist between the weight of the root of the beet and the 
chemical composition of its juice. We have compiled such data 
from varions federal and State bulletins, and have examined them 
by calculating the intensity of such relationships on the — i to + i 
Scale of the coefficient of correlation. We have also written the 
regression equations showing the absolute change in solids, sugar, or 
purity, associated with a unit change in the weight of the beet.° 

We find that composition and purity are very closely correlated 
with the weight of the beet; as the weight increases, total solids, 
purity and sucrose fall rapidly. The following is a representative 
summary showing the rate of fall on the relative scale of — i to + i 
of the coefficient of correlation, and the rate on an absolute scale by 
the second term of the regression equation, where w= weight, 
j=rz sucrose, /) = purity and ;f = total solids. 

Data pertaining to 475 Beets'' 

rwt = — 0.497 — 0.023 ^ = 20.1 19 — 0.096 w 
rws = — 0.576 ± 0.021 s = 17.644 — o. 122 w 
rwp = — 0.474 ± 0.024 p = 88.516 — 0.273 w 

Inasmuch as our results show the necessity of taking into account 
the weight of the individual beets in all studies on composition, and 
because of the bearing of our data on the beet sugar industry, we 
shall publish them in füll in the Journal of Industrial and Engineer- 
ing Chemistry (1913, v, p. 192). 

49. Fasting studies. XI. Note on the composition of the 
muscle of fasting dogs. Paul E. Howe (with H. C. Biddle). 
(Laboratory of Physiological Chemistry, University of Illinois.) 
To be published in füll in the April number of the Biochemical 
Bulletin. 

50. The role of phagocytes in the involuting tail of amphib- 
ian larvae. Max Morse. (Boardman Laboratories, Trinity Col- 
lege, Hartford, Conn.) Barfurth, Metchnikoff, Mercier, and 
others have sought, in the phagocytes, the principal factor in the 
absorption of tissues. This has been held in question mainly by 
Looss, who believes that a chemical dissolution is at the basis of the 

'Harris: American Naturalist, 1910, xliv, p. 693. 

* Nevada Data. Wilson: Bull. 32, Nev. Agri. Exper. Sta., 1896. 



1913] 



Alfred P. Lothrop 



289 



process and that this has no primary relation to the activity of the 
phagocytes. 

It is conceivable that if phagocytosis is the principal factor, the 
blood-counts (differential and with the hemacytometer) would show 
both an increase in the total number of leucocytes and a difference 
between numbers of polymorphonuclear forms, in the bloods of a 
larva in which the process of metamorphosis has not begun and of 
a transforming individual. On account of the great difficulty in 
identifying the various forms of leucocytes in hemacytometer 
preparations, this method of counting was not adopted, but the 
counts were made upon smears, stained with Wright's stain. Three 
sets of individnals were used: (i) Larvse in which the appendages 
had not appeared «and hence no absorption of the tail had begun; 
(2) individuals in which the process of absorption had progressed 
to some extent and (3) those in which the absorption had been com- 
pleted for a number of months, i. e., adult frogs. Twenty-eight 
specimens were used and the percentage results are as f ollows : 





Polymorph 
9.8 

8.6 
18.3 


Basoph 


Eosinoph 

6.5 
7.0 
0.4 


T,arge M 


Small M 


Individuais absorbing the tail 

Non-absorbing 

Adults 


4.2 

4.7 
6.2 


36.1 
20.6 
13.2 


42.4 

S9.0 
61.2 







The polymorphonuclear type runs slightly more numerous in 
the individuals undergoing metamorphosis than in individuals before 
the process has begun, but they are found in much larger quantities 
in adults than in either of the other two groups. As Friedsohn and 
Neumann have shown, however, it is impossible to distinguish young 
polymorphonuclear leucocytes in the blood of amphibian larvse f rom 
young erythrocytes and other forms of leucocytes, since all of the 
corpuscles originate from cells similar in appearance in all cases. 
Hence, the number of large nucleated forms doubtless includes 
young polymorphonuclear leucocytes; and if this were so, it would 
be expected that the number of the large ones would be smaller in 
adults, which is the case as seen in the above column of values for 
large mononuclear leucocytes. For these reasons, it is doubtful if 
there is any decided difference in the number of polymorphonuclear 
leucocytes in any stage of frog development; and therefore it is 



290 Proceedings Columbia Biochemical Association [Jan. 

doubtful whether these bodies play an important röle in the absorp- 
tion of the tail of the tadpole. 

With reg-ard to the basophiles and eosinophiles, it will be seen 
that they occur in smaller numbers in individuals undergoing meta- 
morphosis than in young tadpoles and, so far as the basophiles are 
concerned, they are fewer in number in larvse than in adults. The 
reverse is the case with the eosinophiles, being found in small num- 
bers in the adult. The small mononuclear leucocytes occur in 
smaller numbers in the "absorbing" animals than in either of the 
other types. They are regarded by the investigators mentioned 
above as the young, indifferent, forms of the several types of 
leucocytes and probably, also, of the erythrocytes, but since they 
occur in larger numbers in the blood of the adult, this view is not 
borne out by the results of the present investigation. 

It may be concluded, then, from this set of data, that leucocytes 
do not play an important role in protein and other transfer in the 
involuting tadpole tail nor do they initiate the process. 

51. Laboratory hints. Max Morse. {Boardman Labora^ 
tories, Trinity College, Hartford, Conn.) 

An ULTRA-FILTER. Colloids may be filtered to advantage by 
coating the surface of an alundum filter-disc, placed in a Buchner 
funnel, with a thin collodion Solution and applying the funnel to a 
pump, or the house vacuum. The first drainage through the filter 
will be coarse, the finer suspensions Alling up the interstices of the 
filter disc; afterwards, fine particles filter. 

A CONVENIENT HOT-WATER BOTTLE HOLDER. Sclcct 3. FloreUCC 

flask with a neck whose sides are as nearly parallel as possible for a 
distance and wrap half a yard of quarter-inch twine around the neck, 
spirally ; then wrap electrician's tape or surgeon's tape around the 
whole, again spirally, thus keeping the twine from unwinding. The 
heat does not soften the rubber of the tape sufficiently to cause it to 
leave the twine. If in place of the twine, small round leather belt- 
ing be used, the apparatus is perfectly satisfactory for all time. 
The belting may be obtained in any sewing-machine shop or in a 
belting supply house at small cost. 

52. The diffUsion of iodo-eosin from ether through rubber 
into ether. Jacob Rosenbloom. (Laboratory of Biological 



1913] Alfred P. Lothrop 291 

Chemistry of the University of Pittsburgh.) Boas and P have 
shown that various cholesterol esters diffuse from ether Solution 
through rubber into ether; cholesterol-stearate, for example, with a 
molecular weight of 652.51, diffuses very readily. Recently I found 
that the acid form of iodo-eosin diffuses very quickly under similar 
conditions. This substance, with a molecular weight of 836, has 
the composition indicated by the appended structural formula : 

co<^ • >c< >0 

53. The absence of certain enzymes from the human 
chorion. Jacob Rosenbloom. (Laboratory of Biological Chem- 
istry of the University of Pittsburgh.) Published in füll in this 
issue of the Biochemical Bulletin, page 236. See also abstract 

55- 

IL ABSTRACTS OF PAPERS FROM THE COLUMBIA BIOCHEMICAL 
DEPARTMENT AND AFFILIATED LABORATORIES 

54. The preparation of histon from flounder sperm. Walter 
H. Eddy. Testes, obtained from cold storage flounders and pre- 
served in alcohol, were ground and extracted with 0.8 per cent. 
hydrochloric acid Solution. The histon was obtained from the acid 
extract after the manner of Kossei and Kutscher, and purified by 
washing with water, re-dissolving in 0.8 per cent. hydrochloric acid 
Solution and reprecipitating with ammonia, several times. Like 
ammonia-precipitated thymus histon, this histon was insoluble in 
water and the hydrochloric acid Solution, after dialysis to neutrality, 
gave all the characteristic histon tests. The "ammonia precipi- 
tate," washed with water, alcohol and ether, and dried to constant 
weight at 105° C, contained 18.08 per cent. of nitrogen (one 
determination only, because of lack of pure material). Like that 
of thymus histon, the hydrochloric acid extraCt of the histon from 
flounder testes yielded, when saturated with sodium chlorid, a pre- 
cipitate which was soluble in water. Such a Solution gave the char- 
acteristic ammonia and alkaloidal tests, but was not precipitated 

* Rosenbloom : Biochemical Bulletin, 1912, ii, p. 67 ; Boas and Rosen- 
bloom : Proc. Soc. Exp. Biol. and Med., 1911, viii, p. 132. 



292 Proceedings Columbia Biochcmical Association [Jan. 

by cold nitric acid Solution. Histon from fresh testes is in course 
of prqjaration for comparisons with the product from cold storage 
material. We lare also investigating the presence of histon in 
flounder roe. 

55. A quantitative study of certain enzymes of the övary, 
Uterus, and bladder, of pregnant and non-pregnant sheep. 
Thuisco A. Erpf-Lefkovics and Jacob Rosenbloom. Published 
in füll in this issue of the Biochemical Bulletin, page 233. See 
abstract 53. 

56. Pathological deviations in the chemistry of uremic 
blood.^ Nellis B. Foster. In general it was found that there is 
a considerable increase in the amount of non-protein nitrogen in the 
blood in severe cases of nephritis. However this is not an invariable 
rule, as very severe cases sometimes show an approximately normal 
figure; and a high non-protein nitrogen value has been noted in 
other diseases, such as valvulär heart cases and pneumonia. The 
results must be interpreted in the light of the whole clinical picture. 
When the total non-protein nitrogen is i gram or over, per liter, 
the prognosis is probably to be regarded as extremely grave. The 
percentage of non-protein nitrogen that occurs as urea is extremely 
variable, and seems to bear no constant relation to the total non- 
protein nitrogen. The data are so lacking in concordance that it 
must be left to further investigation, now in progress, to disclose in 
uremic blood chemical substances which either qualitatively or 
quantitatively present a constant divergence from normal. 

57. Effect of phlorhizin on a dog with Eck fistula. Nellis 
B, Foster. It has been stated by Rosenfeld that the administration 
of phlorhizin to dogs with Eck fistula does not induce glucosuria. 
Such a result would have so much bearing upon our ideas of the 
mode of action of phlorhizin that the matter required further study. 
One gram of phlorhizin in olive oil emulsion was given to a dog with 
an Eck fistula. Glucose was found in the urine, in considerable 
quantity, for nine days subsequent to the phlorhizin administration. 

58. The relation of uricolysis to suboxidation.^*' Frederic 
G. Goodridge and Nellis B. Foster. In order to investigate the 

'Foster: Archives of Internal Mediane, 1912, x, p. 414. 
^* Goodridge and Foster : Ibid., 1912, x, p. 585. 



I9I3] Alfred P. Lothrop 293 

subject of diminished oxidation in its relation to uric acid excretion, 
we &tudied cases of poisoning by illuminating gas (people) and by 
potassium Cyanide (dogs). The results show that retardations of 
the oxidizing processes, either by deprivation of oxygen (gas) or 
by interference with cellular functions ( Cyanide), were not followed 
by increased excretion of uric acid. It appears improbable, there- 
fiore, that uric acid destruction in the body is a simple oxidizing 
process. 

59. A demonstration of some of the tinctorial properties of 
pigments produced from thymol by ammonium hydroxid. 
William J. Gies. The phenomena described in a previous com- 
munication on this subject^^ were demonstrated, as an introduction 
to the succeeding communication by Mr. Horowitz. Ammonium 
hydroxid produces from thymol a blue^ pigment (or mixture of 
pigments?). Ether extracts the blue material from the alkalin 
liquid, but in ether Solution the pigment is red. After evaporation 
of the ether from such an extraot, a purplish oily product remains. 
This residue yields a purplish Solution in alcohol, which becomes 
hliie when it is rendered slightly alkalin. Filter paper, soaked in 
such a blue, alkalin, alcoholic Solution, and then dried at room tem- 
perature, assumes a bright red color as the alcohol disappears. 
Treated with alcohol, such red filter paper, particularly if slightly 
moist, becomes bright green. Interesting probabilities suggested by 
these results, and the possible relationship of these color phenomena 
to the pigments in the Monardas^^ and other plants, will be in- 
vestigated. 

60. Experiments on pigments produced from thymol by the 
action of ammonia. Benjamin Horowitz. Professor Gies has 
found that thymol, in contact with ammonia, develops a blue color.^^ 
Under his direotion I have been making a study of this phe- 
nomenon. 

The question early arose as to whether ammonia and thymol 
alone are sufficient for the formation of the pigment. Certain ob- 

"Gies: Biochemical Bulletin, 1912, ii, p. 171. 

"Wakeman: Bulletin of the University of Wisconsin, No. 448; Science 
series, 191 1, iv, p. 81. 

"Gies: Biochemical Bulletin, 1912, ii, p. 171, See also the preceding 
abstract, above. 



294 Proceedings Columbia Biochemical Association [Jan. 

servations by Liebermann^"* (whose work on colored Compounds 
produced from phenols has been of great importa'nce ) , as well as 
the many uses o£ oxidizing agents in the production of phenol pig- 
ments, suggested that 'atmospheric oxygen takes an active part in the 
process. This supposition was confirmed. A current of hydrogen 
passed throiigh an aqneous ammonia-thymol mixture inhibited the 
f ormation of pigment. Nascent hydrogen ( f ormed by the addition of 
Zn dust or sodium amalgam) acted similarly. On the other band, 
the addition of a few drops of hydrogen peroxid greatly accelerated 
the production of pigment by oxidation. A rise has been noticed 
in an aqueous ammonia-thymol mixture inverted over water, in- 
d'icating the absorption of oxygen. 

The addition of Zn dust to the blue Solution in an open vessel 
caused the color to disappear, except near the surface, where blue 
always remained, showing the influence of atmospheric oxygen. 
Ether was added to ascertain whether the reduced substance could 
be extracted by it and whether the chromogen thus removed by ether 
would yield pigment in the presence of oxygen. The bottle was 
now tightly stoppered and allowed to stand. Within 24 hours the 
red ether layer (the blue pigment yields a red Solution in ether) had 
become colorless. On releasing the stopper the ether Solution be~ 
came colored again. This was repeated many times with different 
samples but always with the same result. Attempts have since been 
made to isolate the reduced product by evaporating the ether Solu- 
tion in a current of hydrogen, but so f ar without success. The work 
is in progress. 

61. Influence of uranium nephritis on the excretion of 
Creatinin, uric acid and chlorids, and the effect of Creatinin in- 
jections during uranium nephritis. W. M. Kraus. In acute 
uranium nephritis in dogs, Creatinin was excreted in decreased 
amounts ; uric acid, in increased amounts. In subacute uranium 
nephritis, Creatinin was eliminated in decreased amounts ; uric acid 
and dhlorids, in increased amounts (2 weeks). Creatinin, injected 
in normal dogs, appears to be excreted " in toto." Creatinin, in- 
jected during acute uranium nephritis, is not wholly eliminated. 
Such an injection causes decreased Output of endogenous Creatinin, 

"Liebermann: Ber. d. d. Chem. Gesell., 1874, vii, p. 247; 1875, viii, p. 1649. 



I9I3] Alfred P. Lothrop 295 

also uric acid, chlorids and wäter ; and death may ensue. Creatinin, 
injected during subacute uranium nephritis, is excrelted "in toto" 
and apparently does not affect the excretion of endogenous cre- 
aitinin, uric acid, chlorids or water. 

Two of the animals with acute nephritis died after injection of 
Creatinin, in marked contrast with the apparent lack of effect of 
Creatinin injections in th^ dogs with subacute nephritis. In acute 
nephritis, where Creatinin (endogenous and injec'ted) and all the 
other substances named above were excreted in decreased amounts, 
there was apparently a partial arrest of renal function. In the 
subacute nephritis this did not occur. In acute nephritis there is 
probably not only insufficient intact tubulär epithdium to carry the 
additional bürden, viz., the injected Creatinin, buit this additional 
bürden aggravates the preexisting condition. In subacute nephritis, 
on the other hand, regeneration occurs, thus increasing the func- 
tional possibilities of the kidney, so that injected Creatinin can be 
excreted " in toto." 

Fever increases Creatinin excretion. It is also known that cer- 
tain infeotions, e. g., diphtheria, cholera, pneumonia and colon in- 
fections, cause tubulär nephritides. If, instead of adding hyper- 
creatininemia to a nephritis which is predominatingly tubulär (as 
described for the above experiments with uranium nephritis), there 
should be hypercreatininemia follozved by tubulär nephritis, it is 
probable that a similar reaction would result, namely, a uremia-like 
toxemia, ending perhaps in death. 

Creatinin has been taken only as an example of urinary sub- 
stances normally excreted by the kidney tubules. It seems probable 
that other normal substances, which are increased in fever and ex- 
creted by the tubules, would act in a similar way, i. e., to overtax an 
already overfunctioning kidney in a condition aggravated by a 
tubulär nephritis. This Suggestion as to the production of uremia 
dioes not concern the substances directly causing rt, merely the 
mechanism of their retention. 

62. On the question of protein in expired air. Charles 
Weisman. Rosenau and Amoss^^ recently pu'blished a paper in 
which they stated that expired air contains volatile prcytein. Their 

"Rosenau and Amoss: Journal of Medical Research, 191 1, xxv, p. 35. 



296 Procccdings Columbia Biochemical Association [Jan. 

conclusions were dependent 011 anaphylactic phenomena that ap- 
peared to be obtained with condensations from expired air. At Dr. 
Gies' Suggestion I am repeating their experiments with a view of 
applying the findings to problems in Ventilation and disease. Six 
repetitions of the experiments by Rosenau and Amoss have been 
made thus far, with negative results in each instance. 

We believe that Rosenau and Amoss failed to conduct adequate 
control experiments, both on the toxic effects of blood serum and 
on their anaphylactic tests. Their choice of sites (heart, brain) 
for the injeotions is open to criticism. Injeotions into the heart 
may produce lesions of the heart or lungs as well as pericardial in- 
jury and hemorrhage; and, after injury to the bündle of Hiss, 
resu'ltant Symptoms like the Stokes-Adams Syndrome, with oon- 
sequent difficulty of respiration, may simulate anaphylactic effects. 
Besides, in such injections, there is no assurance that the entire 
amount of injected fluid goes into the heart. As for injections into 
the brain, Rosenau and Amoss themselves say : " When the second 
injection was placed under the dura, through the optic foramen, the 
results were sometimes clouded by the appearance of Symptoms 
which were interpreted to be the result of central Irritation." In 
cur work, the second injection was made intravenously (externa! 
jugular vein) . 

The conclusion by Rosenau and Amoss, that expired air contains 
protein ( " volatile " ) , appears to be unwarranted. The experiments 
are in progress with the Cooperation of Drs. J. Bronfenbrenner and 
S. Gitlow. 

[The proceedings of the February and April meetings of the 
Biochemical Association will be published in the April number of 
the Biochemical Bulletin.] 

Biochemical Laboratory of Columbia University, 
College of Physicians and Surgeons, 
New York. 



FOLIA MICROBIOLOGICA 

Perhaps it will be of some interest for readers of the Biochem- 
ICAL Bulletin to learn that the recently founded Nederlandsche 
Vereeniging voor Microbiologie publishes a quarterly Journal enti- 
tled Folia Microbiologica. The editors are : Professors Beijerinck 
(Delft), Klein (Groningen), Poels (Rotterdam), and Sleeswijk 
(Delft). The subscription price of Folia Microbiologica is Five 
Dollars ($5.00) a year. The first volume is now complete and con- 
tains the following papers : 

Numbers i and 2 : Beijerinck, Mutation bei Mikroben ; Klein, 
Ueber die biologische Analyse des Kaseinantiserums ; Jacobsen, Die 
Kulturbedingungen von Hcematococciis pluvialis. 

Number 3 : Söhngen, Ueber fettspaltende Mikroben und deren 
Einfluss auf Mölkereiprodukte und Margarine; Ross Van Lennep, 
L'influence des substances fixes sur l'anaerobiose dans les milieux 
de culture liquides ; Fresemann Victor, Ueber die proteolytische und 
antiproteolytische, resp. antitryptische, Wirkung des menschlichen 
Blutserums; Reeser, Complement fixation of different sera prepared 
at the State Serum Institute, Rotterdam ; Boeseken und Waterman, 
Ueber die Wirkung der Borsäure und einiger anderen Verbindungen 
auf die Entwickelung von Penicillium glaucum und Aspergillus 
niger. 

Number 4: Eijkman, Untersuchungen über die Reaktionsge- 
schwindigkeit der Mikroorganismen; Beijerinck, Die durch Bakte- 
rien aus Rohrzucker erzeugten schleimigen Wandstoffe; VanCalcar, 
Ueber die Kenntnis des anaphylaktischen Zustandes des tierischen 
und menschlichen Organismus; Waterman, Beitrag zur Kenntnis 
der Kohlenstoffnahrung von Aspergillus niger; Jacobsen, Die Oxy- 
dation von elementarem Schwefel durch Bakterien. 

The editorial introductory notice concludes with these words: 
" If foreign authors should wish the fruits of their researches to 
appear in our columns, they would be most heartily welcomed. 
The Latin title of our Journal indicates that this new publication 
has not sprung from chauvinism. We entertain very earnest con- 
victions on the ' internationalism of science.'" 

C. A. Pekelharing 

University of Utrecht, 
Holland. 



BIOCHEMICAL BIBLIOGRAPHY AND INDEX 

WILLIAM J. Gl ES 

(Biochemical Lahoratory of Columbia University, at the College of Physicians 

and Surgeons, New York.) 

Biochemical literature is becoming so abundant and comprehen- 
sive, and so detailed and miscellaneous, that earnest efforts to assimi- 
late a considerable part of it are apt to induce severe attacks of indi- 
gestion. Ways and means for rapid and accurate sifting and 
Classification of subjects, experimental data, conclusions, and theo- 
ries, gain importance witli the increase of activity, and the growth 
of interest, in biochemistry. Appropriate year books, reviews, Zen- 
tralblätter, and especially the Department of Biological Chemistry 
in Chemical Ahstracts, acquire cumnlative usefulness as biochemical 
research develops in quantity, variety and extent. 

The writer finds it very desirable to maintain a running, quar- 
terly, card index of the titles of the papers in the leading biochemical 
Journals. This index is particularly valuable during the intervals 
between the publication of indices of volumes of abstracts and re- 
views. In the belief that it may be helpful to others, a copy of a 
portion of the index is presented below. Sections of the bibliog- 
raphy and index will be printed regularly in the Biochemical Bul- 
letin, if the writer's opinion on its probable general Utility proves 
to be correct. 

In the appended bibliography, titles are shortened in a free and 
easy manner, redundant words are ignored, common words are ab- 
breviated, surnames of collaborators are connected by hyphens, and 
punctuation marks are omitted, for the sake of condensation. Vol- 
ume numerals are given in Roman, and the Arabic numerals imme- 
diately following them, or placed at the beginning of sections, desig- 
nate respective issues of the volume ; numerals separated by a slanted 
line indicate month and day of issue. The numeral at the end of 
each item is that of the initial page of the corresponding paper. 
The numeral at the beginning of each item indicates sequence in the 

298 



1913] William J. Gies 299 

bibliography. The latter numerals are used in the index of subjects 
at the end (page 305). Abbreviations of words in the index are 
similar to those in the bibliography. The System of notation in the 
index, although a space-saving device, makes it easy to refer to any 
title in the bibliography, The index is a compass not an encyclo- 
pedia; its purpose is achieved if it acts as a convenient guide to the 
sources of information — if it helps the reader speedily to locate and 
follow the main currents through the literature. 

Biochemical bibliography and index: 1912; fourth quarter (Sept.- 
Dec.) ; Journals : Biochemische Zeitschrift, Zeitschrift für physiolo- 
gische Chemie, Journal of Biological Chemistry, Bio-Chemical Journal, 
Biochemical Bulletin. 

Biochem. Zeit. XLIV: 1-2; 9/6. — iHäri Einfl intraven Bluttransf a 
Gaswechs,!. — 2Häri-Pesthy Hat Temp d Nähr Einfl a Gaswechs ?,6. — 
^Rudö-Cserna Wirk intraperiton Blutinfus a Gaswechs,40. — 4Häri Einfl 
Kohlenhyd a Energieums,66 ; 5Wirk intraperiton Blutinfus a Energie- 
verbr,84. — GAlexand^r-Revess Einfl optisch Reiz a Gaswechs Ge- 
hirns,95. — ^Alexander Blutgaswechs Gehirns, 127. — SOrnstein Parenter 
Ernähr,i40. 3-4;9/9. — gQuagliariello Änd H'-konz währ Hitzekoag 
Prot,i57; loH'-konz Blut bei Temp'erhöh nach Wärmestich,i62. — 
iiRohonyi Veränd H'-konz b Pepsinwirk u Saurebind'vermög hydrol 
Spalt'prod Eiweis,i65. — i2Glaser Entwick'arbeit im Fundulusei,i8o. — 
iT^Berczeller Lipolyt Wirk Organextr,i85 ; i4Bestim Fet u Lipoid d 
Blutes u üb Lipol,i93. — i^Verzär Arb Pankr u Einfl a Verbren Kohl- 
enhyd,20i. — i6Beläk Wirk Phloriz a Gaswechs u Nierenarb,2i3. — 
lyTangl Respir'app für Schwein, Schaf ,235 ; iSMinim Erhalt'arbeit 
Schwein i Hunger,252. — igWeiser Ca, Mg, P, N-Umsatz wachs 
Schwein,279. — 2oZunt2 Versuchsergebn v Chauveau Mind'wert d 
Fet Kohl'hyd gegenüb als Energiespend b Musk'arb,290. — 2iSiegfried- 
Zimmermann Bericht, 292. 5-6;9/2o. — 22Pribram Diastas,293. — 
2^Loeb-Beutner Verletz'strom,303. — 2^alladin Eiweissabbau u At- 
mung Pflanz,3i8. — 2^Forssmann-Hintze Heterol Toxiz Antiser,336. — 
26Kopacze'wski Einfl Antisept a Wirk Maltase,349. — 2yTschernoruski 
Wirk V Nucleinsäu u nucl'spalt Ferm im tier Organis,353. — 28Fasal 
Colorim Meth quant Tryptophanbest u üb Tryptophangeh Horngebild u 
Eiweisskör,392. — 2gBierry Verdau Inulin, 402; 3oSaccharos spaltend 
Ferm,4i5; 3iRafiinos u Gentianos spalt Ferm,426; 32Stachyos u Man- 
ninotrios spalt Ferm,446. — ;^^Ohta Hitzebeständ Tryps u Peps,472; 
34Verh Äpfelsäu im Tierkör,48i. — ^^N euherg-Schewket Polarim 
Bestim Glucosamingeh v Ovomucoid u Pseudomucin,49i ; 36Veränd 



300 Biochemical Bibliography and Index [Jan. 

Arzneimit i Licht,495 ; 37Nachw gepaar Glucuronsäu i Harn, 502. — 
2ßAltschul " Agfa "-Lecith, 505. (Pages, 508.) 

XLV:i-2;9/25. — ^gEmbden-Krans Milchsä'bild durchström Leber, 
I. — 4oSchmits Verh Glycerin künst Durchbl Leber, 18. — 41 Oppen- 
heimer Milchsä'bild durchström Leber,30. — 42Embden-Kalberlah-Engel 
Milchsä'bild Muskelpresss,45. — 4^Kondo Milchs'bild Muskelpresss,63. 
—44Kraske Milchs'bild B\ut,8i. —4SKondo Milchs'bild Blut,88.— 46^; 
Noorden Milchs'bild Blut,94. — 4yEmbden-Baldes-Schmits Milchs'bild a 
Traubenz i Tierkör, 108. — 480ppenheitner Einwirk verdün NaOH a 
Glyc'ald u Dioxyaceton,i34. — 4gMasiida Auftret aldehydart Subst b 
Leberdurchblut u üb Acetessigsäu'bild aus Äthylalk, 140. — ^oEmbden- 
Baldes Umwandl Acetaldeh i Äthylalk tier Organism, 157. 3-4;9/30. 
— ^lOhta Acetessigsä'bild aus Dicarbonsä mit 4C-Atom,i67. — ^2Emb- 
den-Schmita-Baldes Glycer'bild Tierkör, 174. — ^T^Embden-Oppenheimer 
Abbau Brenztraub'säu i Tierkör, 186. — $4Höber Verteil Blutzuck auf 
Körperch u Plasm,207. — ^^Langer Heroinaussch u -gewöhn,22i ; 56 
Alkaloidaussch nach Magen unt Einfl i Mag gebracht Salz,239. — 
$yPescheck Einwirk NH4-salz u essig Salz a N-wechs Fleischfr,244. — 
^SCytronberg Cholest'ase Blutkör,28i. — sgDavidsohn Magenlipase,284. 
— 6oEisenberg Formaldehydhämoly,303. — GiMaidorti Blutgiftanäm, 
328. — 62Allemann Bedeut H' Milchgerin,346. 5-6;io/i2. — 6^Endler 
Durchtr Salz durch Protoplas,359. — 64N othmannZuckerhandl Wirk 
Narkot a Plasmaström,4i2. — 6$Tschermak Adaptat Ferm'bild Ver- 
dau'kan,4S2. — 66Bickel-Tsividis Einfl Digital'kör Kurv d Elekt'kar- 
diogr,462. — GyFreudenberg Fettstoffwechs,467. — 6SPreti Katal Einwirk 
Blei a Harnsä'bild u Harnsä'zersetz, 488. (Pages, 502.) 

XLVI:i-2;io/23. — 6gPorges Resp Quot Säurevergift, i. — yoBre- 
dig-Fiske Katal bewirk asym Synth,7. — yiChristiansen Frei u geb HCl 
i Mag'inh,24; y2lbid., 50; yT,Ibid., 71; y4lbid.,S2. — y^Müller Einfl Be- 
handl Milch a Labfähigk,94. — y6Würts Verteil P-säure Harn u Kot, 
103. — yyTschernorutzky Wirk NaaCOg auf Alkaloidsalz u Farbst,ii2. 
— ySLöb Verbal Stärk unt Einfl stillen Entlad,i2i ; 79Pankr'diast,i25. 
— SoMichaelis-Davidoff Elektrom Bestim Blutalk,i3i. — SiKammann 
Pollentox,i5i. — 82Rohland Adsorp Tone,i70. 3-4;ii/9. — S^Galeotti 
Aussch H2O bei Atmung, 173. — 84Gramenitzki Blut- u Hamzuck b 
Adren'inf,i86. — ^^Siegfried-Zimmermann Entst Phenol a />-Kresol i 
Hund,2io. — 86Rosenthal Einfl Osmiumsäu a Receptor'app Erythrocy, 
225. — 8yOhta Bedeut Proteol f spezif Hämol, 247. — 88 Fränkel Lipoid, 
253. — 8gChristiansen Mett Meth u Acid'opt Peps'wirk,257, — goLöb- 
Gutmann Glykoly,288. — giHaas Schick Glyoxylsäu i Tierkör,296. — 



1913] William J. Gies 301 

g2Rona Schicks tierabgeb Eiweisskör i Darm,307. 5;ii/i4. — g^Bat- 
telli-Stern Oxyd />-Phenylendiam i Tiergevveb,3i7; g4lbid.jT,4T,. — 
g^Mohr-Heimann Norm u Eklamp Placent,367. — g6Rohland Ad- 
sorp Tone,374. — gySchuls Kieselsä'gehalt Schilddr,376. — gSvFenyvessy 
Natur u künst Komplem verhalt sich in " Regenerat " ident,393. — 
ggBattelli-Stern Nomenkl Polyphenoloxydas,395. 6;ii/25. — lOoDox- 
Neidig Spalt a- u ^-Methylglucosid dur Asp niger,2g7. — lOiHassel- 
balch Neutral'reg u Reizbark Atemzent i Wirk a COa-span Blut,403. — 
i02Winterstein App Mikroblutgasanal u Mikrorespirom,440. — lO^Pig- 
hini Chem Nerv'syst,450. — lO^Freund-Kaminer Bezieh zw Tumorzel u 
Blutser,47o. — lo^Lehedew Mechan alkohol Gär,483. — io6L6pezSuärez 
HCI-bild Magen,490. — loyBang Erwid,5oo. — loSEissler Physostigmin, 
502. (Pages, 504.) 

XLVII:i ;ii/30. — logPalitzsch-Walbum H"-konz b trypt Gelat'ver- 

flüs.i. — I loKlausner Klausner Serumreak,36. — 1 1 iMichaelis-David- 

sohn Abhäng spezif Fäll'reak von H*konz,59. — ii2Beutner Physikal 

Nat bioelek Potent'dif, y^. 2;i2/g. — ii^Johannessohn Einfl org Sau 

a Hefegär,97. — ii4Loeb Verh Eissigsäu b künst Durchblut Leber,ii8; 

ii5Permeab u antag Elek'lytwirk (neu Meth),i27. — iißHirata Diast 

Kraft Mundspeich,i67. — iiyZaleski-Marx Carboxylas höh Pflanz, 184. 

— iiSStoklasa-Sebor-Zdobnicky Photochem Synth Kohlenhydr,i86. 

3-4;i2/i2. — iigBerrär Chem u quant Bestim Leim, 189. — i2oScaffidi 

Purinstoffwechs,2i5. — i2iChristiansen Mechan Peps'verdau,226. — 122 

Michaelis Isoelekt Punkt,250. — i22,Michaelis-P eckst ein Ibid., Casein, 

260. — 124W olfgang-Faleki: Physikal Zustandsänd Kolloid,269. — 125 

Starkenstein Ferm'wirk u Beeinfl d Neut'salz,3oo. — i26Grafe-Vouk 

Inulinstoffwechs Cichorium intyb L. (Zichorie), 320. — 127a/ Klercker 

Pentos d Guanylsäu,33i. — i28Loezvy-Gerhartz Aussch HoO b Atmung, 

343. 5;i2/i8. — i2gBickel-Pawlo'W Pharm Wirk /J-Oxyphen'äth'am, 

345. — i^oReale C-wechs; Labil u stab C d Harn,355. — i^iSsobolew 

Milchs'bild b antisep Organautol,367. — 12,2V ernon Abhäng Oxydase- 

wirk V Lip'd,374. — i^^Pollini Katal Wirk Eisensalz Leberautol,396. — 

i^4Neuberg-Kerb Zuckerfrei Hefegär,405 ; 135 Ibid,4i^. 6;i2/3i. — 

iTjSMoldovan Wirk Chinins,42i. — i^yMichaelis-Rona Umlag Glucos b 

alkal Reakt; Theor Katal,447. — i^SSasaki Abbau Polypept dur nicht 

verflüss Bakt,462; I39lbid dur verflüss Bakt,472. — i4oSignorelli Ver- 

hältn zwisch Amin-N u Ges-N Harn unt versch Beding,482. (Pages, 

508.) 

Zeit. f. physiol. Chem. LXXX:5;8/28. — i4iOsborne-Mendel- 
Ferry Wachst bei Fütter'vers m isol Nahr'subst,307. — i42Rogosinski 



302 Biochcmical Bibliography and Index [Jan. 

Methylier Clupein,37i. — i^T^Pringsheim Ferm Abbau Hemicellulos ; 
Trisacch Zwisch'prod d Hydrol Mannan,376. 6;io/2. — i^Wacker 
Cholesterin u Begleitsubst i Depotfet b Carcin,383. — i^^Trier Um 
Am'äth'alk (Colamin) i Cholin,409. — i46Letsche Einwirk Hydrox'am 
Blut färb (Methäm),4i2. — I4y Mörner Ovomucoid u Zucker Weiss d 
Vogelei,430. (Pages, 473). 

LXXXI: i-2;io/io. — i4SAbderJialdcn-Fodor Aus Glykok, d-Alan 
u /-Leuc darst struk'isom Tripept,i. — 14(^80 hui ze-Trier Verbreit Cholin, 
53. — i^oEider-Palm Zusam u Bild Enzym; Entwick Hefe in Nahrlös, 
59. — i^ilnonye Entst Kreatin, 71; i52Xan'prot'reak,8o. — is^Grabow- 
ski-Marchlewski Blutfarbst, 86. — 154 Abderhalden Feststel Schwan- 
gersch,90. — i^^Dorner Beeinfl alkoh Gärung d Zell u Zellpresss,99. — 
i^6Buglia-Costantino Muskelch; Formol titr Ges'am'-N d glat, querges 
u Herzm, 109; I57lbid, d N Ext'kt'st,i20; I581bid, frei, Formol titr 
Amin-N,i30; i59Chem Embry; Formol titr Ges'-am'-N d embry Mus- 
kel, 143 ; 160 Ibid, frei Am'-N Musk d Ochsenembr,i55. — i6iCostan- 
tino Muskelch; S d glat, querges u Herzm, sow Myoprot d Säuget,i63. 
— i62Usui Bind Thymol rot Blutz,i75. — i6^Sieber H2O2 als hydrol 
Prinzip, 185. — i64Rohland Tongeruch,200. — i6^Warbiirg Bestim kl i 
HoO-gelös C02-Meng,202. — 166J olles Nachw Glukuronsä diabet 
Harn, 203; i67Nachw Album Harn, 205. — i68Grafe Bericht,2o6. 
3;io/22. — i6gAbderhalden-Weil Nerv'syst Aminosä; Am'sä periph 
Nerv u Leitungsb Rück'mark (weis Subst),207. — lyoAbderhalden- 
Weil Die b Isolier Monoam'sä m Hilf Est'meth entsteh Verlust ; infrei- 
heitsetz Est m Bleihydr,226. — lyiAbderhalden-Hanslian Verh a-Pyr- 
rolid'carb'sä im Organis,228. — iy2Abderhalden-Weil Dreh'gsvermög 
Blutpias u -ser versch Tierart versch Alt u Geschl,233. — lyT^Laquer- 
Brünecke Einfl Gasen (O) Tryp- u Pep'verdau,239. — iy4Siegfried- 
Schutt Absch Am'sä mit Hilf Carbaminoreak,26o. — ly^Kossel-Gazv- 
rilow Frei Amidogr d Prot,274. — iy6Grafe Antw Bemerk v W. Völtz 
üb N-reten u N-gleichgew b Füt NH4-salz,28o. — lyy London Aus An- 
lass Mit'g Folin-Lyman, Prot metab standp bl'd a tis anal; absorp f 
stom,283. 4; 10/30. — lySAbderhalden-Kashizuado Kern Thymus u 
Anaphylax'vers mit Kernsubst (Nuc'prot, Nuc'n, Nuc'nsä),285. — 179 
Abderhaldeti-Kautssch Fäulnisvers rf-Glutam'sä; y^Am'but'sä,294. — 
— iSoAbderhalden Isol Glycyl-phen'alan a Chym d Dünnd; Stud mit 
Hilf versch Abbaustuf Prot u synth darges Polypep,3i5. — iSiAbder- 
halden-Hirsch Füt'vers Gelatin, NH^-salz, vollst abgebau Fleisch u 
aus all bekan Am'sä besteh Gemisch,323. — iS2Pfeiffer-Modelski Verh 
Am'sä u Polypep geg Neut'salz,329. — iSsPekelharing Anorg Salze auf 



1913] William J. Gies 3^3 

Wirk Pankreaslip,355. 5-6 ;i 1/5. — i^^London-Ritvosch-Mepissow- 
Stassow-Dagaew- Masijezvski- Gabrilowitsch- Krym- H olmberg - Wide- 
mann-Gillels-Solowjew Norm u path Verdau (Hund), 369. — iSsHen- 
riques-Gjaldhcsk Plasteinbildung,439. — i86Abderhalden-ValettePetti- 
bone Einfl physik Zust Prot a Raschh ihr Abbau dur Ferm; Bedeut 
Verdau Prot dur Pep-HCl für weit Abbau dur Tryp,458. — iSyAbder- 
halden-Lampe Schicks von in Magendarmk eingeführ Am'sä, Am'sä'- 
gemis, Pepton u Protein,473. (Pages, 507.) 

LXXXII:i-2;ii/i2. — i88Abderhalden-Hirsch Synth Fähigk tier 
Zell; Verwert versch N-quel im Organis,i. — iSgAbderhalden-Lampe 
Ibid,2i. — igoFischer-MeyerBets Porphyrinbildung,96. — igiAbderhal- 
den AnaphylaXjioQ. — ig2Küster Methylier Hämin,ii3. — ig^Abderhal- 
den-Wurm Pyrrol'carb'sä u aus ihr Aufgeb Polypep,i6o; I94a-Am'but'- 
rsä u Derivjiö/. — ig^Arnold Hämatoporphyr'urie b Abdom'typhus,i72. 
3-4 ',11/25. — ig6Hedin Reak zw Enzym u and Subst,i75. — igyJahn- 
sonBlohm Einwirk kol'd Subst auf Hem d Enzymwirk, 178. — igSRinger- 
Schmutzer Quad'ur,209. — iggKylin Färbst d Fucoid,22i. — 20oJegoroff 
Verh Schim'p (A niger, P crustac) z Phytin,23i. — 20iWiener Zusam's 
art u ven Blut,243. — 202Fischer-Krollpfeiffer Einwirk Phthalsä'anhyd 
a Pyrrolderiv,266. — 20^Arnold Darst Hämatoporphyr a CO-Blut,273. — 
204Fanser Einwirk HCl-gas Diastas,276. — 20^Jansen Konstit Cholsä 
verm Bromier,326. — 2o6Jansen Cholsä'resor Hundedarm,342. — 207 
Bürker Nomenkl Blutfarbs'deriv,346. 5;ii/3o. — 2oSGrafe N-reten 
b Füt NH^-salz (Schwein), 347. — 2ogPanser Einwirk HCl Invertas, 
:^'//'. — 2ioFischer-Röse Garfarbst,39i. — 2iiMariconda Funkt Verh 
Darmsegm nach lang Period funkt Untätigk,4o6. — 2i2Marchlewski 
Bemerk, Blutfarbst,4i3. — 2iT,Sieber Bericht,4i4. 6;i2/24. — 214L0M- 
don-Dobroivolskaja Pfortaderbl ; fistel,4i5. — 2i^Starkenstein-H enze 
Nachw Glykog Meeresmol'k (spez b Cephal u Aplys),4i7. — 2i6Kasch- 
izvabara Einfl J Autol,425. — 2ijBuglia-Costantino Muskelch; Ext'kt'st 
u frei Formol titr Am'-N d Musk,439. — 2i8Küster Bilirub u Hämin, 
463. — 2igRinger-Trigt Einfl Reakt auf Ptyarwirk,484. — 22oPincus- 
sohn Abw; Erwid an Gräfe, 502. (Pages, 502.) 

Jour. Biol. Chem. XII:3;9. — 221 Koch Meth guanid urin p'thy'dect 
anim,3i3. — 222Williams Anim calorim ; sm resp cal,3i7. — 223 Williams- 
Riche-Lusk Ibid., met dog aft ing meat larg quant,349. — 22äfLevene- 
Jacobs Guan'hexosid i hydrol thym-nucl ac,377; 225Cerebron ac,38i ; 
226Cerebrosid of brain,389. — 22yVanSlyke-Meyer Am'ac-N bl'd; prot 
assim,399. — 228Levene-Jacobs Struc thym-nucl ac,4ii; 229Guanyl 
ac,42i. — 22,oJacobs Prep glucosid, 427; 23iRemov phos'tung ac fr aq 



304 Biochemical Bibliography and Index [Jan. 

501,429. — 232 Ringer Prot metab i exp diabet,43i. — 2T^;^Anderson Org 
P- ac compd wh bran,447. — 2^4Kendall-Farmer Bact metab,465 ; 235 
Ibid,46g. — 2T,60sborne-Mendel-Ferry Gliad i nutr,473. — 22i7Ringer 
Chem gluconeogen ; quant conv propion ac i glucos,5ii. (Pages, 522.) 
XIII:i;io. — 2^SKober-Siigiura Cu compl'x am ac, peptd a peptn,i. 
— 229UnderhiIl Mech phlorh diabet,i5. — 24oLusk Anim calorim; metab 
aft inges dextr a fat, incl behav H2O, urea, NaCl,27. — 24iFisher- 
Wishart Ibid, absorp dextr a eff on comps brd,49. — 242Kendall- 
Farmer Bact metab,63. — 24^Mendel-Daniels Behav fat-sol dye a st-fat 
in anim org,7i. — 244Menge New compd cholin type; acyl deriv a- 
meth'ch, ")8-homoch" (/?-meth'ch), y-homoch, 97. 2;ii.—24sWhite- 
Thomas Tryp proteol Cynoscion regal,iii. — 246Epstein-Bookman 
Form glyc'ol i body,ii7. — 24yHart-Humphrey-Mornson Comp effic f 
growth total-N fr alfalfa hay a corn,i33. — 248Lusk-Riche Anim 
calorim; infl ing am'ac metab,i55 ; 249lbid, infl mix food-st metab,i85. 
— 2^oM cCollum-Steenbock Creatin metab grow pig,209. — 2$iMcCol- 
lum-H aipin-Drescher Synth lecith i hen a charac of lec produced,2i9. — 
2S2Denis Metab cold-bl anim; urin fish,225. — 2^T,Osborne-Mendel 
Mainten w isol prot,233. — 2^4Levene-Birchard Kyrin f ract part'l hydrol 
prot,277. 3;i2. — 2^^Keyes-Gillespie Gas metab bact; prod ferm 
dextr Bcoli, Btyphosis, Bzvelchii,2gi ; 256Ibid, absor O by grow cult 
Bcoli, B'welchii,2,oS- — 2^7 Anderson Org P- ac cot seed meal,3ii. — 258 
Robertson Bld relat'ship i compos serum prot; compar ser hors, rabb, 
rat, ox, norm a fast,325. — 2^gRettger-Neivell Putrefac, proteus gr,34i. 
— 26oLewis Behav hydant deriv in metab ; hydan a ethyl hydan,347. — 
26iDakin Racemiz prot a deriv resul fr tautom ch'ge,357. — 262Folin- 
Macallum Colorim deter uric ac in ur,363. — 26'T,Hunter-Givens Metab 
endog a exog purin (monk),37i. — 264F olin-Lyman Absorp fr stom; 
repl London,389. (Pages, 391.) 

Bio-Chem. Journ. Vl:4\io.—26'^MacLean Phos'tid kidn,333; 266 
Purif phos'tid, 355. — 26yCooper Relat phenol a w-cresol to prot (mech 
o disinfec),362. — 268Bostock Distr N in autol, spec ref deaminiz,388. — 
26gCoppin Efif purin deriv a org compd grow a cell-div plant,4i6. — 
2'joMelvin Glycol i brd,422. — 2'jiHarris-Creighton Reduc FeClg surv 
organ,429. — 2y2WatkynThomas Act opium alkaloid,433. — 2y:^Harding- 
Ruttan Detec aceto-acet ac b sod nit'prus a NH40H,445. — 2y4Smed- 
ley Fat ac butter,45i. (Pages, 129.) 

Biochem. Bull. II:5;9. — 2'jsWinterstein E Schulze,i. — 2y6Rose 
Lit inos-P- acid (phyt),2i.— 277i/arW3' Artif cell,5o.— 278^^0/^ New 



igi3] William J. Gies 3^5 

func peroxidas, trans orcin i orcein,53. — 27gGies Diff thr rub'r ; lipin 
and lipin-sol subst,55. — 2SoRosenbloom Ibid,64. — 2SiWelker Ibid, Diff 
prot; disin'gr col'dion membr b ethtr, 70.— 282Beal-Geiger Ibid, pigm, 
7S.—2SsKahn-Rosenbloom Col'd-N ur, dog tumor hr'st,8,7.—284Howe 
Fast,go.—2SsBerg Contr str musc; surf tens,ioi.— 286£;(/rf3; Prot 
compd,iii. — 28yRosenbloom-Weinberger Eff intrap inj epineph partit 
N uT,i2:^.—288Hallibtirton Bioch Soc, Eng,i28.—28gMendel Proc See 
(2) Diet hyg a hyg phys'l, 15 Int Con Hyg a Demog,i29. — 2goMandel 
Proc See Bioch ine Phar'col (8d) 8 Int Con Appl Ch,i50.—29iLothrop 
Proc Col Un Bioch As'n, 156. (Pages, 210.) 

Subject index of the foregoing bibliography/ Absorpi 77,206-41-64; 
acetaldehso; acetat57; acet-acii4; acet-acet-ac49,5 1,273; acidi 1,89,1 13; acidosög; 
acyl-deriv244 ; adaptat65; adsorp82,96; agei72; "agfa"-lecith38 ; (i-alani48; alb'in 
167; alcoh49,50,fermi05-55; aldeh49; alfa-hay247; al-tract65,i87; alkali8o,i37; 
alkard56,77,272 ; am-aci69-70-<^-8i-^-7,238-48; a-am'butyr-aci 79-94; am'eth'alc 
145; am-grpi7S; am-Ni40-56-5-9-6o,2i7-27; ammon57,273,saltsi77-8i,2o8; amylas 
79,116; anaphyli78-9i ; anem6i ; antag-actiis; antisep26; antiser25; Aplysiid2i5; 
appi7,i02; A-nigerioo,2oo; assim92,227; asym-synth7o ; autoli3i-i,2i6-68. Bact 
i38-9,2S5-6,metah234-5-42,coli,typhos,welch2S5-6; bil-pig2io; bilirub2i8; Bioch- 
Soc-Eng288; blood3,5,7,io-4,39,40-/-'^-5-<5-9,S4,84,ioi-i4-72-7,20i-j-7-i2-4-27-4i- 
58-7o,alk8o,corp58,i62,gasi02,serio4,transfi; brain6,7,226 ; breast283; brom20S; 
butter274. Cai9; calorim222-j-40-7-^-9 ; carbam-reaci74; carbohydr4, 15,20,1 18; 
CO203; CO2I01-65; C-metabi3o; carbox'asii7; carcini44; cardiogr66; caseini23; 
catal68,70, 133-7; celliS5-88-9,277,div269; Cephalop2i5; cerebron-ac22S ; cerebrosid 
226; cholest'as58 ; cholest'oli44; chol-ac2os-ö ; cholin 145-9,244; chymi8o; clay82, 
96, 164; clupeini42; coag9; colamini45; cordion28l ; colloidi24-97,N283 ; Col-Un- 
Bioch-As29i; complem98; CU238; correci68,2i3; corn-gr247; corp54; cot-seed- 
m'l257; creatini5i,metab25o; cresol, m267,^85 ; cult-medi5o; Cynoscton-regal245. 
Deamin268; developi2; dextr(gluc) ; diabet 1 66,232-^ ; di-acet-ac49,5i,273 ; diastas 
22,79,204; di-carb-ac5i ; diffus63,279-8o-J-2 ; digest29,i2i-73-84-<5; digital66; di- 
oxyacet48; disinfec267; drug36; dye243. Eclamp95; eggi2,whi47; electr23,78,8o, 
1 12-22-5; e'lytli5; embry 159-60; energ4,5,20; enzym30-7-2,65,99, 125-43-50-96-7; 
epid-tis28; epineph84,287 ; erythrocy86; ether28o-i; ethy-hydanto26o ; excr55-<5, 
83,128; extr'tivi57,2i7. Fast258-84; fati4,20,i44,24O-j,metab67,sol-dy243; fat-ac 
274; feces76; fermi 13-34-5,255; FeCls27i ; fish252; fistula2i4; food2,8,i4i,249; 
formald6o, 156-5-9-60,21 7; Fucoidesi99; fungiioo,200. Gas io2-73,metabi, 2,3,6,7, 
16,255-0 ; gelati09-i9-8i ; gentianos3i ; gliad236; gluconeogen237 ; glucosam35; 
glucos47,54,84, 137,237-40-1-55; glucosid23o; glucuron-ac37,i66; d-glutam-aci79; 
glycer40,52; glyc-ald48; gly'col 148,246; glycog2i5; glycolys90,270 ; glyc-phen'alan 
180; glyoxyl-ac9i ; growi4i,247-5o-<5-69; guanid22i ; guan-hexos224 ; guanyl-ac 
127,229. H*9,io-7,62,io9-ii ; hay247; HCl7i-^-5-^,io6,204-9; heart 66,156-7-5-61 ; 
"heat-punc"io; hem'porph 195,203; hemicelluli43; hemin 192,2 18; hemogli46-53, 
207-12; hemol6o,87; heroin55 ; homochol244; hungeriS; hydanto26o; H2O2163; 
hyd roll 1,1 37-43-63,254; hy'ox'ami46. Inj-curr23 ; inos-P-ac276 ; Int-Con,(8)-App 

' See explanation on page 298. 



3o6 Biochemiccd Bibliography and Index [Jan. 

Ch290;(i5)-Hyg-a-Dem289; intest92,i8o,2o6-ii ; inul29,metabi26; invert209; I216; 
ironi33. Kerat28; kidn 16,265; Klausnr-reaciio; kyrin254. Lact-ac39,4i-^-j-^-5- 
6-7,131; Iead68,hydri70; lecith38,25i ; /-leucini48; light36; lipassg, 183; lipin 14,27g- 
80; lipoid 14-88, 132; Iipoli3-^; livr39,40-/-9,l 14-33. Mgig; mainten 18,253; tnal- 
ac34; maltas26; manani43; maninotn32; meati8i,223; metabi,2,3,4,5,6,7,8,i5-ö-5, 
24,57,67, i2o-<5-30-76-7,223-32-^-5-4(>--?-S-9-50-2-5-6-6o-5 ; methemogl 146 ; method 
28,80,115-9-54-65-0-7-70-^,205-30-62-6-73; meth'ation 142-92; meth'chol244 ; meth- 
glucioo; meth-guan22i ; Mett-methSg; mlk75,coag62 ; morsk2i5; muscl42-5, 156-7- 
5-^>-6o-i,2i7-85,contr20,285 ; myoproti6i. Ni9,i40-57,227-47-6S-83-7,foodi88-9, 
met57, 176,208; narcot64; nerv- sy 3103-69; neut-regioi ; nomencl99,207 ; nucleas27; 
nucl-ac27, 178,224-^; nucleini78; nuc-proti78; nutri8,i4i-5o-8i-5-9,208-36-53-5. 
Op'm-alk272; optic6,i72; orcin(ein)278; osm-ac86; ovomuc35,i47; oxidasi32; 
oxidat93-<^; 0173,256; /'-oxyphen'eth'ami29. Pancri5,dias79,lipi83; p'thy'dect22i ; 
parenter-f eed6,8 ; Pen-crustac200; pentosi27; peps 11,33,89,121-73-86; peptid238; 
Pepton 187,238; pernieabii5; peroxid278; phenol85,267 ; ph'endiam93-4 ; phloriziö, 
diabet239; phosphatid265-6 ; P-ac76,233-57 ; P19; phos'tung-ac23i ; pho'ch-syniiS; 
phtal-ac-anhy202 ; physostigioS; phytin200-76 ; pigment77, 199,210-82; placent95; 
plant24,i 17,269; plasma54; plasteini85; polemicio7-76-7,220-64 ; porn-tox8i ; poly- 
pep 138-9-80-^-93; polyphen'oxidas99; port-brd2l4,fist2i4; porphyri9o; potent'l 
112; p'p't'niii; pregnani54; propion-ac237 ; protein9,ii,28,92,i75-8o-6-7,253-<^-5- 
6i-7-8i,assim227,compd286,metab24,i77,232; proteol87,24S ; proteus gr 259; pro- 
toprm63,streani64 ; pseudomuc35; ptyalin2i9; purini20-57,263-9;putrefaci79,259; 
pyrol202; pyrorcarb-aci7i-93; pyr-tart-acS3. Quad'urigS; quini36. Racemizat 
261; receptor86; rafinos3i ; reduc27i ; regen98; renin75; respi7,24,83,i02-28,calor 
222,centrioi.quot69; rub'r279,8o-/-<?. Salivii6; salts56,63, 125-82-5; Schulze275; 
secretös; Sec,Bioch-Pharm'l(8d)290,Diet-hyg,hyg-phys'l289; serumiio,258,prot 
258; sexi72; sil-ac97; sod-carb77,NaCl240,NaOH48,Na-nit'prus273; sp-crdi69; 
stachyos32; st'ch78; stomach56-9,io6-77,264,cont7i-<?-j-^; sucras,sucros3o; sugar 
i47,s-free-fermi34-5; S161; surf-ten285; surv-org27i ; synth7o,i8o-5-9,25i. Tau- 
tomch'ge26i ; temp2,io,33; thymusi78,nucl-ac224-8; thymoli62; thyroid97; tissue 
93-4> 1 77. extr 13 ; toxic25 ; toxin8i ; toleranceSS; tranfusi ; tripepti48; trisac'ari43 ; 
tryps33,i09-73-86,proteol245; tryptoph28; tumor283,celli04; typhusi95. Urea24o; 
uric-ac68,i98,262 ; urin37,76,84,i30-4O-66-7-95,22i-52-62-83-7. Water83, 128,240; 
■wheat-bran233. Xanthoproteic-reaci52. Yeasti34-5-S0,fermentaii3. 



BIOCHEMICAL NEWS, NOTES AND COMMENT 

Contents. I. General: Necrology, 307; in memoriam, 307; honors, 308; re- 
tirements, resignations, declinations and appointments, 310; lectures, 312; endow- 
ments, funds and buildings, 313; societies, associations, etc., 314; officers-elect of 
biological organizations, 315 ; miscellaneous items, 316. II. Columbia University 
Biochemical Association: i. General notes, 321; 2. Proceedings of the Associa- 
tion, 322; 3. Columbia Biochemical Department, 324. 

I. GENERAL 

Necrology. David Axenfeld, professor of physiology at 
Perugia. — Carl Bins, professor of pharmacology at Bonn. — Edzvard 
Curtis, emeritus professor of materia medica and therapeutics at 
Columbia University. — Elie de Cyon, some time professor of physi- 
ology at the Academy of Sciences of St. Petersburg, lately of Paris. 
— Wilhelm Ebstein, professor of internal medicine at Göttingen. — 
Humphrey O. Jones, proiQssor of chemistry at Cambridge. — Oszvald 
Kohts, professor emeritus of diseases of children at Strassburg. — 
Ewen Mcintyre, for many years president of the N, Y. College of 
Pharmacy. — /. W. Mallett, professor emeritus of chemistry at the 
University of Virginia. — Hermann Miink, professor emeritus of 
physiology at the veterinary College in Berlin. — Clarence V. Murphy, 
bacteriologist and medical chemist at the Mass. State Sanatorium, 
Rutland. — Aime Pagnoul, formerly director of the Agricultural 
Station at Pas-de-Calais. — Heinrich RittJmusen, professor emeritus 
of agricultural chemistry at Königsberg. — Preston B. Rose, for- 
merly assistant professor of physiological chemistry and toxicology, 
and lecturer on renal diseases, at the University of Michigan. — O. 
T. Williams, lecturer on pharmacology and demonstrator of bio- 
chemistry at the University of Liverpool. 

In memoriam. Lord Lister. The Lister memorials com- 
prise a " Lister International Memorial Fund," f rom which will be 
drawn from time to time a Lister international award for the most 
notable contribution to surgery in any part of the world, and which 
will also Support fellowships and studentships in surgical research; 
a monument in London; a memorial tablet in Westminster Abbey; 

307 



3o8 Biochemical News, Notes and Comment [Jan. 

a monument in Glasgow; and the preservation of the ward in the 
old building which is now being torn down to make way for a new 
building of the Royal Infirmary. This ward will be arranged as it 
was in Lister's time, furnished with contemporary articles and pro- 
vided with exhibits showing the work that Lister did, and with 
articles of a personal nature associated with the man in his work. 
Contributions may be made to any of the memorials, and may be 
sent to Dr. W. W. Keen, 1729 Chestnut Street, Philadelphia. 
Each contributor is asked to designate the particular memorial to 
which he wishes his contribution to be applied (p. 189), 

Paul C. Freer. The July issue of the Philippine Journal of 
Science was a memorial to the late Dr. Paul C. Freer, director of 
the Bureau of Science of the Philippine Islands, dean of the College 
of Medicine and professor of chemistry at the University of the 
Philippines, founder and editor of the Philippine Journal of Science 
(p. 189). 

Honors. Nobel prizes were presented by the King of Sweden 
at a bancjuet in Stockholm, on December 10. Those to whom 
awards had been made were present, including Dr. Alexis Carrel, 
of the Rockefeller Institute for Medical Research (p. 190). — The 
Nobel prize for chemistry has been divided between Professors 
Grignard, of Nancy, and Sabattier, of Toulouse. — Professor Sabat- 
tier has given his portion of the Nobel prize in chemistry to the 
building fund of the Toulouse Institute of Chemistry. — A reception 
was given in honor of Dr. Alexis Carrel, at New York University, 
on November 16, when President Taft, the French ambassador, and 
others, delivered congratulatory addresses and Dr. Carrel responded. 

Order of merit. Dr. Paul Ehrlich, of Frankfort, and Dr. 
Emil Warburg, president of the "Reichsanstalt" at Charlottenberg, 
have been made members of the Bavarian-Maximilian Order, which 
is the highest Bavarian decoration for scientific Services. 

Corresponding member. Prof. F. E. Lloyd, of McGill Uni- 
versity, has been elected a corresponding member of the Centro de 
Sciencias, Letras, e Artes, Campinas, S. Paulo, Brazil, in recogni- 
tion of his work on the desert rubber plant, guayule. 

Honorary member. At its meeting on December 3 the Acad- 



1913] General 309 

emy of Medicine, of Paris, elected Professor Delezenne of the 
Pasteur Institute an honorary member of the section on anatomy 
and physiology. 

Anniversary celebrations. On October 22 Prof. A. Kossei, 
of Heidelberg, celebrated the twenty-fifth anniversary of his pro- 
fessorship. — The twenty-fifth anniversary of Prof. Charles Richefs 
appointment to the chair of physiology in the Faculty of Medicine, 
of Paris, was celebrated on December 22. Professor Chauveau 
presided at the celebration and presented Dr. Riebet with a Fest- 
schrift to which three score scholars had contributed from different 
countries, among them being Pavloff, Kossei, Verworn, Sherring- 
ton, Chauveau and Bouchard. After the presentation, addresses of 
congratulation were made by Professors Landouzy, Dastre, Gley, 
Langlois, and others. 

CoMPLiMENTARY DINNERS. Dr. Jacques Loeb was the guest of 
honor at the second annual dinner of the Columbia University Bio- 
chemical Association, at the Chemists' Club, N. Y., Nov. 6 (p. 322). 
— Prof. R. H. C hütenden will be the guest of his many pupils and 
friends at a dinner at Delmonico's, N. Y., on March i. 

AwARDs OF MEDALs. By the Royal Society: The Davy medal 
to Prof. Otto Wallach, of Göttingen, for his researches on the chem- 
istry of the essential oils and the cycloölefines; the Buchanan medal, 
to Col. Wm. C. Gorgas, of the U. S. Army, chief sanitary officer of 
the Panama Canal zone. — By the Prussian government: The gold 
medal for science to Dr. Walther Nernst, professor of chemistry at 
Berlin. — By the Swedish Medical Society: The Retsius medal to Dr. 
J. N. Langley, professor of physiology at Cambridge, for his work 
on the nervous System. 

Prizes. The Gedge pri::e of Cambridge University has been 
awarded to Mr. A. V. Hill, of Trinity College, for his essay on the 
heat production of amphibian muscle and of cold-blooded animals. 
— Prize for work on diabetes: The medical society of Carlsbad has 
offered $1,000 for the best work or works on "Treatment of dia- 
betes mellitus, with special reference to balneotherapy." Competi- 
tion is open to physicians of all countries, and any language may 
be used. All Communications should be addressed to the Vereini- 
gung Karlsbäder Aertze, Carlsbad, Austria. The jury consists of 



3IO Biochcmical News, Notes and Comment [Jan. 

Professors von Jaksch of Prague, Lüthje of Kiel, Ortner of Vienna, 
Schmidt of Innsbruck, and Dr. Ganz. Essays must be received by 
Dec. 31, 1913. 

Retirements, resignations, declinations, appointments. Re- 
TiREMENTS : Dr. Frana Pf äff , professor of pharmacology and thera- 
peutics, Harvard Medical School. — Captain R. W. Silvester, for 
tvventy years president of Maryland Agricultural College. (He has 
been made president emeritus and librarian of the Institution.) — Dr. 
G. R. Kraus, professor of botany at Würzburg. 

Declinations. Prof. 'Andrezv Boss, in charge of the depart- 
ment of farm management of the department of agriculture, Uni- 
versity of Minnesota, has declined an offer of the position of director 
of the new government demonstration farms and trial gardens, at 
Mandan, N. D. — Prof. E. M. Freem<in, chief of the division of 
plant pathology and assistant dean and secretary of the faculty of 
the College of agriculture, University of Minnesota, has declined an 
offer of the position of chief pathologist of the Kew Botanical Gar- 
dens. The position carries a salary of $4,700. 

Appointments have lately been announced, as follows ■} 

Alabama Polytechnic Institute: Dr. Joseph S. Caldwell (University 
of Chicago), professor of botany. 

Albany Medical College: Dr. Ralph E. Myers (Harvard Medical 
School), instructor in pharmacology. 

Australian Institute of Tropical Medicine (Townsville) : Dr, Young 
(Lister Institute of Preventive Medicine), biochemist. 

British army medical advisory board : Dr. Leonard Hill, civilian 
physiologist. 

Cambridge University: Dr. W. B. Hardy, university lecturer in 
physiology. 

Carnegie Institution, Nutrition Laboratory: Dr. Sergius Morgulis 
(Sheldon fellow, Harvard University, 1911-12, recently investigator 
in the laboratory of Professor Zuntz, Berlin), associate in animal 
metabolism. 

College of Physicians and Surgeons (Baltimore) : Dr. Bartgis 
McGlone, associate in physiology and embryology. 

Cornell College of Agriculture: Mr. M. J. Prucha, assistant pro- 
fessor of plant physiology (promotion). 

^ In this summary of appointments, institutions f rom which resignations 
occurred are named in parenthesis. See also pages 321 and 324. 



1913] General 3 ^ ' 

Georgetown University: Dr. L. W. Fetzer (U. S. Department of 
Agriculture), associate professor of pathological chemistry. 

Guy's Hospital Medical School: Dr. S. Martin Lowry, lecturer on 
chemistry. 

Harvard University: Dr. R. P. Strong (director of the Government 
Biological Laboratory at Manila, professor of tropical medicine in the 
Philippine Medical School), head of the newly established department 
of tropical medicine. 

Industrial: Dr. H. J. Wheeler (Agricultural Experiment Station, 
Rhode Island State College), manager of the agricultural service 
bureau of the American Agricultural Chemical Company (Boston and 
New York). ■ 

Johns Hopkins Medical School : Dr. B. B. Turner, assistant in phar- 
macology. 

Maryland Agricultural College: Prof. T. H. Spence (vice-presi- 
dent), acting president. 

Montefiore Home (New York City) : Dr. H. D. Dakin, Consulting 
chemist; Dr. Nelson W. Janney, chemist; Dr. Isaac Levin (Columbia 
University), director of the department of Cancer research. 

N. Y. College of Pharmacy : Dr. George C. Diekman, associate dean. 

N. Y. University and Bellevue Hospital Medical College: Dr. A. O. 
Gettler, associate professor of chemistry (promotion). 

Oxford University: Dr. W. H. Perkin (University of Manchester), 
Waynflete professor of organic chemistry. 

Rhode Island Agricultural Experiment Station : Dr. Btirt L. Hart- 
well (Rhode Island State College), director, vice Dr. H. J. Wheeler, 
resigned. 

St. Louis University: Dr. P. M. Carrington (U. S. Marine Hospital 
Service), professor of hygiene. 

Siam : Mr. W. B. Freeman, of Denver, director of the public System 
of Irrigation and drainage. 

State University of Kentucky: Dr. Joseph H. Kastle, director of 
the Agricultural Experiment Station and dean of the College of 
Agriculture. 

U. S. Dep't of Agriculture: Dr. Carl L. Aisberg (Bureau of Plant 
Industry), chief of the Bureau of Chemistry (pages 211 and 329). — Dr. 
L. A. Clinton (Conn. Agricultural Experiment Station, Storrs), direc- 
tor of farm-management investigations for the North Atlantic states. 
— Dr. W. D. Bigelow, member of the board of food and drug inspec- 
tion, vice Dr. R. E. DooHttle, resigned. 



312 Biochemical News, Notes and Comment ■ [Jan. 

U. S. Bureau of Mines : Dr. Reid Hunt, member of the commission 
on the Hygiene and danger conditions in mines. 

University of California: Dr. /. IV. Gilmore (College of Hawaii), 
head of the department of agronomy, College of Agriculture. — Dr. 
H. J. Webher (Cornell College of Agriculture), director of the Citrus 
Experiment Station and dean of the Graduate School of Tropica! 
Agriculture. 

University of Chicago: Appointments necessitated by the death of 
Prof. Waldemar Koch^ — Dr. Fred C. Koch, instructor in physiological 
chemistry; Dr. Shiro Tashiro, assistant in physiological chemistry; 
Miss Mathilde Koch, research assistant in physiological chemistry; 
Dr. G. L. Kite, assistant in pharmacology. (Associate professor S. A. 
Matthews is conducting the course in pharmacology.) 

University of Illinois: Dr. C. W. Allee (University of Chicago), 
instructor in plant physiology. 

University of Kansas : Dr. F. P. Chillengworth, assistant professor 
of physiology. — Dr. C. A. Shull (University of Chicago), assistant pro- 
fessor of plant physiology. 

Yale University: Dr. F. P. Underhill, professor of pathological 
chemistry in the Medical School. 

Lectures. Middleton Goldsmith lectures of the N. Y. 
Pathological Society. Dr. E. F. Bashford, director of the Im- 
perial Cancer Research Fund of England, delivered two lectures on 
"Cancer" at the N. Y. Academy of Medicine, on the evenings of 
October 2 and 4. 

Lectures by visiting members of the 15TH International 
Congress on Hygiene and Demograph y (p. 129). Prof. Max 
Riibner, Berlin: (N. Y. Academy of Medicine), Wesley M. Car- 
penter lecture, Oct. 3, The life of a cell; Harvey lecture, Oct. 5, 
Modern steam sterilization ; (N. Y. University and Bellevue Hos- 
pital Medical College), Herter lectures (5), Energy problems in 
nutrition, Oct. 7-1 1. — Prof. Carl von Noorden, Vienna: (N. Y. 
Post-Graduate Medical School), lectures on New aspects of the 
pathologic treatment of diabetes, and Diagnosis and treatment of 
nephritis, October 29-31; (Johns Hopkins Hospital), The princi- 
ples of treatment of diabetes mellitus, Nov. 2; (St. Louis Medical 
Society), Treatment of acetonuria, Sept. 30. — Prof. Hermann 

^Biochemical Bulletin, 1912, i, pp. Z72 and 522. 



1913] General 3 1 3 

Straiiss, Berlin: (N. Y. Post-Graduate Medical School), Gastric 
secretion from the therapeutic point of view, Oct. 14, and The 
method and purpose of dechlorination in nephritis, Oct. 15. 

HuxLEY LECTURE. Dr. Simon Flexner delivered, at Charing 
Gross Hospital Medical College, on October 31, a Huxley lecture on 
Recent advances in science in relation to practical medicine. 

MiscELLANEOUS ITEMS. Prof. M. T. Bogcrt, President of the 
Society of Ghemical Industry, lectured before the McGill Chemical 
Society, Montreal, Dec. 16, on The Classification of carbon Com- 
pounds, and in the evening addressed the Montreal members of the 
Society of Chemical Industry at a banquet at Coopers Limited. On 
the following day he addressed the Toronto members of the society 
at a banquet at the Engineers' Club, Toronto, on A closer Coopera- 
tion between the universities and chemical Industries. 

At McGill University the annual university lecture for the cur- 
rent year was delivered, Oct. 8, by Prof. F. E. Lloyd, on The arti- 
ficial ripening of bitter fruits. 

Prof. /. /. R. MacLeod recently delivered, at the University of 
London, eight lectures on Carbohydrate metabolism. 

Endowments, funds and buildings. Funds and endow- 
MENTS. The executors of the estate of George Crocker have filed 
their final accounting with the courts which shows that Columbia 
University has received from the estate $1,566,635 for the cancer 
research fund (p. 194). — Mr. AustenChamberlain has received £48,- 
000 tovvards the £100,000 which he is raising for the London School 
of Tropical Medicine. — Mr. George F. Baker, president of the First 
National Bank of New York, has given a large sum, reported to be 
$2,000,000, to effect an alliance between the New York Hospital 
and the Cornell University Medical College. — Dr. John C. Hem- 
meter, University of Maryland, made, at the celebration of academic 
day, November 12, a gift of $10,000 as a beginning on the endow- 
ment of the chair for experimental physiology. — An annual fund 
of $15,000, to Support research in medicine at the University of 
Toronto, has been subscribed for five years by a few Citizens of 
Toronto. 

Buildings and equipment. The cornerstone of the new dis- 
pensary building of the College of Medicine of Syracuse University 



314 Biochemical News, Notes and Comment [Jan. 

was laid on December 14. — A bronze bust of Dr. E. W. Hilgard, 
emeritus professor at the University of California, was recently 
unveiled in the foyer of the new agricultural hall when the building 
was dedicated. The occasion was also marked by the formal inves- 
titure of Prof. Thomas F. Hunt as dean of the department of agri- 
culture. — Mr. Andrew Carnegie has offered to the University of 
Paris the last $20,000 necessary for equipping the new Institute of 
Chemistry in course of erection in the Rue Pierre Curie. 

Societies, associations, etc. American Association for the 
Advancement of Science. A füll account of the proceedings has 
been published in Science, issue of Jan. 10, p. 41. 

Federation of American Societies for Experimental Biol- 
OGY. Proceedings are published in this volume, p. 271. 

RusH Society. This society, established through the initiative 
of the Medical Department of the University of Pennsylvania, was 
organized November 21. Its objects are similar to those of the 
Harvey Society (New York), namely, the diffusion, by lectures, of 
knowledge conceming recent advances in the medical and the gen- 
eral biological sciences, and public hygiene. The officers are : Richard 
M. Pearce, president; Alfred Stengel, vice-president ; William Pep- 
per, secretary-treasurer, and A. E. Taylor, A. C. Abbott and H. H. 
Donaldson, councilors. 

SiXTH International Congress for General and Medical 
Electrology and Radiology. This congress, held in Prague 
during the first week of November, was attended by 760 members. 
About 130 papers were read. An interesting account of the pro- 
ceedings was published in the Journal of the American Medical 
Associatio7i, Dec. 14, p. 2169, 

New Hospital Association. Delegates from twenty-nine 
hospital dispensaries met recently at the N. Y. Academy of Medicine 
and organized an association to be known as the Associated Out- 
Patienf Clinics of the City of Nezu York. The association aims to 
coördinate the work of existing dispensaries and out-patient clinics, 
to eliminate unworthy applicants for treatment, and to promote 
proper Standards of treatment, economy and efficiency in dispensary 
management. 



1913] General 3 1 5 

New Orleans Academy of Science. The newly reorganized 
New Orleans Academy of Science held its first regulär meeting on 
November 12. As now organized it consists of sixteen sections 
with a chairman for each section, among them Biology and Physiol- 
ogy, Gustav Mann, chairman. 

The N. Y. Gastro-Enterological Society was founded, De- 
cember 3, largely through the efforts of Dr. G. R. Lockwood. The 
object of the society, as the name implies, relates to the study and 
discussion of gastro-intestinal diseases. At the meeting for Organ- 
ization, Dr. Max Einhorn was elected president, Dr. G. R. Lock- 
wood, vice President and Dr. Harold Barclay, secretary-treasurer, 
for 19 12-13. The officers constitute the Council. The charter 
members are Drs. Harold Barclay, George E. Brewer, H. S. Carter, 
Max Einhorn, Ellsworth Eliot, Wm. J. Gies, W. Van V. Hayes, 
Lucius W. Hotchkiss, Ludwig Käst, Edward Leaming, G. R. Lock- 
wood, Wm. G. Lyle, Charles Peck, A. R. Stern. The society will 
meet at the homes of members, on the second Mondays of January, 
March, May and November. Harold Barclay, Secretary (68 
West 56th Street, New York). 

Officers-elect of biological organizations. American So- 
ciety OF Biological Chemists (p. 275). 

American Society for Pharmacology and Experimental 
Therapeutics (p. 279). 

American Physiological Society (p. 271). 

American Association for the Advancement of Science : 
President, E. B. Wilson. 

Society of American Bacteriologists : President, C.-E. A. 
Winslow ; vice-president, Chas. E. Marshall ; secretary-treasurer, A. 
Parker Hitchens; Council, W. J. MacNeal, L. F. Rettger, D. H. 
Bergey, H. A. Harding; delegate to Council of American Associa- 
tion for the Advancement of Science, S. E. Prescott. 

American Society of Naturalists : President, Ross G. Har- 
rison ; vice-president, E. M. East ; secretary, B. M. Davis ; treasiirer, 
J. Arthur Harris; additional members of the executive committee, 
A. P. Mathews and A. L. Treadwell. 

American Phytopathological Society: President, F. C. 



3i6 Biochcmical News, Notes and Comment • [Jan. 

Stewart; vicc-prcsident, Haven Metcalf ; secretary-treasurer, C. L, 
Shear; coimcilor, W. J. Morse. 

BoTANicAL Society of America: President, D. H. Campbell; 
vice-president, M. A. Howe; treasiirer, Arthur Hollick; secretary, 
George T. Moore; coiincilors, G. F. Atkinson, R. A. Harper and 
William Trelease. 

American Society of Zoologists : President, Henry B. Ward. 
— Eastern Branch: President, Raymond Pearl; vice-president, Alex- 
ander Petrimkevitch ; secretary-treasurer, Caswell Grave ; additional 
memhers of the executive conimittee, C. E. McClung, R. G. Harri- 
son (elected, 1910), and H. E. Jordan (elected, 191 1). — Central 
Branch: The present officers continue until the next meeting of this 
branch (Biochemical Bulletin, 1912, i, p. 494). 

Society of Chemical Industry : President, Marston T. Bogert. 

Miscellaneous items. Medallion of van't Hoff. The 
Dutch sculptor, Pier Pander (Rome), has executed a bronze medal- 
lion of van't Hoff. Any one desiringto purchase a copy of it should 
address Prof. Ernst Cohen, van't Hoff Laboratorium, University, 
Utrecht, Holland. If 100 copies are sold, the price will be 6.50 
Marks (5.50 Marks if 200 copies are sold). The medallion has 
been executed after a portrait relief in marble by Pier Pander. 

Citrus Experiment Station. The University of California 
has for several years maintained four separate sub-stations in south- 
ern California. These will be united into an enlarged research Sta- 
tion which will probably be located at Riverside. While this Station 
will be designated the Citrus Experiment Station, after the domi- 
nant industry of southem California, the work will relate to all 
crops grown in that region. The coupling of the Graduate School 
of Tropical Agriculture with the Station for Agricultural Research 
will make it unique among our agricultural experiment stations. 

Coroner's CONSULTANTS. Coroner Peter M. Hoffman, of Cook 
County, 111., has named Drs. John H. Long, Walter S. Haines, Lud- 
vig Hektoen and John A. Wesener, and Chief Justice Harry Olsen 
of the Municipal Court, as his Consulting staff. With the aid of this 
staff of Consultants, and the establishment of a chemical laboratory, 
the coroner hopes to reduce the number of fatalities from poison- 



1913] General 3 ' 7 

ing which annually swell the list of deaths, registered as " suspici- 
ous," that require investigätion by the coroner. The salary of the 
chemist in charge of the laboratory will be $2,500 per annum; there 
will be one assistant. Applications may be sent to the Coroner, 
Room 500, County Building, Chicago, 111. 

The Harriman Research Laboratory, which operates a 
building on the grounds of Roosevelt Hospital (N. Y.), has been 
incorporated. It was established in 1910 and is maintained by Mrs. 

E. H. Harriman for the study of chemical problems connected with 
disease. 

International bureau of foodstuffs. Delegates from the 
various governments represented at the international congress for 
the investigätion of methods of analysis have established, in Paris, 
a permanent international bureau of analyses of foodstuffs. 

New JOURNAL of science. The Publishing house of Julius 
Springer, Berlin, announces the publication, beginning January 3, 
1913, of a new weekly Journal, Die Naturwissenschaften, which, 
according to the announcement, " für den deutschen Wissenschafts- 
betrieb ungefähr das leisten soll, was die 'Nature' für den englischen 
und die 'Science' für den amerikanischen leisten." Each number 
will contain about 24 pages; the subscription price is 24 Marks. 
The Naturwissenschaftliche Rundschau,, edited by Prof. W. Sklarek 
and published by Friedrich Vieweg und Sohn, which for twenty- 
seven years has maintained high scientific Standards, will be merged 
in the new Journal. 

" Pawlow." I note with interest Professor Halsted's protest^ 
against the spelling of Lobachevski's name with a "w," a sort of 
scientific Wellerism which Teutonic influence has foisted upon the 
English language. Is it too much to hope that some day we may 
find American physiologists referring to Pavloff instead of to Paw- 
low, or is it true that in such mixed crosses, as the heredity experts 
would say, German pedantry is prepotent over common sense? /. 

F. Abbott (Science, 1912, xxxvi, p. 595). 

Artificial milk produced from soya beans. An artificial 
milk manufactured from soya beans, which is said to contain "all the 

' Halsted : Science, 1912, xxxv, p. 736. \ 



3i8 Biochemical News, Notes and Comment ' [Jan. 

Clements" of the best milk and can be used for the same purposes, 
was recently shown to a gathering of scientists in London. The 
artificial milk is said to be more digestible than ordinary milk and 
its Cream more nourishing. It can be used for all cooking purposes 
and good cheese can be made from it, but it will not yield butter. 
As it is germ-free it will keep longer than milk. The discovery is 
the work of three Germans who spent three years in perfecting it. 
The process of manufacture is simple and always produces the same 
result. The "milk" is not touched by band or exposed to atmos- 
pheric influence until it is poured into bottles for delivery. The 
"milk " can be sold at 6 cents a quart, which is 2 cents cheaper than 
the cost of London milk, and the cheese at 6 cents a pound {Journ. 
Amer. Med. Assn., 1912, lix, p. 1637). 

BiocHEMisTRY IN ENGLAND. During the past year a move- 
ment toward the Organization and closer social affiliation of those 
biologists and chemists who are interested in the investigation of 
Problems common to the two branches of science has resulted in the 
formation of the Biochemical Club of England. . . . The move- 
ment cannot fail to lead to meetings which will be stimulating and 
füll of interest, if one may judge by the success which has attended 
the similar Organization, the American Society of Biological Chem- 
ists, since its Organization six years ago. Chemical points of view 
are rapidly gaining a preeminent position in the biological and 
medical sciences. The closer association and Cooperation of investi- 
gators in medicine with scientists who are attacking allied problems 
in other fields, such as agriculture, plant physiology and pathology, 
microbiology in its industrial applications, etc., is certain to afford 
advantages of mutual value. In the United States, compared with 
Germany for example, there has always been less tendency for the 
Student of chemistry in medicine to hold aloof from the biochemist 
proper. Theconsideration of medical problems from a more strictly 
biological point of view is a timely attitude, and the new English 
Organization with its broad affiliations is a commendable one {Edi- 
torial: Journ. Amer. Med. Assn., 1912, lix, p. 1803). 

Inventor of GELATIN CAPSULES. It has bccu incorrcctly as- 
sumed that the apothecary Gross von Figely, of Vienna, in 1865, 
introduced gelatin as a vehicle for medicines. The Londoti Journal 



1913] General 3 ^ 9 

of Arts (August, 1848, page 42) contains an article which shows 
that the real inventor of gelatin capsules was James Murdoch, of 
London. In England he was granted a patent in May, 1848, enti- 
tled "An invention for preserving medicines, etc., in solid, liquid, 
or powdered form, protected from the air." The description fol- 
lows. "The capsule consists of two parts, which fit together; one 
part forms a case to receive the substance to be preserved, and the 
other forms the cover, which fits tightly over the case; by simply 
moistening the edges, the capsule can be closed airtight. The most 
suitable form is a cylinder with hemispherical ends. They are made 
as follows: "Polished metallic rods, of the form and size of the 
case and cover, are dipped by pairs in a Solution of gelatin, which 
is drawn off from the rods after drying. In order to facilitate the 
loosening of the capsules, the rod may be slightly smeared with oil 
or fat. Each rod must have an opening from end to end, to allow 
the air to escape after dipping in the gelatin. In addition to animal 
jellies, starch paste and other mucilaginous liquids can be used. For 
medicinal substances Iceland moss is best." F. M. Feldhaus (Chem- 
iker Zeitung, No. 74, 1912), 

The prevention of senility. Dr. Metchnikoff recently ad- 
dressed a letter to a leading Hungarian daily paper in which he pub- 
lished the results of his latest investigations. His scientific discov- 
eries, he says, have been so exaggerated in lay papers that he has 
resolved henceforth to write direct to the public. After mentioning 
his early theory that the length of lifeamonganimals variesinversely 
with the length of the large intestine, and his later theory that sen- 
ility is the consequence of the effects of toxins (chiefly phenols) 
produced by the intestinal bacteria, Metchnikoff refers to experi- 
ments in which he actually succeeded in producing, in apes, senile 
degeneration by giving them for some time small doses of />-cresol. 
These were the fundamental investigations which led to the Solution 
of the question as to how the action of the intestinal bacteria might 
be checked or diminished. The lactic acid bacillus has proved to 
be the best for this purpose. An obstacle to the work of the lactic 
acid bacilli has been their need of sugar, which does not reach the 
rectum in a usable form. Dr. Metchnikoff and Dr. Wollman, his 
pupil, have overcome this difficulty by cultivating bacteria that pro- 



320 Biochemical News, Notes and Comment ■ [Jan. 

duce sugar from starch. It is now possible to supply a diet capable 
of supporting the bacillus that limits the action of the intestinal 
flora. Of course, concludes Metchnikoff, the struggle against senility 
is not concluded. Whether these discoveries will actually tend 
toward the lengthening of human life is a question of the future, 
but it cannot be denied that a beginning has been made, and we have 
reason to hope that from these investigations mankind may derive 
practical benefit (Journ. Amer. Med. Assn., 1912, lix, p. 815). 

Electrons. Abstract of an address before the American Philo- 
sophical Society at Philadelphia, Nov. i, by Sir William Ramsay. 
The actual existence of electrons in motion has been conclusively 
demonstrated ; the mass of an electron is not far from one iSßoth of 
that of an atom of hydrogen; and as the mass of an atom of hydro- 
gen is now known with fair accuracy, that of an electron is nearly 
0.8X10'^'^ gram. Electrons in motion are negative electricity; 
they constitute a form of matter, which, at present, has more claim 
to the term "elementary" than have most of the " Clements." In- 
deed, metals must be regarded as Compound substances, of which 
one component consists of one or more electrons; these electrons 
are, as a rule, not very firmly attached, as is evident from the gener- 
ally easy oxidation of most metals. Non-metals are also composed 
partly of electrons, not so easily detached. The " combination of 
Clements with each other" consists in the shifting of one or more 
electrons from the more metallic to the less metallic dement; no 
doubt it will some day be possible to give " structural formulae " to 
the Clements, showing the relationship in position, or in directed mo- 
tion, between the true Clements and their attached electrons. The 
word " electricity" has a dual meaning; it may mean first, an assem- 
bly of electrons, stationary or in motion; or second, waves in the 
ether, produced by the stopping or starting of electrons in motion. 
The motion of electrons constitutes one factor of electrical en- 
ergy; wave-motion in the ether can be used as a means of gene- 
erating electrical energy, by employing the waves in making elec- 
trons move. Progress in man's command of natural forces has 
been made by learning how to direct and control the motion of 
masses — in other words, by acquiring a knowledge of mechanics; 
progress in the future will consist in acquiring the power to control 



1913] Columbia Biochemical Association 321 

and direct the motions o£ electrons. This has already been largely 
achieved by electric contrivances : it is, however, only by the use of 
concrete ideas regarding the "material" used, viz., electricity, that 
the progress of invention and discovery can be hastened {Science, 
191 2, xxxvi, p. 684). 

IL COLUMBIA UNIVERSITY BIOCHEMICAL ASSOCIATION 

I. General notes 

Marriages. On October 2, Miss Rowena Farmer and Dr. 
Oscar M. Schloss. — On October 15, Miss Ethel P. Willcox and Dr. 
Harold E. Woodward. 

Appointments. Columbia University : Dr. Russell L. Cecil, 
Proudfit fellow in medicine; Dr. Wm. H. Woglom, assistant pro- 
fessor engaged in cancer research (page 201). — Cornell University 
Medical College : Dr. Stanley R. Benedict, professor of chemistry 
(promotion). — Johns Hopkins Medical School: Dr. Edwards A. 
Park (Columbia University), assistant professor of pediatrics. — 
Massachusetts Agricultural College: Dr. H. D. Goodale (Carnegie 
Institution, Station for Experimental Evolution), research biologist. 
department of poultry husbandry. — Turck Research Laboratory 
(New York) : Dr. Anton R. Rose (Columbia University), chemist. 
— University of Texas (Austin) : Mary E. Gearing (Public 
Schools, Houston, Tex.), professor of domestic science; Anna E. 
Richardson (Agnes Scott College, Decatur, Ga.), assistant pro- 
fessor of domestic science. — U. S. Department of Agriculture, 
Bureau of Chemistry: Carl L. Aisberg, chief chemist (p. 211) ; H. 
E. Buchbinder, assistant chemist. (P. 324.) 

New members and ofEcers-elect of societies. American So- 
ciety of Biological Chemists : Members, Louis Baumann, Samuel 
Bookman, Ernest D. Clark, Isidor Greenwald, Alfred P. Lothrop; 
Nominating Committee, Carl L. Aisberg, P. B. Hawk and Alfred 
N. Richards; Lipoid nomenclature committee, Wm. J. Gies; Com- 
mittee on the Organization of a federation of biological societies, 
Wm. J. Gies (p. 277) . — American Society of Naturalists : E. Newton 
Harvey. — American Society of Zoologists : E. Newton Harvey. — 
Phi Lambda Upsilon: National president, A. D. Emmett; national 
secretary, H. L. Fisher; registrar, George D. Beal. — Society for 



322 Biochemical News, Notes and Comment . [Jan. 

Clinical Serology and Hematology (New York) : Secretary, D. J. 
Kaliski. — Nu Sigma Nu Alumni Association (New York) : Execu- 
tive Committee, Wm. K. Terriberry; Nominating Committee, Ralph 
G. Stillman. 

2. Proceedings of the Association 

Second annual dinner. The second annual dinner of the As- 
sociation (and the first meeting of the Association for the academic 
year I9i2-'i3) was held at the Chemists' Club, 52 East 4ist Street, 
on Wednesday evening, November 6.^ About 125 members and 
their guests were present, the attendance being so large, in fact, that 
the main dining room of the Club was filled to capacity and it was 
necessary to use a room on the floor above for the accommodation 
of about 20 members. 

The dinner was given in honor of Dr. Jacques Loeb, of the 
Rockefeiler Institute for Medical Research. AdeHghtfulhalf hour's 
infomial social gathering preceded the banquet. The president, Dr. 
Walter H. Eddy, was the toastmaster and appropriately introduced 
the Speakers of the evening. Prof. Lafayette B. Mendel, of Yale 
University, spoke briefly on the characteristics of scientific men. 
Prof. Marston T. Bogert, of Columbia University, President of the 
Society of Chemical Industry, brought to the Association, and its 
guest of honor, the greetings of that Society. Profs. C.-E. A. Wins- 
low, of the College of the City of New York, and Graham Lusk, 
of the Cornell University Medical College, also spoke informally 
and in a very entertaining way. The main address was delivered 
by Dr. Loeb, who gave a very interesting and instructive account of 
the results of some of his recent work on the permeability of cells. 

Prof. Max Morse, of Trinity College, proposed that Dr. Loeb 
be elected an honorary member of the Association. The motion was 
seconded by Dr. E. Newton Harvey, of Princeton University, and 
unanimously carried by a rising vote. 

The names of those present and the groupings at the tables are 
indicated on pages 323 and 324. 

* An account of the first annual dinner, in honor of Prof. R. H. Chittenden, 
was published in the Biochemical Bulletin, 191 i, i, pp. 334-339. 



I9I3] 



Columbia Biochemical Association 



323 



*Charles Baskerville 
*Marston T. Bogert 
Walter H. Eddy 
*Jacques Loeb 
*Graham Lusk 
*S. J. Meltzer 
*Lafayette B. Mendel 
*C-E. A. Winslow 

Ella Hazel Clark 
Helen Gavin 
*Marie L. Minor 
Helen G. Russell 
Emily C. Seaman 
Mary B. Stark 
Helen S. Watt 
*Mary D. Womack 

J. J. Bronfenbrenner 
tJ. G. M. Bullowa 
A. J. Goldfarb 
Benjamin Horowitz 
Louis Hussakof 
fMax Kahn 
fWilliam Weinberger 
Charles Weisman 

*Ch'lotte G. Bultman 
Mary C. de Garmo 
*tMary B. Kirkbride 
Jessie A. Moore 
*Fairfax T. Proudfit 
*Mird D. Schlesinger 

Tula Lake Harkey 
*Israel S. Kleiner 
Alfred P. Lothrop 
Anton R, Rose 
*Mary D. S. Rose 

* Guest. 



* James Ewing 
*Cyrus W. Field 
Nellis B. Foster 
F. G. Goodridge 
*P. A. Levene 
♦William H. Park 
*E. E. Smith 
fAlexander Smith 
*Karl Vogel 

*John Auer 
C. Stuart Gager 
*Walter A. Jacobs 
*F. H. McCrudden 
H. O. Mosenthal 
*Edgar W. Olive 
*D. D. Van Slyke 

*Jerome Alexander 
*tJ. P- Atkinson 
*Edwin J. Banzhaf 
Charles F. Bolduan 
fWm. B. Boyd 
fF. T. Van Beuren 
Harry Wessler 
tH. B. Wilcox 

B. C. Gruenberg 
Max Morse 
Raymond C. Osburn 

*Louisa Bruckman 
*Harriet C. Jacobson 
Marguerite T. Lee 
Helen McClure 

Arthur Knudson 
♦fAlbert Plaut 
Edward Plaut 
*Eugene Unna 



*E. H. Bartley 
♦Walter A. Bastedo 
*tS. P. Beebe 
♦Charles A. Doremus 
♦Henry C. Sherman 
Matthew Steel 
♦Frederick H. Sykes 
♦H. T. Vulte 

Ula M. Dow 
Ada M. Field 
Ruth S. Finch 
♦Edith C. Keefer 
♦Ethel MacMillan 
♦Alice E. Skinner 
♦Wilhelmina Spohr 
Helen B. Thompson 

♦Ronald M. Ferry 
T. Stuart Hart 
Paul E. Howe 
♦Ewing H. Rand 

Anna M. Connelly 
♦Mathilda L. Mayer 
Elizabeth Rothermel 
Ethel W. Wickwire 

Harvey B. Clough 
Samuel Gitlow 
Fred W. Hartwell 
C. A. Mathewson 

Donald Gordon 
John L. Kantor 
Daniel R. Lucas 
Ralph G. Stillman 



t Detained or obliged to leave before the conclusion of the dinner. 



324 



Biochcmical Ncivs, Notes and Comment' 



[Jan. 



*Orabel Chilton 
♦Helen Ide Gray 
Alice H. McKinney 
*Bessie G. Pond 

S. R. Benedict 
R. A. Cooke 
*M. S. Fine 
*V. C. Myers 

*tM. I. Falk 
*R. G. Reese 
fWm. W. Tracey 
G. W. Vandegrift 



Will H. Chapman 
*Fred'k H. Morrison 
*Harold E. Smith 

Ernst Boas 
Ernest D. Clark 
Harry L. Fisher 
Ross A. Gortner 
Isidor Greenwald 
E. Newton Harvey 
Michael Heidelberger 
*John J. Kenny 
J. Buren Sidbury 
Louis E. Wise 



William J. Gies 
Joseph S. Hepburn 
Walter F. Hume 
Walter M. Kraus 
S. Kubushiro 
E. G. Miller, Jr. 
P. W. Punnett 
*T. B. Reed 
Grover Tracy 



Proceedings of the eighth scientific meeting. The second 
meeting of the Association for the academic year 1912-13 was held 
at the Columbia Medical School, on Friday, Dec. 6, at 4.15 p. m., 
instead of the regulär weekly departmental seminar. At Dr. Gies' 
Suggestion the Association began, with this meeting, a series of 
quarterly sessions for the presentation of the results of research 
by its members. As presiding officer at the seminars, Dr. Gies out- 
lined the plan and purpose of these scientific sessions and formally 
turned over the meeting to the Association. President Eddy then 
took the chair. The scientific program and abstracts of the papers 
are given on page 284. 

The remaining meetings of this series, for the year I9i2-'i3, 
will be held on February 7, April 4 and June 2. Abstracts of the 
Communications will be published in the succeeding issues of the 
BiocHEMicAL Bulletin, 

Alfred P. Lothrop, Secretary. 



3. Columbia Biochemical Department 

Resignations and appointments. Stapf. The following 

further changes in the stafT, for the year I9i2-'i3, \vere officially 

authorized during the quarter ending Dec. 31 : Dr. Jacob Rosen- 

hloom, associate, resigned to accept the assistant professorship of 

* Guest. 

t Detained or obliged to leave before the conclusion of the dinner. 



1913] Columbia Biochemical Association 325 

biochemistry at the University of Pittsburgh. — Dr. Herman O. 
Mosenthal, instructor, appointed associate, vice Dr. Rosenbloom re- 
signed. — Dr. Max Kahn appointed instructor, vice Dr. Mosenthal 
promoted. — Dr. Clayton S. Smith, instructor, resigned to accept an 
assistantship in pharmacology in the Bureau of Chemistry, U. S. 
Department of Agriculture, Washington. — Dr. Louis E. Wise ap- 
pointed instructor, vice Dr. Smith resigned (page 203). 

The retirement of Drs. Rosenbloom and Smith from the depart- 
ment as noted above, after active and very successful terms of Serv- 
ice, occasioned deep regret among their associates at Columbia, 
whose hearty good wishes attend them in their new fields of use- 
fulness. ( See bibliography, below. ) 

Students. Arbuckle Sugar Co. (Brooklyn) : Abraham Gross, 
research chemist. — Harriman Research Laboratory (Roosevelt Hos- 
pital, N. Y.) : Marston L. Hamlin, research assistant. — Industrial 
School (New Bedford, Mass.) : Constanze C. Hart (Teachers Col- 
lege), assistant. — State Normal School (Truro, N. S.) : Blanche R. 
Harris (Teachers College), assistant. — Texas (North) State Nor- 
mal School: Blanche E. Shuffer (Teachers College), professor of 
home economics. — N. Y. University and Bellevue Hospital Medical 
College: Percy W. Punnett, assistant in chemistry. — ^University of 
Kentucky: Mary E. Sweeny, head of extension department. — Uni- 
versity of Porto Rico; L. A. Robinson, professor of psychology. — 
Washington State College (Pullman) : Louise McDanell, instructor 
in domestic science. — West High School (Rochester, N. Y.) : David 
F. Renshaw, instructor in chemistry. 

Dr. Rosenbloom's career. Jacob Rosenbloom was born in 
Braddock, Pa., on Feb. 25, 1884. His early education was received 
in the public schools and high school of North Braddock, Pa. At 
the end of a four-year course at the University of Pittsburgh he 
received the degree of B.S. in chemistry, in 1905. From 1905 to 
1909 he was a Student here at the College of Physicians and Sur- 
geons, receiving the degrees of M.D. and Ph.D. in 1909. His ma- 
jor subject for the Ph.D. degree was biological chemistry, with 
Professor Gies. 

Dr. Rosenbloom was assistant in this department for the year 
1909-19 10; associate (also assistant pathologist to the German 



326 BiocJicmical News, Notes and Commcnt [Jan. 

Hospital, N. Y.), during 1910-12. The summer of 1912 was spent 
at Johns Hopkins University in clinical medicine. Last October he 
received and accepted appointment as assistant professor of bio- 
chemistry in the University of Pittsburgh. 

Dr. Rosenbloom is a Fellow of the American Association for the 
Advancement of Science, a member of the American Society of Bio- 
logical Chemists, Society for Experimental Biology and Medicine, 
American Chemical Society, Chemists' Club of New York, Sigma 
Xi, International Psychoanalytic Association, and the Society for 
Biological Research of the University of Pittsburgh. He married, 
in June, 191 1, Miss Merla Cohen of Baltimore, Md. 

Dr. Rosenbloom's publications. 1905. A colorimetric method 
for the determination of tungsten; Thesis for the degree of B.S., Univ. 
of Pittsburgh. 

1907. Some azolitmin Compounds of mucoids, nucleoproteins and 
other proteins, with exhibition of products (with Wm. J. Gies) ; 
Proc. Amer. Soc. Biol. Chem., i, p. 48; Jour. Biol. Chem., iii, p. xxxix. 

1909. A contribution to the study of the nature and origin of the 
Bence Jones protein, with bibliography ; Dissertation, Columbia Uni- 
versity. Pp. 64. 

19 IG. On the effects and fate of injected connective tissue mucoid 
(with Wm. J. Gies) ; Proc. Amer. Soc. Biol. Chem., i, p. 271 ; Jour. 
Biol. Chem., vii, p. Iviii. — Is the Bence Jones protein produced from 
osseoalbumoid ? ; Ibid., p. 227 and p. xiv. — A study of the duodenal 
Contents in man (with M. Einhorn) ; Arch. Internal Med., vi, p. 666; 
Int. Beitr. z. Path. 11. Ther. d. Ernähr. Stoffw. u. Verd'krank., ii, p. 
184. 

191 1. A histological and chemical study of the fatty matter of 
normal and cryptorchid testes (with F. M. Hanes) ; Jour. Exp. Med., 
xiii, p. 355. — A study of the nitrogen metabolism in three cases of 
duodenal alimentation (with M. Einhorn) ; Amer. Jour. Med. Sei., 
cxlii, p. 7 ; Int. Beitr. z. Path. u. Ther. d. Ernähr. Stoffw. u. Verd'krank., 
iii, p. 5. — ^A new process for the purification of lipins, with demon- 
strations (with Wm. J. Gies) ; Proc. Amer. Soc. Biol. Chem., ii, p. 8; 
Jour. Biol. Chem., ix, p. xiv. — A demonstration of the osmotic pressure 
exerted by fat (with Wm. J. Gies) ; Proc. Soc. Exp. Biol. and Med., 
viii, p. 71. — The effects of intraperitoneal injections of epinephrin on 
the partition of nitrogen in the urine of dogs (with W. Weinberger) ; 
Ibid., p. 131. — Experiments on the diffusibility of cholesterol esters and 



1913] Columbia Biochemical Association 327 

of lecithan Compounds (with E. Boas) ; Ibid., p. 132. — The importance 
of the colloidal nitrogen in the urine in the diagnosis of Cancer (with 
M. Einhorn and M. Kahn) ; Anier. Jour. of Gastro-Enter., i, p. 12; 
Arch. f. Verdauungskrank., xvii, p. 557. — On the lipins of the heart 
muscle of the ox; Science, xxxiv, p. 221 ; Biochem, Bull., i, p. 114. — 
The effect of pregnancy on the Upins of the ovary and corpus luteum 
of the cow; Ibid., p. 222 and p. 115. — ^A proposed chemical Classifica- 
tion of lipins, with a note on the intimate relation between cholesterols 
and bile salts (with Wm. J. Gies) ; Ibid., p. 51. — Intracellular lipins; 
Ibid., p. 75. — A review of the history of Bence Jones protein and mul- 
tiple myeloma; Ibid., p. 161. — The older theories of edema; Ibid.. 

P- 275- 

1912. Osseoalbumoid as a possible precursor of Bence Jones pro- 
tein; Arch. Internal Med., ix, p. 236. — Spontaneously precipitated 
Bence Jones protein in urine; Ibid., p. 255. — The glycyltryptophan 
and tryptophan tests for Cancer of the stomach (with C. H. San- 
ford) ; Ibid., p. 445. — A note on the distribution of chlorate in a woman 
fatally poisoned by potassium chlorate; Biochem. Bull., i, p. 483. — 
A study of the diffusibility of lipins from ether through rubber mem- 
branes into ether ; Ibid., ii, p. 64. — The colloidal nitrogen in urine from 
a dog with a tumor of the breast (with M. Kahn) ; Ibid., p. 87. — Effects 
of intraperitoneal injections of epinephrin on the partition of nitrogen 
in urine from a dog (with W. Weinberger) ; Ibid., p. 123. — A quanti- 
tative study of the lipins of bile obtained from a patient with a biliary 
fistula; Ibid., p. 182. — A disturbing factor in Lieben's and in Gunning's 
test for acetone in urine ; Jour. Amer. Med. Assn., lix, p. 445. — A report 
of some new chemical analyses of urinary calculi, with indications for 
treatment (with M. Kahn) ; Ibid., lix, p. 2252. — The diffusion of iodo- 
eosin from ether through rubber into ether ; Proc. See. Exp. Biol. and 
Med., X, p. 48. 

[Dr. Rosenbloom's papers in this issiie (pp. 22g, 2^^, 2^6, and 2<)Q) 
were submitted for publication in December 1912.] 

Awards of degrees at Columbia. -Mr. Anton R. Rose re- 
cently passed a public examination for the Ph.D. degree, thus com- 
pleting the requirements for that degree in biological chemistry. 
His dissertation is entitled Biochemical studies of phyto-phosphates. 
— Miss Clara W. Hasslock and D. F. Renshaw completed on Oct. 10 
the requirements for the degree of A.M. 

Miscellaneous items. Professor Gies delivered a lecture in 



328 Biochemical News, Notes and Comment [Jaa 

the autumn series at the New York Botanical Garden, October 26, 
on The chemical production of albuminous substances in plants. 
On October 7 he addressed the Section on Research of the First 
District Dental Society of the State of New York, at the Academy 
of Medicine, on Recent developments in the study of dental caries. 
Dr. Lothrop followed with a paper on the work he has been doing 
in this connection on salivary mucin.^ — Professor Gies was one of 
the Organizers of the New York Gastro-Enterological Society 
(P- 315)- He is Secretary of a Committee of Twenty-five, of Prof. 
R. H. Chittenden's pupils, in charge of a dinner to be given at Del- 
monico's on March i in honor of Professor Chittenden, and of a 
fund to be given to Yale in the name of Professor Chittenden. 

•Gies: Journal of the Allied (Dental) Societies, 1912, vii, pp. 397 and 478; 
Lothrop : Ibid., p. 410. 



EDITORIALS 

A year's experience in the conduct of the Biochemical Bulle- 
tin has induced us to change our plan of quarterly issue. The 
manuscript of future numbers will be sent to the printer on the first 

New plan of ^^Y ^^ ^^^^^ natural quarter; the months of issue 
quarterly issue of will be January, April, July and October; and 

the Bulletin ^^g contents of each issue will pertain to the quar- 
ter preceding the month of issue. Volume II will close with the 
July number. 

We congratulate President Taft and the country on the appoint- 
ment of Dr. Carl L. Aisberg, in succession to Dr. Harvey W. Wiley, 
as Chief of the Bureau of Chemistry. Dr. Alsberg's training in 

chemistry, in general biology and in medicine has 
been unusually broad and deep. His chemical 
knowledge, his sanitary comprehension, his scientific wisdom, and 
his zeal as an investigator, have had exceptional fruitage through- 
out his entire professional career. Admired as a gentleman by all 
who know him and respected by his colleagues everywhere as a sci- 
entist of eminent capacity, Dr. Aisberg is also universally esteemed 
for his habitual fidelity to duty, his moral integrity and his high 
professional purpose. We look forward with great confidence to 
a career for Dr. Aisberg which will be distinguished by a patriotism, 
a zeal in public Service, a personal courage, a common sense, a sci- 
entific exactness, an aggressiveness in the detection of violations of 
law, an executive capacity, that will be the delight of all his biochem- 
ical colleagues and the pride of his countrymen — and in this faith 
we tender him our felicitation and support. 

As we are about to close this issue of the Bulletin, we learn 
that the following testimonial (as proposed by Prof. Graham Lusk), 
which was sent a few days ago (Jan. 21) to about 275 of Dr. 
Alsberg's fellow workers in the American Physiological Society and 

329 



330 Biological Che mists in Hospitals [Jan. 

the American Society of Biological Chemists, has already been 
signed by nearly all of them : 

To Dr. Carl L. Aisberg: We who, like yourself, are active workers 
in the field of experimental biological science, congratulate the country 
and yourself on your appointment as Chief of the Bureau of Chemistry 
in the Department of Agriculture. We wish to express our confidence 
in your ability and integrity. We desire for you a successful admin- 
istration which shall promote the public welfare, shall jealously guard 
the public health, and shall uphold the dignity of the science which 
you represent. 

We are greatly indebted to Dr. Adler for the biographical and 
bibliographical facts, pertaining to Dr. Aisberg, on pages 211-216 
of this issue. 

During the past winter a number of cases of stock poisoning 

due, apparently, to the feeding of spoiled or moldy silage, were 

brought to our attention. At that time we were unable to give the 

Stock poisoning niatter due consideration. During the present 

due to spoiled si- winter, however, we shall be in a position to make 

läge. Help! some preliminary studies to ascertain the cause 
of toxicity in silage. Our work would be greatly facilitated if 
those readers of the Biochemical Bulletin who know of such 
cases of stock poisoning would bring this matter to the attention of 
the owners so that samples of the silage might be forwarded to the 
Chemical Section of the Iowa State College. The samples should 
be accompanied by füll particulars regarding the apparent cause of 
spoilage and the Symptoms exhibited by the animals to which the 
silage was fed. Arthur W. Dox. 



Until recently the pathological work in our larger hospitals has 

been done by attending physicians. The growth of this special field, 

however, has made it impossible for any one to keep abreast with 

Demand for bio- ^^ ^"^ ^^ anything eise. The progress of medical 

logical chemists in science constantly tends toward what is most re- 

the hospitals fined and subtle. As the microscope revealed 

new fields of research, so chemistry has opened previously unim- 



1913] Editoriais 33 ' 

agined paths for investigation. The directors of Hospitals are alive 
to this fact, as is shown by the increasing demand for biological 
chemists to cooperate with pathologists in the investigation of dis- 
ease. Up to now relatively few men have been adequately trained 
to be hospital chemists. What is required of them, and what will 
be demanded of them more and more, is not the making of routine 
analyses at the Suggestion of some attending physician, who in all 
probability has but a vague idea of what he wants; but rather that, 
unguided, they shall be able to discern in any disease process a defi- 
nite problem for investigation, and shall be competent to establish 
the conditions and conduct the details of suitable experimental re- 
search thereon. In order to do this with any degree of success, a 
hospital chemist must, in addition to his knowledge of biological 
chemistry, have a fairly good understanding of general pathology 
and bacteriology. It is not necessary that he be a physician, since 
technical familiarity with the clinical aspects of disease will not be 
reqnired of him. 

In the next decade, if not sooner, there will be a great demand 
for this type of highly trained biological chemist. The beginning 
of this demand is seen now in the growing number of biological 
chemists attached to the main hospitals of our larger cities. These 
chemists are on the same footing with the pathologists and the bac- 
teriologists. Although at present their remuneration is not what it 
should be, this will be remedied as soon as their value to hospitals 
is clearly shown. This field of work should be particularly attrac- 
tive to those who do not care for an academic career but who are 
devoted to biochemical research and averse to commercial chemistry. 



In the fall of 191 1, when it was seriously proposed to merge the 

American Society of Biological Chemists into the American Physio- 

logical Society, we were among the many who objected to the plan 

Federation of °^ ^^^ ground that such a merger would be detri- 

American Societies mental to biological chemistry as a science and 

for Experimental as a profession — sufficient reason for dissenters, 

10 ogy ^^^^ •£ ^ mej-ger were ordered, to maintain the 

existence of an independent American Biochemical Society.^ In 

^Editorial: Biochemical Bulletin, 1911, i, p. 364. 



332 Electrons [Jan. 

presenting these objections informally to our colleagues \ve empha- 
sized, however, the desirability of more intimate affiliation between 
the leading biological societies, in harmony zvith the policy of the 
American Society of Biological Chemists front its estahlishment, 
and suggested the Organization of a " federation" of independent 
societies for the attainment of that purpose and other mutually 
advantageous objects. There was much discussion but no decision. 

The Organization of the Biochemical Society of England, mean- 
while, with all that its existence implies,^ gave added weight to the 
objections that were raised against the assimilation of the Amer- 
ican Society of Biological Chemists by the American Physiological 
Society. 

During the past year the " federation " idea has won its way into 
unanimous acceptance, as is indicated by the account in this issue of 
the Organization of the Federation of American Societies for Ex- 
perimental Biology (page 269). The Federation is, in effect, an 
embryonic American Biological Society, the independent societies 
being its working divisions. The plan of federation has not weak- 
ened the independence or impaired the autonomy of any of the con- 
stituent societies. 

We heartily commend to the attention of all our readers the 
Mathews plan for the Organization of the American Biological So- 
ciety, which is published in füll in this issue (page 261 ). We believe 
that the logical development of the Federation would secure all the 
many desirable results at which Professor Mathews' excellent and 
far-reaching plan is aimed, including the establishment and success- 
ful conduct of a Biological Abstract Journal. We hope to present 
the views of some of our colleagues on this and related subjects in 
our July issue. 

The knowledge of nature as it is — not as we imagine it to be — 

constitutes true science. — Paracelsus. 

„ Liability to error is the price we pay for forward 

Electrons -^ o- , • i. 

movement. — o idgwick. 

The secret of all who make discoveries is to look upon nothing 

as impossible. — von Liehig. 

^Halliburton: Biochemical Bulletin, 1912, i, p. 484; ii, p. 128; 1913, ii (this 
issue), p. 318. 



BOOKS RECEIVED 

The BiocHEMicAL Bulletin will promptly acknowledge, under this heading, 
the receipt of all publications that may be presented to it. From time to time, 
selections will be made for review on pages of the volume to be appropriately 
indicated here. Reviews will be matter-of-fact Statements of the nature and 
Contents of the publications under consideration, and will be intended solely to 
guide possible ptirchasers. The wishes or expectations of publishers or donors 
of volumes will be disregarded, when they are incompatible with our convictions 
regarding the interests of our colleagues. The sises of the printed pages are 
indicated, in inches, in the appended notices. 

Glycosuria and allied conditions. By P. J. Cainmidge. Pp. 467 — 4 X 6.;4 ; 
$4.50 net. Longmans, Green & Co.. New York; Edward Arnold, London, 1913. 

The chemical Constitution of the proteins: Part II, Synthesis, etc. 26. ed. 
(One of the Monographs on Biochemistry.) By R. H. A. Plimmer, Univ. reader 
and ass't prof. of physiological ehem., University Coli., London. Pp. 107 — 4^ X 
7l/i ; $1.20 net. Longmans, Green & Co., 1913. 

Microscopy and the microscopical examination of drugs. By Chas. E. 
Gabel, microscopical food and drug analyst, Iowa State Dairy and Food Commis- 
sion. Pp. 116 — 4X6H', $1.00. Kenyon Co., Des Moines, la., 1911. 

Collected papers: Laboratory of physiological chemistry, Sheffield Sci- 
entific School, Yale University. 1911-1912. (35 reprints.) 

Medical and surgical report of Bellevue and Allied Hospitals in the City 
of New York. By Van Home Norrie, John A. Hartwell, A. Alexander Smith 
and Charles E. Nammack. Vol. iv, 1909-1910. (55 reprints.) 

Report of the laboratories of the University of BufFalo, medical depart- 
ment; including the third Harrington lecture (Hektoen). No. 4. 1912. (8 
reprints.) 

Contributions from the physiological laboratory of the Medico-Chirurgi- 
cal College, Phila. By Isaac Ott and John C. Scott. Part xix of Ott's con- 
tributions to physiology, 1912. (13 reprints.) 

Report of the Pellagra Commission of the State of Illinois. Pp. 250 — 4]4 
X 7. Nov., 1911. 

Practical physiological chemistry. A book designed for use in courses in 
practical physiological chemistry in schools of medicinc and of science. By 
Philip B. Hawk, professor of physiological chemistry and toxicology in the 
JeflFerson Medical College of Philadelphia. Fourth edition, revised and en- 
larged. Pp. 475—45^X8; $2.50 net. P. Blakiston's Sons & Co., Philadelphia, 
1912. 

The protein dement in nutrition. (One of the International Medical Mono- 
graphs.) By Major D. McCay, professor of physiology, Medical College, Cal- 
cutta. Pp. 216 — 4X7. with 8 füll page portraits of human subjects; $2.00 net. 
Longmans, Green and Co., New York; Edward Arnold, London, 1912. 

Oxidations and reductions in the animal body. (One of the Monographs 
on Biochemistry.) By H. D. Dakin, The Herter Laboratory, New York. Pp. 
135 — 4)/^X8; $1.40 net. Longmans, Green and Co., 1912. 

Researches on cellulose. III (1905-1910). By C. F. Gross and E. J. Bevan. 
Pp- U3 — 3/^X6; $2.50 net. Longmans, Green and Co., 1912. 

An introduction to the study of the protozoa, with special reference to 
the parasitic forms. By E. A. Minchin. professor of protozoology in the Univer- 
sity of London. Pp. Si/— 4X7l^; $600 net. Longmans, Green and Co., New 
York; Edward Arnold, London, 1912. 



OFFICERS OF THE BIOCHEMICAL DEPARTMENT OF 
COLUMBIA UNIVERSITY, 1912-1913* 

OFFICIAL REGISTER, DEC. 31, 1912 
William J. Gies: Professor and Chairman of the Stoff; Consulting chemisl, 

New York Botanical Garden; Pathological chemist, Bellevue Hospital; Mem- 

bcr of the Faculties of N. Y. Teachers College and N. Y. College of 

Pharmacy. [B.S., Gettysburg College, 1893 and M.S., 1896; Ph.B., Yale 

University, 1894 and Ph.D., 1897. Instructor, i898-'o2; adjunct professor, 

1902-05; Professor, 1905-.] 
Paul E. Howe: Assistant Professor. [B.S., University of Illinois, 1906; A.M., 

1907 and Ph.D., 1910. Assistant professor, 1912-.] 
Nellis B. Foster: Associate; Associate Physician, New York Hospital ; Chemist, 

St. Luke's Hospital. [B.S., Amherst College, 1898; M.D., Johns Hopkins 

University, 1902. Instructor, i9o6-'o8; associate, 1908-.] 
Walter H. Eddy : Associate and Secretary of the Staff. [B.S., Amherst Col- 
lege, 1898; A.M., Columbia, 1908 and Ph.D., 1909. Assistant, i9o8-'io; 

associate, 1910-.] 
Alfred P. Lothrop: Associate and Departmental Registrar. [A.B., Oberlin, 

1906 and A.M., 1907 ; Ph.D., Columbia, 1909. Assistant, i9o8-'o9 ; instructor, 

i909-'i2; associate, 1912-.] 
Herman O. Mosenthal : Associate; Assistant Attending Physician, Presbyterian 

Hospital; Assistant Physician, Vanderbilt Clinic; Instructor in medicine. 

[A.B., Columbia, 1899 and M.D., 1903. Assistant, i9o8-'09 ; instructor, i909-'i2 ; 

associate, 1912-.] 
Emily C. Seaman: Instructor. [B.S., Adelphi College, 1899; A.M., Columbia, 

1905 and Ph.D., 1912. Tutor, i90(>-'io; instructor, 1910-.] 
Max Kahn: Instructor. Director of the chemical and physiological laboratories 

of Beth Israel Hospital. [M.D., Cornell University Medical College, 1910; 

A.M., Columbia, 191 1 and Ph.D.. 1912. Instructor, 1912-.] 
Louis E. Wise: Instructor. [A.B., Columbia, 1907 and Ph.D., 191 1. Instructor, 

1912-.] 
Edgar G. Miller, Jr. : Assistant, 1911-. [B.S., Gettysburg College, 1911.] 
Frederic G. Goodridge: Assistant, 1912-. [A.B., Harvard University, 1897; 

M.D., Columbia, 1901.] 
Arthur Knudson: Assistant, 1912-. [A.B., University of Missouri, 1912.] 
Ethel Wickwire: Assistant, 1912-. [A.B., Tri-State College, 1909.] 
Tula L. Harkey: Assistant, 1912-. [A.B., Colorado College, 1909.] 
Benjamin Horowitz: Assistant, 1913-. [B.S., Columbia, 1911 and A.M., 1912.] 
Christian Seifert: Laboratory assistant, 1898-. 
Stella Waldeck : Recorder, 1908-. 

Blanche E. Shaffer: Laboratory assistant. summer Session, 1912. 
Joseph S. Hepburn : University fellow, 1912-13. [A.B., Central High School, 

Philadelphia, 1903 and A.M., 1908; B. S., University of Pennsylvania, 1907 

and M.S., 1907.] 

*The work of the department was inaugurated in October, 1898, by Prof. 
R. H. Chittenden (lecturer and director), Dr. William J. Gies (instructor), 
Messrs. Alfred N. Richards and Allan C. Eustis (assistants), and Christian 
Seifert (laboratory assistant). 



COURSES OFFERED BY THE BIOCHEMICAL DEPARTMENT OF 
COLUMBIA UNIVERSITY. 1911-1913 

Courses 51, 105 and 215 are given during the first half-year only. Course 
loi is given during the first half-year and is repeated (102) during the second 
half-year. Courses 104 and iio (52) are given only during the second half 
year. All other courses are conducted throughout the entirc academic year. 
All courses not otherwise specified are given at the College of Physicians and 
Surgeons. 

(Abbreviations: C, Conference; D, demonstration ; L, lecture; Lw, labora- 
tory work; R, recitation.) 

ORGANIC CHEMISTRY 
51. Elkmentary ORGANIC CHKMisTRY. Introductory to courses loi, 102 and 
HO (52). {Required of first year siudents of medicine.) L, i hr. D, i hr. 
R, 2 hr., each section (2). Lw. 6 hr. each section (2). Profs. Gies and Howe, 
Drs. Wise and Goodridge, and Messrs. Miller and Knudson. 

NUTRITION (PHYSIOLOGICAL AND PATHOLOGICAL CHEMISTRY) 

101-102. General physiological chemistry. A course in the eletnents of 
normal nutrition. (Teachers College, School of Practical Arts.) L, 2 hr. R. l 
hr., each section (2). Lw, 5 hr., each section (2). Prof. Gies, Dr. Seaman and 
Misses VVickwire and Harkey. (This course is designated "Chemistry 51" and 
"Household Arts Education 125" in the Teachers College Announcement.) 

This course is designated "Chemistry s 51 " in the Teachers College Division 
of the Summer School Announcement. The course was given last summer by 
Prof. Gies, Dr. Seaman and Miss Shaffer. 

104. General patholooical chemistry. Lectures on nutrition in disease. 
(Teachers College, School of Practical Arts.) L, i hr. Prof. Gies. (This 
course is designated "Chemistry 52" in the Teachers College Announcement.) 

HO (52). General physiological chemistry. A course in the Clements of 
normal nutrition. (Required of first year students of medicine.) L, 2 hr. R. i 
hr.. each section (2). Lw, 6 hr., each section (2). Profs. Gies and Howe. 
Dr. Wise, and Messrs. Miller and Knudson. 

This course is designated "S — 104" in the Medical Division of the Summer 
School Announcement. It was given last summer by Prof. Gies and Dr. Smith. 

209-210. Chemistry of nutrition. (School of Pharmacy. Required of 
candidates for the Degree of Doctor of Pharmacy.") L, 1 hr. Prof. Gies. 

211-212. General biological chemistry. Specially adopted to the needs 
of secondary school teachers of biology. L, i hr. Lw, 4 hr. Dr. Eddy. 

213-214. Advanced physiological chemistry, including methods of re- 
SEARCH IN nutrition. (Teachers College, School of Practical Arts.) L, i hr. 
Lw, 5 hr. Prof. Howe, Dr. Seaman and Mr. Horowitz. (This course is desig- 
nated " Household Arts Education 127" in the Teachers College Announcement.) 

215. General biological chemistry. A course in the eletnents of normal 
nutrition. L, i hr. Lw, 7 hr. Prof. Gies, Dr. Lothrop and Messrs. Miller and 
Knudson. 

217-218. BlOCHEMICAL methods OF RESEARCH, INCLUDING CLINICAL METHODS 

AND URiNARY ANALYsis IN GENERAL. L, I hr. Lw, 7 hr. Profs. Gies and Howe, 
Dr. Lothrop, and Messrs. Miller and Hepburn. 

219-220. Nutrition in health. A laboratory course in advanced physio- 
logical chemistry. L, 2 hr. Lw, 14 hr. Profs. Gies and Howe, and Dr. Lothrop. 



Courses in Nutrition (continued) 

221-222. Nutrition in Disease. A lahoratory course in advanced patholog- 
ical chemistry. L, 2 hr. Lw, 14 hr. Prof. Gies. 

223-224. Nutrition in Disease. L, i hr. Profs. Gies and Howe, and Drs. 
Foster, Mosenthal, Kahn and Goodridgc. 

225-226. Advanced physiological and patholocical chemistry, including 
ALL PHASES OF NUTRITION. Research. C, I hr. (individual students). Lw, 16 hr. 
Profs. Gies and Howe, and Dr. Lothrop. 

fm^COLOGY 
231-232. Effects and detection oi«*poi.sons, including food preservatives 
and adulterants. Lw, 6 hr. Prof. Gies and Mr. Miller. 

BOTANY 
235-236. Chemical piiYsiOLOGY of PLANTS. (New York Botanical Garden.) 
L, 1 hr. Lw, 5 hr. Prof. Gies. 

BACTERIOLOGY 
241-242. Chemistry of microorganisms: fermentations. putrefactions 
AND THE behavior of enzymes. An introducHon io sauitary chemistry. L, i hr. 
Lw, 7 hr. Prof. Gies. 

SANITATION 
105. Sanitary chemistry. (Teachers College, School of Practical Arts). 
L. I hr. Lw, 3 hr. Dr. Seaman and Miss Harkey. (This course is designated 
" Chemistry 57 " and " Household Arts Education 129 " in the Teachers College 
Announcement. ) 

BIOCHEMICAL SEMINAR 
301-302. Biochemical Seminar, i hr. Prof. Gies. 

RESEARCH IN BIOLOGICAL CHEMISTRY 
Biochemical research may be conducted, by advanced workers, independently 
or under guidancc, in any of the departmcntal laboratories. 

LABORATORIES FOR ADVANCED WORK IN BIOCHEMISTRY 
The laboratories in which the advanced work of the biochemical department 
is conducted are situated at the College of Physicians and Surgeons, Teachers 
College, New York Botanical Garden and Bellevue Hospital. Fach lahoratory 
is well equipped for research in nutrition and all other phases of biological 
chemistry. 

BIOCHEMICAL LIBRARY 
Prof. Gies* library occupies a room adjoining the main biochemical lahora- 
tory at the College of Physicians and Surgeons and is accessible, by appoint- 
ment, to all past and present workers in the Department. 

COLUMBIA UNIVERSITY BIOCHEMICAL ASSOCIATION 

The Biochemical Association holds scientific meetings regularly on tiie first 
Fridays in December, February and April, and on the first Monday in June. 
These meetings are open to all students in the University. 

SUMMER SCHOOL COURSES 
Summer Session courses are mentioned in the foregoing references to 
Courses 101-102 and iio (52). Prof. Gies will have charge of both courses next 
Summer. He will also conduct a special lecture course in nutrition. The labora- 
tories will be open for research throughout the summer. 



ANNOUNCEMENTS 
Professional Assistance Offered to Biological Chemists 
The Columbia University Biochemical Association will be glad to 
cooperate confidentially with all who desire the Services of biological 
chemists and with all who seek positions in biological chemistry. 
Address inquiries to William J. Gies, 457 West sgth St., New York. 

Joumalistic 

New JOURNAL. Physiological Researches. To appear at irregulär 
intervals. Edited by Burton E. Livingston, Manager, Johns Hopkins 
University ; jDamV/ T. MacDoiigal, Carnegie Institution of Washington ; 
Herbert M. Richards, Columbia University. " The recent rapid advance 
of physiological science has been accompanied by a realization of the 
Community of interest and uniformity of method which characterize the 
physiology of plants and of animals, and it has seemed highly desirable 
that the general physiological field thus indicated should possess an 
Organ of publication in which its more comprehensive and technical 
papers might appear. This need is emphasized by the fact that pres- 
ent facilities for publication in physiology are generally taxed beyond 
their capacity and papers are consequently subject to long delays in 
appearance. It has therefore been decided to inaugurate a new series 
of scientific papers which will embrace contributions towards the 
advance of fundamental physiological knowledge." 

" The plan of publication of the new series, for which the title of 
Physiological Researches has been adopted, is one in which practical 
ownership is vested in the contributors. It is hoped that the project 
will receive the interest and support of biologists of all classes. (Each 
volume will contain about 450 pages; each number will contain but a 
Single contribution ; and the numbers will be issued irregularly). Pub- 
lication of the first contribution ma)'^ be expected in a short time. Sub- 
scriptions will be received by the volume, the price being $5.00 per 
volume, payable in advance. Subscriptions to volume I, which are 
received prior to the date of publication of the first research, may be 
made at the reduced price of S4.00. At the date of the appearance of 
the first research the price will automatically become the regulär one. 
Remittances should be made payable to Physiological Researches, and 
all correspondence should be addressed to Physiological Researches, 
Station N, Baltimore, Maryland, U. S. A." (Editors' announcement.) 

Reduced subscription price of the Journal of Biological 
Chemistry, The directors of the Journal of Biological Chemistry have 
announced that "beginning with the February issue of 1913 (Vol. 14, 
No. i) the subscription price of the Journal to domestic subscribers 
will be reduced f rom $4.00 to $3.00 per volume ; to foreign subscribers, 
$3.25. Any one engaged in biochemical work who subscribes for the 
Journal at this rate (beginning with Vol. 14) may secure Volumes 1-13 
for $20.00, plus cost of transportation. The price at which a complete 
set has hitherto been sold is $50.00. Subscribers for the Journal who 
wish to complete their files may secure early volumes for $1.50 each, 
plus cost of transportation. Address : Alfred N. Richards, Secretary, 
University of Pennsylvania. 



Meetings of Societies and Congresses 

A-MERicAN Chemical Society: Annual meeting (47th) at Mil- 
waukee, Wisconsin, March 25-28. Charles L. Parsons, Secretary, Box 
505, Washington, D. C. At the last meeting of the Society the Council 
authorized the formation of a Div^ision of Biological Chemistry. At 
that meeting the details of Organization of the Division were entrusted 
to a committee. The committee will report in Milwaukee, the final 
Organization of the Division will be perfected, and officers will be 
elected. 

Tenth International Congress of Agriculture: Ghent, Bel- 
gium, June 8-13. Secretary-general, Dr. P. de Vuyst, 22 Avenue des 
Germaines, Brüssels. American committee: Dr. L. O. Howard, mem- 
ber of the International Commission on Agriculture and chief of the 
Bureau of Entomolog>'; and Dr. A. C. True, director, Mr. John Ham- 
ilton, specialist in farmers' institutes, Dr. C. F. Langworthy, chief of 
nutrition investigations and Dr. J. I. Schulte, assistant agriculturist, of 
the Office of Experiment Stations. 

American Medical Association, Annual meeting: Minneapolis, 
Minn., Jiine 17-20. General secretary, Geo. H. Simmons, 535 Dear- 
born Ave., Chicago. 

General meeting of the International Association of Botan- 
iSTs: Copenhagen, June 23. Secretary-general, J. P. Lotsy, Haarlem, 
Holland. 

Seventeenth International Congress of Medicine: London, 
Aug. 6-12. General secretary, Dr. W. P. Herringham, 13 Hinde St., 
London, W. 

FouRTH International Congress on School Hygiene: Buflfalo, 
N. Y., Aug. 25-7,0. Secretary-general, Prof. Thomas A. Storey, Col- 
lege of the City of New York. 

NiNTH International Physiological Congress: Groningen, Hol- 
land, Sept. 2-6. American Secretary, Prof. W. T. Porter, Harvard 
Medical School. 

Third International Congress of Refrigeration : Washington, 
D. C, Sept. if) (opening meeting) ; Chicago, Sept. 17-23 (business 
and scientific meetings). Secretary-general, Mr. J. F. Nickerson, 431 
So. Dearborn St., Chicago. 

The Biochemical Bulletin 

The Biochemical Bulletin is a quarterly biochemical review. 
It publishes results of original investigations in biological chemistry 
and presents miscellaneous items of personal and professional in- 
terest to chemical biologists. Original contributions to research, 
preliminary reports of investigations, abstracts of papers, addresses. 
reviews, descriptions of new methods and apparatus, practical sug- 
gestions to teachers, biographical notes, historical summaries, 
bibliographies, quotations, news items, proceedings of societies, 
personalia, views on current events in chemical biology, etc., are 
solicited. 

Suhscription priccs. Vol. I : $6.00 (No, i, $1.50 ; No. 2, $2.50 ; 
No. 3, v$2.oo; No. 4, $1.50). Vol. II: $2.75 (domestic) ; $3.00 
(foreign) ; $6.00 aftcr July i, igi^ (No. 5, $1.25 ; No. 6, $1.00). 

Remittances, manuscripts and correspondence should be addressed 
to the Biochemical Bulletin, 437 West 59th St., New York City. 



Vol. II April, 1913 No. 7 

Biochemical Bulletin 

Edited, for the Columbia University Biochemical Association, by the 

EDITORIAL COMMITTEE: 

ALFRED P. LOTHROP, Chairman, 

PAUL E. HOWE, Secretary, WILLIAM J. GIES, Treasurer, 

WALTER H. EDDY, JOSEPH S. HEPBURN, H. O. MOSENTHAL. 

NELLIS B. FOSTER, MAX KAHN, EMILY C. SEAMAN, 

F. G. GOODRIDGE, ARTHUR KNUDSON, ETHEL WICKWIRE, 

TULA L. HARKEY, EDGAR G. MILLER, JR., LOUIS E. WISE, 

all of the Staff of the Biochemical Department of Columbia University. 



CONTENTS 

Heinrich Ritthausen (Portrait) 334 

Appreciation. Thomas B. Osborne ; 335 

BiBLiOGRAPHY. Lewis IV. Fetzer. 339 

Dinner To Professor Chittenden : Testimonial by his pupils, '94S. 349 

Society for Experimental Biology and Medicine : Tenth anniversary meeting 

and dinner. Nineteen O. Three 358 

Methods for the Electrometric Determination of the Concentration of 

Hydrogen IONS IN BiOLOGiCAL FLUIDS. K. A. Hasselbalch 367 

A Method for the Determination of Tryptophan derived from Protein. 

Jesse A. Sanders and Clarence E. May 373 

Physical Chemistry of Muscle Plasma. Filippo Bottazzi '. 379 

Fasting Studies : h. A Note on the Composition of Muscle from Fasting Dogs. 

H. C. Biddle and Paul E. Howe 386 
Some Notes on the Form of the Curve of Carbon-dioxide Excretion Result- 

iNG FROM Muscular Work Following Forced Breathing. G. O. Higley... 390 
The Influence of Barometric Pressure on Carbon-Dioxide Excretion in Man. 

G. O. Higley 393 
The Relation of Acapnia to Shock, and a Consideration of the Mechanical 

Effects of Artificial Hyper-Respiration upon the Circulation. 

Henry H. Janeway and Ephraim M. Ewing 403 
Cleavage of Pyromucuric Acid by Mold Enzymes. 

Arthur W. Dox and Ray E. Neidig. 407 

Analysis of the Ash of the Castor Bean. Marston Lovell Hamlin 410 

Notes on the Chemical« Nature of the " Tannin Masses " in the Fruit of 

the Persimmon. Ernest D. Clark 412 

HiSTON AND ITS Preparation. Walter H. Eddy 419 

Did von Wittich Antedate Ostwald in the Definition of Enzyme Action ? 

William N. Berg 441 

The Biochemical Society, England 446 

Scientific Proceedings of the Columbia University Biochemical Association. 

Alfred P. Lothrop, Secretary 452 

Biochemical Bibliography and Index. William J. Gies 470 

Biochemical News, Notes and Comment 476 

Editorials : Including numerous quotations from letters on the Mathews plan for 

the Organization of an American Biological Society 487 



NEW YORK 

Columbia University Biochemical Association. 

Entered as second-claas matter in the Post Office at Lancaster, Pa. 



MEMBERS OF THE COLUMBIA UNIVERSITY 
BIOCHEMICAL ASSOCIATION 

Honorary Members 

PROF. R. H. CHITTENDEN, First Director of the Columbia University De- 
partment of Biological (Physiological) Chemistry; Director of the Shef' 
field Scientific School of Yale University 

PROF. HUGO KRONECKER, Director of the Physiological Institute, Uni- 
versity of Bern, Switserland 

PROF. SAMUEL W. LAMBERT, Dean of the Columbia University School of 
Mediane 

DR. JACQUES LOEB, Member of the Rockef eller Institute for Medical Re- 
search; Head of the Department of Ex perimental Biology 

PROF. ALEXANDER SMITH, Head of the Department of Chemistry, Co- 
lumbia University 

Corresponding Members 

PROF. LEON ASHER, University of Bern, Switserland 

PROF. FILIPPO BOTTAZZI, University of Naples, Italy 

PROF. VLADIMIR S. GULEVIC, University of Moscow, Russia 

PROF. W. D. HALLIBURTON, King's College, London 

PROF. S. G. HEDIN, University of Upsala, Sweden 

PROF. FREDERICO LANDOLPH, University of La Plata, Argentina 

PROF. A. B. MACALLUM, University of Toronto, Canada 

PROF. C. A. PEKELHARING, University of Utrecht, Holland 

PROF. S. P. L. SÖRENSEN, Carlsberg Laboratory, Copenhagen, Denmark 

Members Resident in New York City 

Brooklyn Botanic Garden, — C. Stuart Gager. 

College of the City of New York. — ^Wm. B. Boyd, Louis J. Curtman, 
Benj. G. Feinberg, A. J. Goldfarb. 

Columbia University: Departments. — Anatomy: Alfred J. Brown, H. von 
W. Schulte; Bacteriology: James G. Dwyer; Biological Chemistry: Walter H. 
Eddy, Nellis B. Foster, William J. Gies, F. G. Goodridge, Tula L. Harkey, 
Joseph S. Hepbum, Benjamin Horowitz, Paul E. Howe, Max Kahn, Arthur 
Knudson, Alfred P. Lothrop, Edgar G. Miller, Jr., H, O. Mosenthal, Emily C. 
Seaman, Chris Seifert, Ethel Wickwire, Louis E. Wise; Botany: E. R. Alten- 
burg, C. A. Darling; Cancer Research: William H. Woglom; Chemistry: A. M. 
Buswell, R. P. Calvert, Gustave Egloff, Harry L. Fisher, Percy W. Punnett, 
A. W. Thomas; Clinical Pathology: Edward Cussler, Peter Irving; Diseases of 
Children: Herbert B. Wilcox; Gynecology: Wilbur Ward; Mediane: T. Stuart 
Hart, I. Ogden Woodruff; Pathology: B. S. Oppenheimer, Alwin M. Pappen- 
heimer; Pharmacology: Charles C. Lieb; Physiology: Russell Burton-Opitz, 
Donald Gordon, Leander H. Shearer, Wm. K. Terriberry; Surgery: Hugh 
Auchincloss, William Darrach, Rolfe Kingsley, Adrian V. S. Lambert, F. T. 



Members resident in New York (con.) 

Van Buren, Jr. ; Therapeutics: Maximilian Schulman; University Physician: 
Wm, H. McCastline; Vanderhilt Clinic: F. Morris Class, Julius W. Weinstein; 
Zoology: John D. Haseman, H, J. Muller, Charles Packard. 

Colleges. — Barnard: Helene M, Boas, Ella H. Clark, Ruth S. Finch, Louise 
H. Gregory; College of Pharmacy: Charles W. Ballard; Teachers College: 
Mary G. McCormick, Mrs. A. P. McGowan, Sadie B. Vanderhilt. 

Students. — Graduate: Cora J. Beckwith, Sidney Born, O. C. Bowes, Helen 
B. Davis, Mary C. de Garmo, Frank R. Eider, Louis J. Hirschleifer, Shojiro 
Kubushiro, Victor E. Levine, Darwin O. Lyon, W. A. Perlzweig, Edward Plaut, 
Geo. S. Rosenthal, Edward C. Stone, Jennie A. Walker, Charles Weisman, C. A. 
Wells, Isabel Wheeler. — Teachers College: Anna M. Connelly, Ula M. Dow, 
Ada M. Field, Helen McClure, Alice H. McKinney, Elizabeth Rothermel, Mary 
B. Stark, Helen B. Thompson. — Medical: Louis Berman, Ernst Boas, David C. 
Bull, Will H. Chapman, Robert T. Corry, Calvin B. Coulter, Joseph Felsen, 
Joseph Goldstone, Julius Gottesman, Leon M. Herbert, Martin Holzman, Walter 
F. Hume, Julius Hyman, M. V. Miller, Nathan Rosenthal, A. V. Salomon, Harry 
J. Seiff, Jacob Shulansky, H. J. Spencer, Henry A. Sussman, Wm. W. Tracey, 
Grover Tracy. 

CoRNELL University Medical College. — Stanley R. Benedict, Ernest D. 
Clark, Robert A. Cooke, Jessie A. Moore, Charles R. Stockard, Geo. W. 
Vandegrift. 

EcLECTic Medical College. — David Alperin. 

Harriman Research Laboratory. — Marston L. Hamlin. 

Hospitals. — Babies': Morris Stark; Bellevue: Edward C. Brenner, Edward 
M. CoHe, Jr., Ralph W. Lobenstine; City: Henry H. Janeway, Louis Pine; Flush- 
ing: Eimer W. Baker; General Memorial: Clinton B. Knapp; German: H. G. 
Baumgard, Alfred M. Hellman, Melvin G. Herzfeld, Frederick B. Humph'cies, 
Charles H. Sanford, Fred S. Weingarten; Jewish: Abraham Ravich ; Lebanon: 
Samuel Gitlow, M. J. Gottlieb, William Weinberger; Lutheran: Daniel R. Lucas; 
Mt. Sinai: George Baehr, Samuel Bookman, Leo Buerger, Burrill B. Crohn, 
Simon S. Friedman, David J. Kaliski, John L. Kantor, Leo Kessel, Reuben Otten- 
berg, Harry Wessler; N. Y.: James C. Greenway, Ralph G. Stillman; N. Y. 
Nursery and Child's: Oscar M. Schloss; Presbyterian: Herbert S. Carter, Russell 
L. Cecil, Arthur W. Swann; Roosevelt: J. Buren Sidbury; St. Ltike's: Norman 
E. Ditman, Edward C. Kendall, W. S. Schley, Chas. H. Smith. 

Long Island Medical College. — Matthew Steel. 

Montefiore Home. — Isidor Greenwald. 

Museum of Natural History. — Louis Hussakof, Israel J. Kligler. 

N. Y. Aquarium. — Raymond C. Osburn. 

N. Y. Association for Improving the Condition of the Poor. — Donald B. 
Armstrong. 

N. Y. Botanical Garden. — Fred J. Seaver. 

N. Y. City Department of Education. — Boys' High School: Frank T. 
Hughes; Brooklyn Training School: ehester A. Mathewson; Commercial High 
School: Walter J. Donvan, Benj. C. Gruenberg; DeWitt Clinton High School: 
Frank M. Wheat; Rastern District High School: Gertrude S. Burlingham; 
Girls' High School: Marguerite T. Lee; High School of Commerce: Harvey B. 
Clough, Fred W. Hartwell; Jamaica High School: Ella A. Holmes, Charles H. 
Vosburgh; Manual Training High School: Anna Everson; Morris High School: 



Members resident in New York (con.) 

Charles A. Wirth; Ncwtozvii High School: Nellic P. Hewins; Wadleigh High 
School: Helen Gavin, Elsie A. Kupfer, Helen G. Russell, Helen S. Watt. 

N. Y. City Department of Health. — Charles F. Bolduan, Alfred F. Hess. 

N. Y. City Normal College. — Beatrix H. Gross. 

N. Y. Eye and Ear Infirmary. — Harold M. Hays. 

N. Y. Milk Committee. — Philip Van Ingen. 

N. Y. PoLYCLiNic Medical School. — Jesse G. M. Bullowa, Mabel C. Little. 

Post Graduate Medical School. — Louis E. Bisch, Arthur F. Chace. 

Pratt Institute. — Grace MacLeod. 

RocKEFELLER INSTITUTE. — Jacob Bronfcnbrcnner, Alfred E. Cohn, George 
\V. Draper, Frederic M. Hanes, Michael Heidelberger, Gustave M. Meyer. 

Russell Sage Institute of Pathology. — Eugene F. DuBois. 

TuRCK Institute. — Anton R. Rose. 

Vettin School. — Laura l. Mattoon. 

Leopold L. Falke, 5316 Thirteenth Avenue, Brooklyn ; Mahel P. Fitzgerald, 
416 East 65th Street, Manhattan; Abraham Gross, c/o Arbuckle Sugar Co., 
Brooklyn; Alfred H. Kropff, 619 Kent Avenue, Brooklyn. 

Non-Resident Members 

Allegheny General Hospital (Pittsburgh). — James P. McKelvy. 

Carnegie Institution (Cold Spring Harbor, L. I.). — Ross A. Gortner. 

Cornell University (Ithaca). — Jean Broadhurst. 

Drake University Medical School (Des Moines, la.). — E. R. Posner. 

Forest School (Biltmore, N. C). — Homer D. House. 

Iowa University Hospital (Iowa City). — Louis Baumann. 

Isolation Hospital (San Francisco, Cal.). — L. D. Mead. 

Jefferson Medical College (Phila.). — P. B. Hawk, Edward A. Spitzka. 

Johns Hopkins University (Baltimore). — John Howland, W. M. Kraus, 
Burton E. Livingston, Edwards A. Park. 

Lehigh University (Bethlehem, Pa.). — William H. Welker. 

Leland Stanford University (Palo Alto, Cal.). — Hans Zinsser. 

MacDonald College (Quebec). — Kathryn Fisher. 

Mass. Agricultural College (Amherst). — H. D. Goodale. 

New Mexico Agricultural College (State College). — R. F. Hare. 

N. J. Agricultural Experiment Station (New Brunswick). — Carl A. 
Schwarze. 

Ohio Agricultural Experiment Station (Wooster). — A. D. Selby. 

Princeton University (N. J.). — E. Newton Harvey. 

Psychopathic Hospital (Boston). — Herman M. Adler. 

Rensselaer Polytechnic Institute (Troy, N. Y.). — Fred W. Schwartz. 

Secondary Schools. — Brock Port State Normal School (N. Y.) : Ida C. Wads- 
worth; Hermoit High School (N. Y.) : Sidney Liebovitz; Indiana State Normal 
School (Terre Haute): Roscoe R. Hyde; Ingleside School (New Milford, 
Conn.) : Mary L. Chase; Knox School (Tarrytown, N. Y.) : Clara G. Miller; 
New Bedford Indiistrial School (Mass.): Constance C. Hart; North Texas 
State Normal School (Benton) : Blanche E. Shaffer; Passaic High School 
(N. J.) : Hazel Donham, Helene M. Pope; Rochester High School (N. Y.) : 
David F. Renshaw; State Normal School (Truro, N. S.) : Blanche E. Harris. 



Non-resident members (con.) 

Trinity College (Hartford, Conn.). — Max Morse, R. M. Yergason. 

TuLANE University (Ncw Orleans, La.). — Allan C. Eustis. 

U. S. Department of Agriculture. — Carl L. Alsbcrg, W. N. Berg, H. E. 
Buchbinder, William Salant, Clayton S. Smith. 

U. S. Food and Drug Inspection Laboratory (Phila.). — Harold E. Wood- 
ward. 

University of Alabama Medical School (Birmingham). — Richard A. Bliss. 

University of California (Berkeley). — William T. Hörne. 

University of Chicago. — Mathilde Koch. 

University of Georgia Medical School (Atlanta). — William D. Cutter. 

University of Illinois (Urbana). — George D. Beal, Isabel Bevicr, A. D. 
Emmett. 

University of Indiana (Bloomington). — Clarence E. May. 

University of Kentucky (Louisville). — Mary E. Swecny. 

University of Michigan (Ann Arbor). — A. Franklin Shull. 

University of Montana (Missoula). — J. E. Kirkwood. 

University of Pennsylvania (Phila.). — A. N. Richards. 

University of Porto Rico (Las Pietras). — L. A. Robinson. 

University of Tennessee (Memphis). — Edwin D. Watkins. 

University of Texas (Austin). — Mary E. Gearing, Anna E. Richardson. 

University of Toronto (Canada). — Olive G. Pattcrson, 

University of Utah (Salt Lake City). — H. A. Mattill. 

University of Wisconsin (Aladison). — W. H. Petersen. 

Vassar College (Poughkeepsie, N. Y.). — Winifred J. Robinson. 

Washington State College (PuUman). — ^Joscphine T. Berry, Louise 
McDanell. 

Wesleyan University (Middletown, Conn.). — David D. Whitney. 

West Penn Hospital (Pittsburgh). — Jacob Rosenboom. 

Williams College (Williamstown, Mass.). — John S. Adriance. 

Yale University (New Haven, Conn.). — Lorande Loss Woodruff. 

Albert H. Allen, Saranac Lake, N. Y. ; Emma A. Buehler, Newark, N. J.; 
George A. Geiger, West Orange, N. J.; Edward G. Griffin, Albany, N. Y. ; F. C. 
Hinkel, Utica, N. Y.; Cavalier H. Joüet, Roselle, N. J.; A. E. Olpt>, West 
Hoboken, N. J.; Adeline H. Rowland, Pittsburgh, Pa. ; William A. Taltavall, 
Redlands, Cal. ; David C. Twichell, Saranac Lake, N. Y. 



EDITORS OF THE BIOCHEMICAL BULLETIN 

The editorial committee 
with the collaboration of the members and the 

SPECIAL CONTRIBUTORS: 

DR. JOHN AUER, Rockefeiler Institute for Medical Research 

PROF. WILDER D. BANCROFT, Cornell University,- Ithaca 

DR. CHARLES A. DOREMUS, 55 W. 53d St., New York City 

DR. ARTHUR W. DOX, Iowa State College Agric. Experiment Station, Arnes 

PROF. JOSEPH ERLANGER, Washington Univ. Medical School, St. Louis 

DR. LEWIS W. FETZER, U. S. Dep't of Agriculture, Washington, D. C. 

PROF. MARTIN H. FISCHER, University of Cincinnati 

DR. MARY LOUISE FOSTER, Smith College, Northampton, Mass. 

DR. V. J. HARDING, McGill University, Montreal, Canada 

DR. R. H. M. HARDISTY, McGill University, Montreal, Canada 

DR. K. A. HASSELBALCH, Finsen Institute, Copenhagen, Denmark 

PROF. G. O. HIGLEY, Ohio Wesleyan University, Delaware 

DR. VERNON K. KRIEBLE, McGill University, Montreal, Canada 

PROF. FRANCIS E. LLOYD, McGill University, Montreal, Canada 

PROF. JOHN A. MANDEL, A^. Y. Univ. and Bellevue Hospital Med. College 

PROF. ALBERT P. MATHEWS, University of Chicago 

PROF. SHINNOSUKE MATSUNAGA, University of Tokyo, Japan 

PROF. LAFAYETTE B. MENDEL, Yale University 

PROF. VICTOR C. MYERS, N. Y. Post-Graduate Med. School and Hospital 

DR. THOMAS B. OSBORNE, Conn. Agric. Experiment Station, New Haven 

DR. AMOS W. PETERS, The Training School, Vineland, N. J. 

PROF. R. F. RUTTAN, McGill University, Montreal, Canada 

DR. E. E. SMITH, 50 East 4ist St., New York City 

DR. A. E. SPAAR, City Hospital, Trincomalee, Ceylon 

PROF. UMETARÖ SUZUKI, University of Tokyo, Japan 

MISS ANNA W. WILLIAMS, University of Illinois, Urbana, lll. 

PROF. E. WINTERSTEIN, Polytechnic Institute, Zürich, Switzerland 

DR. JULES WOLFF, 26 Rue Dutot, Paris 




HEINRICH RITTHAUSEN. 



BiocHEMiCAL Bulletin 



Volume II APRIL, 191 3 No. 7 



IN MEMORIAM 

HEINRICH RITTHAUSEN 
Born January 13, 1826 Died October 16, 1912 

Bv the death of Heinrich Ritthansen in Berlin, on October 16, 
1912, at the age of eighty-six, a long life spent in biochemical 
research was terminated. Beginning as a Student under Liebig, and 
inspired by this great teacher, he niade agricnltural chemistry bis 
life profession. His first work was as assistant to Professor Erd- 
mann at Leipzig. From 1854 to 1856 he was director of the scien- 
tific department of the experiment Station at Moeckern. He theo 
became director of the Station at Saaran in Schlesien. In 1858 he 
was appointed professor of chemistry and physics in the Royal 
Agricnltural Academy at ^^"aldau ; in 1867, professor of chemistry 
and director of the experiment Station at Poppeisdorf; and in 1873, 
professor of chemistry at Königsberg, where he remained until 
1899. when his active career was concluded by advancing years. 
The latter part of his life was spent in Berlin. 

At the present time, when the development of agricnltural sci- 
ence has made piain the value of the agricnltural experiment Station 
not only to the f armer jjut to the entire Community, the life of one 
who commenced his career in the first established institution of this 
kind is of special interest. 

A\'hen Ritthausen began his work, the brilliant writings and 
lectures of Liebig had directed the attention of the whole world to 
the importance of applying the discoveries of science to the practice 

't -1 - 



3,3^ ^Ic'mricli l\inh(nisrn lApril 

of ai;ricu]lui"c. As i»nly ihe nidinicMils of a kiiowlcdge of llie 
cliemical and ])hysical ])r()l)lenis of ihe i^rowlh and niaintenance of 
plants and animals liad l)een ac(|nire(l, it seemed a sim])le matter to 
instrucl ihe farnier in proper melhods to l)e eniployed in raising liis 
crops and stock. No dou1)t remained in the minds of the early 
scientists \\\\t) promoted this propaganda that ihe practical retiirns 
of their efforts wonld soon l)e reahzed. 

Planis were to l)e fed with carbonic acid, nitrogen, and inorganic 
salts, and ihe proper (jnaniity of each essential dement of plant 
food was to l)e determined l)}- chemical analysis of the tissnes and 
ash of the plant, the fertilizer snpplied and the soil in w hich it grew. 

Animals were supposed to be composed of substances directly or 
indirecll}" assimilated from their foods. Their heat and mechanical 
energy was snpplied by the carljohydrates and fats ; the all)nmin, 
fibrin. casein and gelatin ])resent in their Ijlood, mnscle, milk, etc., 
\vas fnrnished !)}■ identical proteins contained in the \arions vege- 
table ])r()dncts \\\{h wliich they were fed. Here again chemical 
analysis was to fnrnish the gnide to proper practice, which shonld 
replace crude and wastefnl methods fonnded in ignorance of the 
real factors involved. 

I.ittle did the enthusiasts who established the first agricultural 
experiment Station realize what was before them nor what dis- 
coxeries in e\'ery Ijranch of l)ii)logical science their efforts were to 
lead to. Little did they dream of enzymes and liydrolyses, of 
toxins and antitoxins and specificity of li\-ing tissues, of optical 
isomers and tantomeric componnds, of nncleic acids and pyrimidines, 
of amino-acids and Polypeptides, of colloids and surface tension, 
nor of mnltitndes of other discoveries ^vhic]^ have followed chiefly 
from the inspiration which Liebig imparted to those who worked 
Avitli liim. 

All tliese disco\-eries had to ])e made l^efore the practical jjrob- 
lems of agricnltnre conld be satisfactorily dealt with by the scientist ; 
and it is evident that Ritthausen was one of tlie first to realize this, 
for we find him, after a short experience in attempting an immediate 
application of chemistry to the feeding of cattle, tnrning bis atten- 
tion to a carefnl study of the protein constituents of their vegetable 
foods. 



1913I Thomas B. Osbonie 337 

His earliest paper on this subject, which appeared in the Jonnial 
für praktische Chemie in 1862, described the proteins of wheat ; 
and for five successive years he pnblished papers on this same sub- 
ject. In the conrse of this work he isolated glutaminic acid from 
the products of hydrolysis of the gluten proteins, a discovery which 
ranks among the more important made by biochemists. He then 
extended his investigations to seeds of importance for nutrition, 
hardlv a year passing when he did not contribute two or more papers 
on the resnlts of his work. 

In 1872 he pnl)hshed a review of his earher work nnder the 
title Die Eiz^'cisskörpcr der Getreidearten. HiUseufrüeJiteii und 
Oelsamen. This was the first attempt made to furnish an account 
of wliat had been learned respecting the properties of proteins of 
vegetable origin. Akhongh this work contained nmch of vahie to 
animal physiologists, and was suggestive in man\- ways in con- 
nection with the problems then claiming their attention, few of them 
appear to have read it with the care that it deserved. Authors of 
text-books on physiological chemistry, for many years after, dis- 
covered in it nothing more than the fact that Ritthausen employed 
dikite alkahne Solutions in isolating his preparations, and con- 
sec[uently dismissed his results with the Statement that all his 
products were altered in their preparation and so deser\'ed little 
consideration on the part of physiologists. Although such a criti- 
cism did not apply to the proteins soluble in strong alcohol which 
Ritthausen had described, and which after fifty years have l)ecome 
of much importance in the study of problems of nutrition. these 
remained for man\- years unknown to nearly every [)hysiological 
chemist. After the pulilication of this review, Ritthausen con- 
tinued his work until it included most of the seeds used for feeding 
men and animals. 

When Hoppe-Seyler and \\'e}l introduced neutral saline S(j1u- 
tions as solvents for many of the proteins of animal and vegetable 
origin, Ritthausen's results were regarded with increasing disfavor 
by physiological chemists. Undiscouraged by the unfair treatment 
accorded him, Ritthausen re-examined ])y the aid of salt solutions 
nearly all of the seeds which lie had previously studied, and also 
showed that most of his earlier preparations were still soluble in 



33^ llc'uiricli RittJuuiscii [April 

neutral sah snliuiun and were nnaltered in elcnienlar\- cumposition. 
Linie attention was, howex-er. paid \n this later work, although bis 
pa])ers were filled witli int'orniatiun that has proxed morc helpful 
in de\"el()i)ing" nur present knowledge of the ehemistrv uf proteins 
in general than has mosl of that ftn'nished l)\' bis critics. 

Aniong the many proteins which he acenralely described were 
several that could easily be obtained in well-formed crystals, a fact 
which. at that tinie. was of great importance in i)rotein chemistry. 
Thns. in 1881, he described a crystalline protein of the hemp-seed 
and the method for its preparation, which is essentially that now in 
nse. For more than twenty years this protein remained almost 
unknown although in recent years, under the name of edestin, it 
has been employed in hundreds of i)hysiological experiments in con- 
nection with a great variety of problems in protein chemistry. As 
a result of bis later w^rk he proved that wide difTerences exist 
between different food proteins; and he was the first to direct at- 
tention to this fact, and to discuss its probable bearing on their 
relative value in nutrition. 

Ritthausen's studies were not confined solely to the \-egetable 
proteins, as is evident from bis extensive bibliography which ap- 
pears at page 339. He made man}- im-estigations of other con- 
stituents of seeds. obtaining \icin and convicin from vetch seeds, 
and discovered in the cotton-seed " melitose " now known as 
raffinose. 

If we are to judge Ritthausen's work fairly we must remember 
that it was begun under the influence of Liebig's erroneous assump- 
tion that only a few forms of protein existed ; that at that time 
organic chemistr}- was in its infanc}-; that few methods were known 
by which jjroteins could be isolated from the tissues containing them, 
or by which the different proteins could be separated from one 
another and be purified : that the only means for preventing the 
changes caused by bacteria and enzymes were low temperatures ; 
and that the facilities for conducting such investigations were very 
limited. "Ko the writer, who has had a long experience in this same 
field. under the vastly more favorable conditions pre\'ailing a gen- 
eration later, it is astonishing that Ritthausen accomplished so much, 
and that the data he secured were in the main so accurate. What- 



1913] Lewis W. Fcfccr 339 

ever may have been the sh(jrtcomings of Ritthaiisen's work, the 
fact remains that he made a niost valuable contribiition to biological 
chemistry : and that instead of criticism, he deserves our gratitude 
and admiration for his patience and perseverance in one of the 
most difficult fields of investigation. 

Thomas B. Osborne, 

Connecticut Agricultiiral Experiment Station, 
N'ew Haz'cn. 



PUI5LICATI(JXS OF PROFESSOR HEINRICH RITTHAUSEX 

I. Journal für praktische Chemie 

Ueber die Aschenbestandtheile einiger Lycopodiumarten : Lvc. coiii- 
planatum, Lyc. Chaiiicrcyparissiis, Lyc. clavatuiii. sowie über die 
Säure von Lyc. coinplaiiatum : 1851, 53, 413. 

Xeue Analysen der Aschen einiger Lycopodiumarten: 1853, 58, 133. 

Ueber die Einwirkung des Sahniaks auf Kupfer; 1853. 59, 369. 

Zersetzung des Sahniaks durch Zink; 1853, 60, 473. 

Ueber einige Kohlenwasserstoffe des leichten Steinkohlentheeröls ; 

1854. 61. 74. 

Chemische L'ntersuchung der Runkelrübe; 1855, 65, i. 

Chemische Zusammensetzung des rothen und schwedischen Klees (Tr//. 

pratcusc und Trif. hyhriduni ) in verschiedenen \'egetations- 

Perioden ; 1855, 65, 8. 
A'eränderungen des Heus von Rothklee durch Auswaschung von Regen : 

1855, 65, 13. 

Ueber den Einfluss der Düngung mit Asche und Gyps auf die chemische 
Zusammensetzung des Klees; 1855, 65, 15. 

Destillations-Rückstände von der Spiritus-Fabrikation aus Kartoffeln 
(Schlempe) ; 1855. 66, 289. 

Rückstände von der Spiritus-Fabrikation aus Getreide; 1855, 66, 308. 

Rückstände, welche bei der Bierproduktion gewonnen werden (mit H. 
Scheven) ; 1855. 66, 311. 

Analysen der Asche von Gerstenmalz, Trebern und Malzkeimen; 1855, 
66, 315. 

Ueber die Bestandtheile des Weizenklebers ; 1862, 85, 193. 

Ueber die Zusammensetzung des Pflanzenleims und das \^erhalten des- 
selben zu \\ asser ; 1862, 86, 257. 



340 Heinrich Rillhaiiscii [April 

Ueber die Ziisanunensetzung des Pflanzenleims, 1863, 88. 141. 

Reactionen des l'flanzenleinis : 1863. 88. 142. 

Zur Darstellung des Pflanzenleinis ; 1803. 88, 145. 

Cholesterin im Fett des Weizens; 1863, 88. 145. 

Trimethylamin aus Weizenbrand; 1863, 88, I-17. 

L'eber die P>estandtheile des Weizenklebers; 1864, 91, 296. 

L'ntersncliungen ueber einige liestandtbeile des Roggensamens ; 1866, 

99. 439- 
Ueber die Cilutaminsiiure ; 186O, 99, 454. 
Ueber die Bestandtheile des Weizenklebers ; 1866, 99, 462. 
Ueber einige Bestandtheile des Roggensamens : In Weingeist lösliches 

Gummi; Cholesterin und Palmitinsäure im Fette des Roggens; 

Buttersäuregährung des Roggenmehls; 1867, 102, 321. 
Dolomitreicher Mergel; 1867, 102, 369. 

Lithionhaltiger Mergel und Boden in Ostpreussen ; 1867, 102, 371. 
Bildung von Mvianit im Grunde einer Düngergrube; 1867, 102, 373. 
Blasenstein (eines Ochsen) von Kieselerde; i8C>7, 102, 374. 
Soda als sogenannter Mauersalpeter; 1867, 102, ;i,j=^. 
Portland-Cement von Powunden ; 1867, 102, ^/(). 
Ueber das Pflanzen-Casein oder Legumin ; 1867, 103, 65, 193, 273. 
Ueber die Zersetzungsprodukte des Legumins und des Prote'inkörpers 

der Lupinen und Mandeln beim Kochen mit Schwefelsäure; 1868, 

103. 233- 
Ueber die (dutansäure, das Zersetzungsprodukt der ( llutaminsäure 

durch salpetrige Säure; 1868, 103, 239. 

Asparaginsäure und (Glutaminsäure, Zersetzungsprodukte des Legu- 
mins beim Kochen mit Schwefelsäure; 1869, 106, 445. 

Prote'instofife des Maissamens; 1869. 106. 471. 

Asparaginsäure und Glutaminsäure, Zersetzungsprodukte des Legu- 
mins und Conglutins beim Kochen mit Schwefelsäure; 1869, 107, 
218. 

Ueber die Säuren der Samen der gelben Lui)inen ; 1870, 2 ( n. f.), 339. 

Lieber das A'orkommen von Amygdalin und eine neue dem Asparagin 
ähnliche Substanz in Wickensamen (mit L^. Kreussler ) ; 1870, 2 

Ol. f.). 333- 
Leucin aus Pflanzenproteinstoffen (mit l". Kreussler) ; 187 r, 3 (n. f.), 

307. 
Lieber die \^erbreitung der Asparaginsäure und Glutaminsäure unter 
den Zersetzungsprodukten der Proteinstoffe (mit R. Pott) ; 187 1, 
3 (n. f-)- 3U- 



1913] Lc7\.'is JJ\ Fctzer 341 

A'erbindungen der Prote'instoffe mit Kupferoxyd ( Legumins, Con- 

glutins. GlutenL.-seins) ; 1872, 5 (n. f.), 215. 
Ueber das Drehungsvermögen von Glutan- und Aepfelsäure ; 1872, 5 

(n. f.), 354- 

Untersuchungen über A'erbindungen der Eiweisskörper mit Kupfer- 
oxyd (mit F. Weger); 1873, 7 ( n. f.), 361. 

Notiz ueber die asparaginähnhche Substanz im Wickensamen; 1873, 7 

(n- f-). 374- 

Ueber die Bestimmung des Stickstoffs der Eiweisskörper mittelst Na- 
tronkalk ; 1874, 8 (n. f.), IG. 

Neue Methode zur Analyse der ^vlilcli und ein vom Milchzucker ver- 
schiedenes Kohlehydrat in der Kuhmilch; 1877. 15 (n. f.), 329. 

Nachtrag hierzu; 1877, 16 (n. f. ), 22)7- 

Krystallinische Eiweisskörper aus verschiedenen Oelsamen ; 1881. 23 
(n. f.). 481. 

Ueber \'icin und eine zweite stickstoft'reiche Substanz der Wickensa- 
men. Convicin : 1881, 24 (n. f.), 202. 

Ueber die Einwirkung von Salzlösungen auf Conglutin und Legumin ; 
1881, 24 (n. f.), 221. 

Ueber die Eiweisskörper von Oelsamen; 1881, 24 (n. f.), 257. 

Ueber die A'erbreitung der ^lyronsäure in den Samen von Brassica 
iiapiis und ra[^a; 1881, 24 (n. f.), 2/2,. 

Zusammensetzung der Eiweisskörper der Hanfsamen und des krystal- 
lisirten Eiweisses auf Hanf- und Ricinussamen ; 1882, 25 ( n. f.), 
130. 

Ueber die Zusammensetzung des Krystallisirten Eiweisses aus Kürbis- 
samen ; 1882. 25 (n. f.), 137. 

Ueber das A'erh alten des chromsäuren Bleis bei A'erb rennungen und 
zu Sauerstoff; 1882. 25 (n. f.), 141- 

lieber das A'erhalten des Conglutins aus Lupinensamen zu Salzlösun- 
gen ; 1882. 26 (n. f.), 422. 

Ueber die Eiweisskörper der Pfirsichkerne und der Pressrückstände 
von Sesamsamen; 1882, 26 (n. f.), 440. 

Ueber das A'erhalten des Legumins zu Salzlösungen; 1882, 26 (n. f.), 

504- 
Ueber ^lelitose aus Baumwollensamen; 1884, 29 (n. f.), 351. 
A'orkommen von Citronensäure in verschiedenen Leguminosensamen ; 

1884. 29 (n. f.), 357. 
\'orkommen von \'icin in Saubohnen (U/cfa /aöa) ; 1884,29 (n. f.), 359. 
Lieber die Löslichkeit von Pflanzenproteinkörpern in salzsäurehaltigem 

Wasser; 1884, 29 ( n. f.), 360. 



342 Heinrich J\illliaiiscii [April 

Ueber Zusainiiicnsct/.uni^- der mittelst Salzlösung (laryestellten luweiss- 
körper der Saubohnen { ricia falhi) und weissen iUjhnen { I'hasc- 
olus), 1S84. 29 (n. f. ), 448. 

Ueber Betain aus Pressrückstiinden der lUiuniwollsanicn (mit Prcnss) ; 
1884. 30 (n. f.), 32. 

L'eber die Fdweisskörper des Weizenklebers oder Glutens ; 1890. 59 

(n. f.).474- 
Löslichkeit von Eiweisskörpern in Glycerin ; 189g, 59 ( n. f.), 479. 
Ueber die Zusammensetzung des Vicins ; 1899, 5g (n. f.), 480. 
Ueber Divicin ; 1899, 59 (n. f.), 482. 
Zusammensetzung des Convicins aus Wicken- und Saubohnensamen ; 

1899, 59 (n. f.), 487- 

2. Berichte der Versuchs-Station zu Möckern^ 

Untersuchungen des Grünfutters von dem amerikanischen Zahnmais 
und dem österreichischen Mais (mit E. Wolff ) : 1854, 3, i. 

A'ergleichende Untersuchung des schwedischen und des gewöhnlichen 
rothen Klees (mit E. Wolff) ; 1854, 3, 11. 

Chemische Untersuchung von Gras, Heu und Grummet ( mit E. Wolff) ; 

1854. 3. 18. ^ . 

Chemische Untersuchung der Runkelrübe: (a) Einfluss des Blattens 
auf die Zusammensetzung; (b) Einfluss der Grösse auf die Zu- 
sammensetzung: (c) Einfluss der A'arietät auf die Zusammenset- 
zung (mit E. Wolff") : 1854, 3, 22. 

Beobachtungen über die ~\Iilchproduktion bei dem Uebergang von der 
Winterfütterung zu der Grünfütterung (mit J. G. Bahr und E. 
Wolff ) : 1854, 3, 38. 

Ueber den Einfluss des im Dampf gekochten Futters auf die Alilch- 
produktion (mit J. G. Bahr) : 1855. 4, i. 

Ueber den Einfluss der Zuckerrüben auf Milchproduktion (mit J. G. 
' Bahr) : 1855. 4, 13. 

Düngungsversuche mit Knochenmehl, guanisirtem Knochenmehl, Blut- 
dünger, und Guano (mit J. G. Bahr und W. Knop) : 1855, 4, 15. 

\'ersuche mit Ueberdüngung von Chilsalpeter, Kochsalz und Guano bei 
Weizen und Roggen (von J. G. Bahr, mitgetheilt von H. Ritt- 
hausen) : 1855, 4- 22. 

^ Compiled from Nobbe's Quellenverzeichniss der hauptsächlichsten in den 
Jahren 1852 liis 1877 von den Versuchs-Stationen veröffentlichten wissenschaft- 
lichen Arbeiten, in Ilntii'ickhi)tg 11. TJüitigkcit d. laiuln'irtscliaftlichcii l'crsuchs- 
Sfafioiicii. etc.. 1877, pp. 284-435 (Berlin). 



1913] Lewis W. Fctzcr 343 

Düngung des Roggens mit Peruanischem Guano, Chilsalpeter, gebrann- 
tem, reinen Knochenmehl und Polenz'schen Guano (von J. G. 
Bahr, mitgetheilt von H. Ritthausen) ; 1855, 4, 29. 

N'ersuche mit verschiedenen Sorten Guano und Knochenmehl, Raps- 
kuchenmehl und Stallmist zu Weizen und Kartoffeln (von J. G. 
Bahr, mitgetheilt von H. Ritthausen) ; 1855, 4. 31. 

Einfluss der Düngung des Klee's mit Asche und Gyps ; 1855, 4, 41. 

l'ntersuchungen des schwedischen und rothen Klees; 1855, 4, 65. 

\"eränderungen des Heus von Rothklee durch Auswaschung von Regen ; 

1855. 4. 72>- 

Vergleichende Untersuchung der \\'intergerste, Annat- und Probstei- 
ger ste : 1855,4, 76. 

Ueber den Einfluss der Lupinen auf die ^lilchproduktion, ein Fütter- 
ungsversuch (mit J. G. Bahr) ; 1856, 5, i. 

Ueber die Zusammensetzung und den Nahrungswerth einiger in der 
Landwirtschaft als Futtermittel angewendeter Fabrikationsrück- 
stände (Kartoffelschlempe, Malz, Presshefe, Getreideschlempe) ; 

1856, 5, 15. 

Ueber einige Eigenschaften von Kulturpflanzen, die in gleicher Vege- 
tationszeit einen verschiedenen (irad der Entwickelung zeigen (mit 
H. Scheven) : 1856, 5, 67. 

Entwicklung und Thätigkeit der land- und forstwirthschaftlichen Ver- 
suchs-Stationen in den ersten 25 Jahren ihres Bestehens ; Fest- 
schrift zur Feier des 25 jährigen Jubiläums der A'ersuchs-Station 
Möckern ; 1877, p. 56. (A statement, made by Professor Ritt- 
hausen at the request of Prof. G. Kühn, in regard to work con- 
ducted by or under him while Director of the Experiment Station 
at Möckern.) 

3. Jahresberichte der Versuchs-Station Ida Marienhütte (bei 
Saarau) nach Breslau, gegründet A. D. 1857 

(Aus den Mittheilungen des landwirtschaftlichen Centralvereins für 

Schlesien )- 

Zusammensetzung" der Kuhmilch ; i, 59. 
Lieber Dünger-Fabrikation; i, 60. 

Analysen des Bodens der Ida-Marienhütte (mit P. Bretschneider) ; 
I, 82. 

" Compiled f rom Nobbe's agricultural bibliography, 1877. See footnote. 
page 342. The numerals for the years of publicatinn are not given in Nobbe's 
bihlioQrapli>-. 



344 II einrieb RiithiUiscn I April 

\'ersuclio über SaiiieiKlungung ; i. 85. 

\'ersiiche mit Ueberdüngung des Roggens ; i, 95. 

Düngungsversiicbe bei Rüben (Beta ) ; i. 104. 

Untersuchung von in gleicher A'egetationszeit, ungleich entwickelten 

Kulturpflanzen; i. 134. 
Untersuchung eines Torfes und seiner Asche; i, 145. 
Bestimmung der Asche in \'egetabilien ; i. 147. 
Bestimmung der Phosphorsäure; i, 148. 
Bestimmung der Kieselsäure in Pflanzenaschen; i. 149. 
Analysen des Bodens der Ida-Marienhütte (mit P. Bretschneider ) ; 

2. 36. 

Untersuchung von Zuckerrüben; 2, 66. 
Untersuchung von Zuckerrüben ; 4. 59. 

Analysen des Bodens der Ida-Marienhütte (mit P. Bretschneider) ; 6. 
100. 

4. Sächsiches Amts- und Amzeigeblatt 

^'ersuche über den Xahrungswerth der Kartofl:'elschlempe in A'ergleich 
zu Kartofi'eln und ]\Ialz, und süsser Maische, bei gleichen Mengen 
Rohmaterial (Kartoffeln und Malz) : Fütterungsversuche mit 
Kühen ( mit J. G. Bahr) ; 1856, p. 87. 

Fütterungsversuclie mit Kühen ueber den Einfluss von geschrotenem 
und gekochtem Getreide auf Milchproduktion (mit J. (i. Bahr) ; 
1856, p. 96. 

Ueber die Zusammensetzung einiger Wurzelgewächse (Rüben, Kohl- 
rüben und Strunkkraut) und den Einfluss der Grösse und Schwere, 
sowie starker Düngung auf die Zusammensetzung derselben; 1857, 

\'ersuch über die A'erdaulichkeit der Holzfaser des Futters beim Rind 
(mit H. Scheven) ; 1858, p. 58. 

5. Landwirtschaftlichen Versuchs-Stationen 

Orittheilungen aus dem Agriculturchemischen Eaboratorium der Fni- 

versität Königsberg i. Pr.) 

Untersuchungen ueber den Einfluss einer an Stickstoff' und Phosphor- 
säure reichen Düngung auf die Zusammensetzung der Pflanze und 
der Samen von Sommerweizen (mit R. Pott) ; 1873. 16, 384. 

Ueber die Einwirkung freier Phosphorsäure auf kohlensauren Kalk ; 
1877, 20. 401. 

I'eber den Fettgehalt der käuflichen K]eber]irä]:)arate ; 1877, 20. 408. 



iyi3] Lci^'is IV. Fct::cr 345 

Analysen einiger Futtermittel: 1877, 20. 409. 

Ueber den angeblichen (Jehalt des Roggensamens an Stearinsäure; 
1877, 20. 412. 

Berichtigung zu der Mittheilung von ]\I. von Sivera : Ueber den Stick- 
stoffgehalt des Torfbodens; 1880. 25. 169. 

l'eber Zerstörung von Fett durch Schimmelpilze Unit H. Baumann) ; 

1896, 47, 389- 
Ueber die Berechnung der Proteinstoft'e in den Wanzensamen aus dem 

gefimdenen Gehalte an Stickstoff; 1896. 47, 391. 

6. Berichte der deutschen chemischen Gesellschaft 

Ueber Mein: Ijestandtheil der Samen von l'icia sativa : 1876, 9, 301. 

^^'assergehalt und Reaktion des Alloxantins : 1896, 29. 892. 

Ueber Alloxantin als Spaltungsprodukt des Convicins aus Saubohnen 

(Ficia faba minor ) und Wicken (Jlcia satkv) : 189C). 29, 894. 
Ueber Galactit aus den Samen der gelben Lupine: 1896. 29. 896. 
Reaktionen des Alloxantins aus Convicin der Saubohnen und Wicken ; 

1896, 29, 2106. 
Mein ein Glycosid ; 1896, 29. 2108. 
Ueber Leucinimid. ein Spaltungsprodukt der Eiweisskörper beim 

Kochen mit Säuren; 1896, 29, 2109. 

7. Archiv für die gesammte Physiologie (Pf^üger) 

Die Eiweisskörper der Ptlanzensamen : 1877, ^S- -^V- 

Ueber den Stickstoffgehalt der Pflanzen-Eiweisskörper nach den ^leth- 

oden von Dumas und Will-\'arrentrapp (mit H. Settegast) ; 1878, 

16, 293. 
Ueber die Zusammensetzung der Proteinsubstanz der Bertholletia- 

(Para-) Nüsse; 1878, 16. 301. 
Ueber den Stickstoft'gehalt der Pflanzen-Eiweisskörper nach den ]\Ieth- 

oden von Dumas und WlU-Varrentrapp : 1878. 18, 236. 
Ueber die Eiweisskörper der Ricinussamen. der Proteinkörper, sowie 

der Krystalloide dieser Samen; 1879. 19. 15. 
Ueber die Eiweisskörper verschiedener Oelsamen ; 1880, 21, 81. 

8. Chemiker-Zeitung 

In Weingeist lösliches (iummi aus Roggen: Secalin : 1897, 21. 717. 
Zur Darstellung der Alkaloide der gelben Lupinen (Lup. Intens) ; 1897, 
21, 718. 



340 llciurich Ritlluuiscu [April 

9. Book 

Die lü\vcisski"ir])er ^\Q\■ ( ictreideartcn, I liilscii fniclite und (MsaiiK'n : 
Beiträge zur i'hysiologie der Samen der Kullurgewäehse. der 
Nahrungs- und Futtermittel; pages 252. Jionn (Max Cohen und 
Sohn), 1872. 

10. Miscellaneous publications 

Versuciie über Düngung von Rüben: Chemische .Ickeyniami, 1858, p. 

130; abs. in Jahresber. Ayr. Cheiii.. 1858, i, 226. 
Die Aschen einiger Futterpflanzen; M ift/ieiliiiu/en aus Jf'aldaii, 1859, 

p. 91 ; abs. in Jahresber. Jgr. Cheiii.. 1850. 2. 84. 
"Mug," ein Dungmittel; JJ'ocJieiiblatt der Annalcii der Laiid-ccirfseliaff, 

1861, p. 8; abs. in Jaliresber. Ayr. Chciii., 1861, 4, 195. 
Das A'erhalten der freien Ph^sphorsäure der Superphosphate ; Laiid- 

7cirtsch. Zeitung für das iiordöstlich.e Deutselüand, 1875, 11; abs. 

in Jahresber. Agr. Chein., 1875, ^^> 5^- 
Verlust an DüngstofTen im PJoden einer Düngerstätte (mit Ritsch- 

mann ) ; . Igrieulfureh. Ceiifralbl., 1876, p. 35; abs. in Jaliresber. 

Agr. Chein.. 1876, ig, 40. 
Ueber Proteinkörner, Krystalloide und krystallisirtes Ei weiss. Schriften 

der Physikalisch-Ökonouiisclieu Gesellschaft .::u Königsberg ; 1881, 

22, 15. 

Lewis W. Fetzer. 

Office nf E.vpcriincnf Statioits, 

U. S. Drparliiicnt of Agricultitre, 
und Ccorgcfon'ii t^iii'c'ersify, 
Jl'ashingtoii. D. C. 



DINNER TO PROFESSOR CHITTENDEN 
Testimonial by his pupils 

In December, 191 1, a nnml^er of the former pupils of Prof. 
Russell H. Chittenden, at a Conference in P)altimore. concluded " that 
the time had arrived when it would be appropriate to provide some 
formal expression of the esteem in which Professor Chittenden is 
held by those who appreciate his contributions to physiological chem- 
istry and education." Drs. S. W. Lambert, F. S. Meara, Holmes 
C. Jackson, S. P. Beebe, and William J. Gies were requested to 
serve as a provisional committee of five, to consider the matter 
further and to proceed with Organization and execution of plans, 
"if some step in this direction seemed appropriate, after further 
consideration." 

The pro\-isional committee of five decided to organize Professor 
Chittenden's pupils for the purpose indicated, and invited twenty 
additional former pupils of Professor Chittenden's to serve with 
them as a Committee of Twenty Five, as follows : John A. Hartwell, 
chainiiaii, T. S. Arbuthnot, S. P. Beebe, Joseph A. Blake, Harvey 
Cushing, H. H. Donaldson, Isadore Dyer, P. B. Hawk, Theodore 
C. Janeway, Elliott P. Joslin, J. H. ^[. Knox, Samuel W. Lambert, 
P. A. Levene, Frank S. ]Meara, Lafayette B. Mendel, Charles Norris, 
Thomas B. Osborne, Alfred N. Richards, E. AV. Rockwood, W. T. 
Sedgwick, W. Gilman Thompson, H. Gideon Wells, E. B. Wilson, 
Holmes C. Jackson, trcasiircr, and William J. Gies, sccrctary. The 
Committee of Twenty-Five rec[uested Drs. Hartwell, Beebe, Jane- 
way, Jackson, and Gies to serve as a subcommittee for the execu- 
tion of the committee's plans. 

The general committee has invited Professor Chittenden's pupils 
to coöperate in raising a Russell H. Cli'ittcndcn Fund, to be pre- 
sented to the Yale corporation without any other condition than that 
it be used for the advancement of the work of the department of 
physiological chemistry in the Sheffield Scientific School. 

The committee also authorized the secretary to invite Professor 

349 



350 Dinner lo Professor Chili enden [April 

Chittenden to be the gnest of bis pupils at a dinner in Xew York on 
^larcli T. In tlic formal imitation, the secrttar\- w n itc to Professor 
Chittenden. in part. as follows : 

The above-named Committee of Twenty Five has instructed nie to 
invite you to be the gnest of your many pupils and friends at a dinner 
in yonr bonor in Xew York City on Alarcb i, 1913. It is onr desire 
not only to bave the pleasure of yonr Company but also to extend to 
you onr personal and professional greetings. anfl to evidence onr friend- 
ship and respect. 

The dinner in bonor of Professor Chittenden was held at Del- 
monico's. on Satnrday evening, March i, and proved to be a delight- 
fnl event in evcrv particnlar. Althongh man}- who expected to be 
present were unable to attend, and sent letters of regret, about 
seventy-fi\-e pnpils and a dozen in\-ite(l friends comprised the en- 
thusiastic Company tbat made the dinner a cordial testimonial of 
affection and esteem for Professor Chittenden. 

Among those who were unable to accept invitations to be present 
at the dinner. and to speak afterwards. were President Hadley, of 
Yale, and Prof. William H. Welch, of Johns Hopkins University. 
In a letter expressing regret for bis unavoidable absence, President 
Hadle}' wrote. in part. as follows : 

When }ou are having the Chittenden dinner, I shall be three thou- 
sand miles away. But I do not want to let the occasion go by without 
a word of greeting. ( )ur universities are on the lookout for men who 
are either discoverers or teachers or Organizers. In Chittenden Yale 
has a man who is all three. 

Professor \\'elcb sent a teleoram in which he said : 



-fc>' 



Deeply regret unavoidable absence from banquet. Affectionate 
greetings and heartiest congratulations to Chittenden — the man and 
friend, the great teacher, investigator, administrator — who has ren- 
dered inestimable service to science, Yale, and countr}-. ^fay many 
vears of health and vigorous work be bis. 

Seated at the Speakers' table were Professors Harvey Cushing, 
Henrv H. Donaldson, lohn A. Hartwell. Russell H. Chittenden. 



THE MEMBERS OF THE GOVERNING BOARD 

Oh THF 

SHEFFIELD SCIENTIFIC SCHOOL 
OF YALE UNIVERSITY 

EXTENÜ CONGRATÜI.ATIUNS TO THEIR CüLLEAG.UE 
THE DIRECTOR OF THE (GOVERNING BOARD 

PROFESSOR 

RUSSELL HENRY CHITTENDEN 

PH. D..SC.D.,LL.D. 

ON THE OCCASION OF THE TESTIMONIAL DINNER 

GIVEN IN HIS HONOR BY HIS PUPILS 

AND FRIENDS ON 

MARCH FIRST NINETEEN HUNDRED AND THIRTEEN 

IN NEW YORK CITY 



THE EVENT WHICH HAS CALl.F.D KORTH THIS PERSONAL MANIFESTATION OF ESTEEM AFFORDS 
AN Ol'PÜRTUNlTY TO THK MHMKERS OF THE (»VKRNING BOARD TO GIVE FORMAL EXPRES- 
SION TO THEIR HIGH ESTIMATK OF l'ROFESStDR CHITTHNDF.NS CONTRIBUJI'JNS TO SCIENCE AND 
EDUCATION AND THEIR GRATIFICATION AT THE FAVORABLE RECOGNITION WHICH HAS BEEN 
ACCORDEDTO HIS DISTINGUISHED SERVICES IN THE PROMOTION OF PHYSIOLOGICAL RESEARCH. 

THE SUCCESS VVITH WHICH PROFESSOR CHITTENDEN HAS FURNISHED INSPIRATION FOR THE 
LIFE WORK Ol" 0THER5 IS WORTHY OF COMMFNDATION: HIS ENERGY RECALLS THE VVORDS 
OF ANOTHER EMINENT STUDENT OF NUTRITION : 

•■THE GRFATFST J<;Y OF THOSE WHr> ARE STF.EPED IN WORK AND WHO HAVE 
SUCCEEDED IN FINDINi:. NEW TRUTHS AND IN UNDERST ANDING THE RELATION 
OF THINGS TO FACH OTHER. LIES IN WORK ITSELF.' 

TO THE (iRATITÜDR AND REGARD OF PROFESSOR CHITTENDEN^S PUPILS HIS COLLEACUES OF 
THE GOVERNING BOARD NOW DESIRE TO ADD THE CORDIAL ASSW^ANCE OF THEIR BEST 
WISHES AND THEIR PERSONAL GRFJETINGS. 



•■ «.f» 



Cera*ti. 



Greetings to Professor Chittenden by bis CoUeagues of tbe Governing Board 

of tbe Sbeffield Scientific Scbool 



I9I3] 



'94 S 



OD 



3 



Frank S. Meara, Graham Lusk, William T. Sedgwick. William T. 
Porter and Elliott P. Joslin. 

At the conckision of the clinner, the chairman of the committee, 
Dr. Hartwell, extended tu Professor Chittenden the affectionate 
greetings of his pupils and friends, and informed him of the estab- 
lishment of the Russell H. Chittenden Fund for the Adzrinceinent 
of Pli\siologieal Cheniistry in the Sheffield Seientific Sehool. Dr. 




Faces of the golcl medal presented to Professor Chittenden by the National 

Institute of Social Sciences 



Hartwell stated that the amount of the fnnd and the time of its 
presentation to the Yale corporation will he annonnced at an early 
date. Dr. Hartwell conclnded his remarks by introducing the toast- 
master, Dr. Meara. who officiated in the graceful and inimitable 
manner in which he is accustomed to preside on such occasions. 
Informal after-dinner addresses were then made by Drs. Cushing, 
Donaldson, Joslin, Levene, Lusk and Sedgwick, to which Professor 
Chittenden responded. 

The Speakers who preceded Professor Chittenden paid eloquent 
tribute to the personality, influence, ser\-ice, and achievements which 
have made Professor Chittenden the Dean of American biological 
chemists. Professor Chittenden replied earnestly and wilh deep 
feeline to the cordial tribute which had been conveved in the senti- 
ments of the Speakers, in the abundant evidence of warm approval 
with which each address was received, and in the evident heartiness 
of his own reception. 



354 Dinner lo l'rofrssor Cliillriulrii [\\)r\\ 

Immedialcly aficr ilic conclusion of Professor C "liiUeiidcirs ad- 
dress, ilie loaslniaster announcc'd iliat ilie National InsiiUite of Social 
Sciences liad xoted a t^'old nicdal lo Professor Chillenden in reco^s^- 
nition of tlie distinciion lic lias aitained in original investigation in 
llic lield of plnsiological cbemistrv. Dr. H. Holljrook Curtis, sec- 
rclar}- oi ilie Inslitnte. made the presentation. The faces of Ihe 
niedal are shown on page 353-. 

Prof. Lafayette B. Mendel followed Dr. Cnrtis witli a presenta- 
tion of engrossed congratulatory resolutions A\liich had Ijeen adopted 
])}• Professor Chittenden's associates in the Governing Board of the 
Sheffield Scientific School. (See page 351. ) 

The accompanying group portrait \\as made jnst after seats 
had been taken at the tables. The names of all in attendance, ar- 
ranged in taljle groiips, are appended :^ 

Speakers' table 

Harvey Cushing Russell H. Chittenden William T. Sedgwick 
H. H. Donaldson Frank S. Meara William T. Porter 

John A. Hart well Graham Lusk Elliott P. Joslin 

Table i. Lafayette B. Mendel, S. J. Meltzer, Jacques Loeb, 
Yandell Henderson, Simon Flexner, P. A. Levene, Frederic S. Lee. 

Table 2. Henry Hun, William Browning, H. H. Curtis, 
Henry Ling Taylor, W. M. Kenna, Harry Saltzstein, Frank C. 
Gephart, H. G. Barbour. 

Table 3. John Rogers, Wm. L. Culbert, R. H. Wylie, William 
Armstrong, Joseph A. Blake, W. L. Griswold, G. Wyckofif 
Cummins. 

Table 4. Theodore C. Janeway, J. H. M. Knox, S. P. Good- 
hart. Chas. H. Studio, George S. C. Badger, Joseph S. Wheelwright, 
N. R. Norton, Joseph H. Pratt. 

Table 5. Charles Norris, A. N. Richards, E. K. Dunham, 
John A. Mandel, H. D. Dakin, George B. Wallace, W^illiam J. Gies, 
Holmes C. Jackson, William C. Lusk. 

^ The men at tables 1 1 and 12, when the photograph was taken, subsequently 
reassembled at tables 9, 10 and 2. One or two additi(3nal rearrangements account 
for the disagreement between the indications of the portrait and the table lists. 



I9I3] '94 S 357 

Table 6. S. P. Beebe, G. A. Hanford, A. L. Dean, Frank P. 
Underhill, Benjamin White, Oswald T. Avery, Leo F. Rettger, S. 
R. Benedict. 

Table 7. Isadore Dyer, Wm. C. Wurtemberg, Robert Taylor 
W'heeler, Donald Guthrie, Henry H. Janeway, A. W. Elting, 
Charles L. Scudder. 

Table 8. Lewis F. Frissell, Seth M. Milliken, Robert P. Wad- 
hams, William P. Healy, Norman E. Ditman, W. W. Herrick, M. 
Heminway Merriman, Cyrus W. Field. 

Table 9. Frank C. Yeomans, Alfred Jerome Brown, J. L. 
Bendell, Isaac F. Harris, Stanley D. Beard, Frank E. Haie, E. 
JMonroe Bailey, Otto G. Hüpfel. 

Table 10. Israel S. Kleiner, Lewis H. \\'eed, Orville H. 
Schell, Simon B. Kleiner, Morris S. Eine, Victor C. ]\Iyers, Warren 
W. Hilditch, Henry C. Courten. 

Adjournment occurred at a very late hour, bnt many tarried to 
discuss informally the happy events of the evening and to talk over 
" old times" in Chittenden's laboratory. 

'94 s. 

-Vc^z^' York City. 



SOCIETY FOR EXPERIMENTAL BIOLOGY AND 

MEDICINE 

Tenth anniversary meeting and dinner 

The tenth anniversary of the estal)Hshment of the Society for 
Experimental Biology and Aledicine was celebrated in Xew York 
on the iQth of February. The fifty sccond regulär scientific meet- 
ing was held, at 4 p. m.. at the College of Physicians and Surgeons, 
in the lecture room adjoining the main biochemical laboratory. 
After the conclnsion of the scientific meeting. at about 7 ]). m., the 
members adjonrned to the Hofbräu Honse (39th Street and Broad- 
way), where, in a body, they feasted on a beefsteak dinner prepared 
nnder the auspices of a committee of which Prof. Graham Lusk 
was chairman. 

The scientific session was the most interesting and important 
in the history of the society. The nature of the proceedings is 
shown by the appended copy of the official program : 

G. X . Calkiiis: Further light on the conjugation of paramecium. — 

JJ\ H. Maincaring and J. Bronfcnhrciuier: On lysis of tubercle bacilli 

(II) ; '^'On chemotherapeutics of tuberculosis. — U\S. Halstead: Hyper- 

trophy of the thyroid ; Partial occlusion of the aorta by bands of living 

tissue. — G. H. A. Cloivcs: Hay fever, with demonstration. — *J. /. 

Ringer: Further studies on the fate of fatty acids in the diabetic organ- 

isni. — R. M. Pcarcc and P. F. Williams: Experience with Abderhalden's 

test for pregnancy. — L. L. JJ'oodruff: The kernplasma relation during 

the life of a pedigreed race of O.vyfliricha f alias. — Richard JJ'cil: A 

new factor in anaphylaxis. — E. E. Biiffcrficld: The reaction between 

oxygen and hemoglobin. — *F. S. Lee and S. Everinglmin: The myo- 

neural junction in fatigue. — F. H. Pike: A demonstration of the effects 

of electrical Stimulation of the labyrinth of the ear. — E. L. Scott: The 

relation of pancreatic extract to the sugar of the blood. — A. F. Hess: 

The pancreatic lipase of infants in acute intestinal disturbances. — JV. 

H. Park, L. ]]'. Fainiilcucr and E. J. Banzhaf: Influence of protein 

concentration on absorption of antibodies in sul)cutaneous injections. — 

* On the ofificial program, but not abstracted in the Proc. Soc. Ex[>. Bio!, 
aiid Med.. 1913. x. pp. 65-122. 

;8 



J3' 





c^ 




Portrait of the Foundcr ot the Society for Experimental Biology and Medicine. 
Reproduced from \'olume II ( 1904-05 ) of the Society's Proceedings 



1913] / Ninetccn O. Tlircc 361 

G. C. Robinson: The inflnence of the vagits nerves 011 tlie faradized 
auricles in the dog heart. — B. S. Oppenheimer and H. B. Williams: Pro- 
longed complete heart block with freqiient changes in the idio-ventricu- 
lar complexes. — C. J. Wiggers and E. F. DnBois: Methods for the pro- 
duction of temporary valvulär lesions.— .^. /. Gold färb: The influence 
of the central nervons System on regeneration ; The effect of salinity 
lipon regeneration. — B. T. Terry: Variations in the amount of trans- 
formed atoxyl (trypanotoxyl) produced by varying the strength of 
atoxyl inciibated with blood. — IV. J. MacNeal and A. F. Chace: Some 
observations on bacteria of the dnodenum. — A. F. Colin: The effects 
of morphin on the mechanism of the dog heart after removal of one 
vagus nerve. — T. S. Githens: The influence of temperature on the mini- 
mal dose of strychnin and the onset of tetanus in the frog. — '■'/. E. 
McWhorter and F. Prime (by invitation') : Cinematographic demon- 
stration of the growth of tissues. — '''F. S. Lee: Cinematographic dem- 
onstration of the beating heart. — "7?. Bnrton-Opitz: Demonstration of 
the vasomotor nerves of the liver. — *//. B. IVilliams: Demonstration 
of the electrovagogram. — Wm. de B. MacNider: The difl^erence in the 
efifect of Grehant's anesthetic and of morphin-ether on the total output 
and composition of the urine in normal dogs. — Sntherland Simpson: 
The rate of growth in the dog. — Andrew Hnnter: The influence of ex- 
perimental cretinism upon nitrogenous metabolism in the sheep. — De- 
Witt Stettcn and Jacob Rosenblooni: ]\Ietabolism studies in a case of 
hypopituitarism, with infantilism of the Lorain t}pe. — /. P. Atkinson 
and C. B. Fitspatrick: On the presence of pressor substances in experi- 
mental immunity. — G. H. A. Clowcs, Francis C. Goldsborongh and F. 
West: On a complement-deviation reaction exhibited in pregnancy. — 
G. H. A. Clozces and Francis C. Goldsborongh: On the antitryptic reac- 
tion exhibited in pregnancy. 

Prior to adjournment an election of officers for 191 3-' 14 oc- 
curred, with the following results : President, Janics Ezcing (re- 
elected ) ; vice president, Cyrns U\ Ficld; secretary, Hohnes C. 
Jackson; treasurer, Charles Morris (reelected). The new members 
elected were Russell L. Cecil, Gary Eggleston, K. George Falk, 
Davenport Hooker, Paul E. Howe and Charles J. West. 

The dinner was a very enjoyable event. The accompanying 
group Portrait shows the condition of the party at midnight. All 

* On the official program, but not abstracted in the Proc. Soc. E.vp. Bio!, and 
Med., J')i3, X. pp. 65-122. 



?62 



Society fov Exf^criiucuhil Hioloj/y aiuf Mcdiclnc 



April 



iliose at tlic rcadcr's cxnxnic Icfl aiid ri^lii wlio did not gel inlo the 
piclurc wcrc under ihe table wlieii ihe jjluHograpli was taken.'' 

The ])resi(lent, Prof. James r^wing-, ably and entenainingh- eon- 
cliKied llie afler-dinner proceedings. Infonnal speeclies were made. 
at the call of the President, l)y the distinguished fonnder, Dr. S. J. 
iMeltzer. also \)\ 1 )rs. (iraham T.usk. I'"rederic S. Lee. G. H. A. 
Clowes. Jac(|ues Loeb. William II. Park and William J. Gies. 

The Speakers felicitated Dr. Meltzer on the happiness of the idea 
that led Ihm to found the societ}' ; they also complimented him on 
the societ}''s past ser\ice. and on its rigor and effectiveness at the 
tenth anniversary of its birth. There was a strong note of con- 
gratulation of the society itself on the prospect of steady growth 
in efficiency and nsefnlness. 

The names of the members present at the meeting or at the 
dinner, or both, are appended : 



J. P. Atkinson 
John Auer 
T. H. Austin 

F. W. Bancroft 
S. P. r.eebe 

Jacob üronfenbrenner 
E. E. Butterfield 

G. X. Calkins 
G. H. A. Clowes 
A. E. Cohn 
Riifus I. Cole 

J. W. Draper 
E. F. Du Rois 
E. K. Dunham 
A. B. Eisenbrey 
C. A. Eisberg 
Haven Emerson 
James Ewing 
L. W. Famulener 
Cyrus W. Field 
C. B. Fitzpatrick 
Simon Flexner 
N. B. Foster 
W^illiam J. Gies 



T. S. Githens 
A. J. Goldfarb 
W. S. Halstead 
Isaac F. Harris 
Alfred F. Hess 
Paul E. Howe 
H. C. Jackson 
Walter A. Jacobs 
H. H. Janeway 
Don R. Joseph 
Ludwig Käst 
L S. Kleiner 
R. A. Lambert 
Frederic S. Lee 
P. A. Levene 
Isaac Levin 
Charles C. Lieb 
Jacques Loeb 
\\\ F. Longcope 
(iraham I,usk 
W. G. MacCallum 
W. T. AFacXeal 
F. H. ^IcCrudden 
A. R. Mandel 



John A. ^Mandel 
W. H. Manwaring 
S. T. ^leltzer 
G. '^r. Meyer 
H. O. :\Iosenthal 
John R. Ahndin 
j. B. ALn-phy 
A". C. Apvers 
Hideyo Xoguchi 
Charles Xorris 
B. S. Oppenheimer 
A. M. Pappenheimer 
William H. Park 
R. ^L Pearce 

F. H. Pike 
A. L Ringer 

G. C. Robinson 
Pevton Rons 
E.'L. Scott 

( r. G. Scott 

H. D. Senior 
M. Sittenfeld 
Edna Steinhardt 
H. A. Stewart 



^ Drs. Auer, Bancroft, Dunham, Eisenbrey, Field, Hess, Jackson, Mandel 
brothers, Norris, Oppenheimer, Park, Pearce. Senior, Swift, Wadsworth, Wal- 
lace, Wood. Anticipating the fate of the Michigan editor in the Roosevelt water- 
wagon case, we wish to add that \ve do not l3elie^"e this situatiiMi implics any- 
thing more than the facts themselves indicate. 







c\ 



. — I 






3 






o 

CO 



I9I3] 



Nineteen 0. Three 



365 



H. F. Swift 
B. T. Terry 
D. D. Van Slyke 
A. B. Wadsworth 



George B. Wallace 
Richard Weil 
C. J. West 
C. J. Wiggers 



Anna W. Williams 
H. B. Williams 
Francis C. Wood 



The Society for Experimental Biology and Medicine has been an 
important influence in the development o£ biological and medical 
science, particularly in New York. It has stimulated aspiration, 
quickened activity, increased productivity, afforded a congenial and 
ready means of expression, and opened a suitable Channel for com- 
munication, during a period of awakening in the biological and 
medical sciences in New York. It continues in this röle as an in- 
fluential factor in the advancement of science in this country. 

The growth of the society is indicated by the appended tabula- 
tion of its total membership at the end of each successive academic 
year since its foundation in 1903 : 



Year 


Total 


Increase 


Year 


Total 


Increase 


Year ; Total Increase 


1903 
1904 

1905 
1906 


19 

55 

87 

119 


36 
32 
32 


1907 
1908 
1909 
1910 


140 
162 

185 
205 


21 
22 

23 
20 


1911 

1912 

1913: 

(Feb. 19) 


222 
239 

255 


17 
17 

16 



Biological chemists may be interested in the following Statistical 
summary relating to the Society for Experimental Biology and 
Medicine : 

Of the seven men at the Conference in Prof. Graham Lusk's 
home preliminary to Organization, on January 19, 1903, four were 
biological chemists. 

The society was formally organized at a meeting in the bio- 
chemical laboratory of Columbia University, at the College of 
Physicians and Surgeons, N. Y., on Feb. 25, 1903. 

Two of the three authors of the Constitution, and two of the first 
five officers, were biological chemists. 

The folloAving members of the American Society of Biological 
Chemists are members of the Society for Experimental Biology and 
Medicine : 



J. J. Abel, J. G. Adami, H. M. Adler, C. L. Aisberg, J. P. Atkinson, 
E. J. Banzhaf, S. P. Beebe, F. G. Benedict, S. R. Benedict, W. N. Berg, 



366 Society for Experimental Biology and Mediane [April 

F. J. Birchard, Russell Burton-Opitz, R. H, Chittenden, A. C. Craw- 
ford, H. D. Dakin, E. K. Dunham, C. W. Field, Otto Folin, N. B. 
Foster, C. Stuart Gager, R. B. Gibson, William J. Gies, Shinkishi Hatai, 
R. A. Hatcher, P. B. Hawk, Paul E. Howe, W. H. Howell, Reid Hunt, 
Andrew Hunter, H. C. Jackson, W. A. Jacobs, Walter Jones, J. H. 
Kastle, I. S. Kleiner, Oskar Klotz, J. B. Leathes, P. A. Levene, Jacques 
Loeb, A. S. Loevenhart, Graham Lusk, A. B. Macallum, J. J. R. Mac- 
leod, W. deB. MacNider, J. A. Mandel, F. H. McCrudden, L. B. 
Mendel, G. M. Meyer, J. R. Murlin, V. C. Myers, F. G. Novy, T. B. 
Osborne, Franz Pfaff, A. N. Richards, A. I. Ringer, T. B. Robertson, 
Jacob Rosenbloom, William Salant, P. A. Shaffer, H. C. Sherman, 
Torald Sollmann, L. B. Stookey, A. E. Taylor, F. P. Underhill, D. D. 
Van Slyke, G. B. Wallace, H. G. Wells, C. G. L. Wolf. 

Of the 775 Communications to the Society for Experimental 
Biology and Medicine at its first fifty-two meetings, 430 — more 
than half — were largely er entirely biochemical in character. 

NiNETEEN O. ThREE 

New York City 



METHODS FOR THE ELECTROMETRIC DETERMI- 
NATION OF THE CONCENTRATION OF HYDRO- 
GEN IONS IN BIOLOGICAL FLUIDS 

K. A. HASSELBALCH 
(Finsen Institute j Copenhagen, Denmark) 

(WITH PL ATE 3) 

The great importance of the reaction of the medium in many 
biological processes has long been appreciated and has led to a series 
of more or less successful endeavors to measure its degree. It is 
only within the most recent years, hovvever, that the methods of 
measurement have been so far perfected as to enable us to say that 
the "true reaction" of biological fluids can now be measured with 
sufficient accuracy for most purposes. 

This is due, in the first instance, to the insight into the nature 
of the question which has been derived from the electrolytic dis- 
sociation theory : the " true reaction " of a liquid is not determined 
by its concentration of free acid or alkali, but by its concentration 
of hydrogen and hydroxyl ions — or, practically speaking, by the 
concentration of hydrogen ions alone, for the product of the two 
is a constant. Since the dissociation of acid or base in a liquid, and 
thereby also its hydrogen-ion concentration, is in many respects 
dependent on the nature of the dissolved substances, the true reac- 
tion of the liquid cannot be determined by merely measuring the 
quantity of alkali or acid that must be added to a certain quantity 
of the liquid in order to effect a particular change of color in the 
indicator used. As is now known, such a titration shows only that, 
at the moment, a certain hydrogen-ion concentration, to which the 
indicator reacts, has been reached, the original hydrogen-ion con- 
centration of the liquid remaining unknown. 

Thus, the determination of the true reaction of a liquid requires 
some procedure by which the concentration of the hydrogen ion is 

367 



3(38 Hydro gen Ions in Biological Fluids [April 

not altered. Only two methods of the latter kind are in practical 
employment, namely, the colorimetric and the electrometric. 

The colorimetric method is based on the above-mentioned fact, 
that a series of indicators shows certain color nuances with known 
hydrogen-ion concentrations, which miist be determined electro- 
metrically, so that the electrometric determination of the hydro- 
gen-ion concentration miist at any rate be considered as the funda- 
mental method. The colorimetric method has been indicated by 
Friedenthal and Salm ;^ its field has been considerably widened and 
its trustworthiness assured by the thorough investigations and im- 
provements of S. P. L. Sörensen and his collaborators. I shall not 
discuss the technical details of the method but merely refer to Sören- 
sen's latest smnmary of his work.^ 

We owe the electrometric method orginally to Nernst.^ It was 
first applied to biological fluids by Bugarsky and Liebermann, ^ and 
by Höber.^ It is based on the fact that a hydrogen-saturated metal 
electrode in a hydrogen-saturated liquid gives rise to a difference of 
Potential between the electrode and the liquid, which is dependent 
on the hydrogen-ion concentration according to known laws. The 
determination of this difference of potential thus makes it possible 
to determine the hydrogen-ion concentration of the liquid. 

The experimental method generally employed to measure the 
difference of potential between the hydrogen-saturated electrode 
and the hydrogen-saturated liquid has been so often described in its 
main features, most recently by Sörensen*^ in the above-cited work, 
that it needs no attention here. The present paper deals with the 
difficulty of obtaining the condition presupposed by the method, 
vis., Saturation of the electrode and liquid zvith hydrogen, without 
any alteration in the hydrogen-ion concentration of the liquid. 

Biological fluids, as is well known, usually contain volatile acids 
(or bases) which determine, in great part, their hydrogen-ion con- 
centration, so that the normal electrometric method, by which liquid 
and electrode are saturated with a current of hydrogen bubbled 

^ Friedenthal and Salm: Zeitschr. f. Elektroch., lo, 1904; 12, 1906; 13, 1907. 

^Sörensen: Ergebnisse der Physiologie, 12, 1912. 

^ Nernst : Zeitschr. f. physikal. Chemie, 4, 1889. 

* Bugarsky and Liebermann : Pflüger's Arch., 72, 1898. 

° Höber: Pflüger's Arch., 81, 1900. 

' Sörensen : Loc. cit. 



I9I3] K. A. Hasselbaich 369 

through the fluid, cannot be used. Let us take an extreme case and 
see what even a slight carbonic-acid tension — according to ordinary 
ideas — may mean for the hydrogen-ion concentration o£ a liquid. 
Sea-water and the surrounding atmosphere have the same carbonic 
acid tension — ca. 0.04/100X760 = 0.3 mm. On driving all the 
carbonic acid f rom the sea-water we should cause the hydrogen-ion 
concentration to sink from ca. lo"^ to ca. lO"^ or, using Sörensen's 
terminology/ the hydrogen-ion exponent, pn-, would rise from 8 
to 9. If, therefore, in this case we saturated the liquid and elec- 
trode with a current of hydrogen, quite an erroneous result would be 
obtained. 

A similar error, though less in amount, would also arise if we 
had recourse to the auxiliary method applied in such cases at the 
beginning of this Century, namely, if the hydrogen-saturated elec- 
trode remained in contact with the liquid and we waited until the 
Potential became constant, i. e., until equilibrium had been attained 
in the diffusion between the liquid and the hydrogen atmosphere. 
For instance, a sample of sea-water (kept in a bottle for nearly a 
year), whose p-a: was in reality 7.55, showed /'h- = 7.74 on using 
this method. The error may be reduced if, with Michaelis,^ we take 
a small quantity of hydrogen and let the electrode only just touch 
the surface of the liquid. But the most satisfactory method of 
proceeding seems to me the f ollowing :^ 

A current of pure hydrogen, saturated with moisture, is led 
through the vessel containing the electrode until the latter has be- 
come saturated with hydrogen. The experimental liquid, which is 
stored in such a way that it retains its natural tension of volatile 
acid (or base), is now led into the vessel in such a quantity that 
the electrode reaches more or less deeply into the fluid (see below) 
and the vessel is then closed. By shaking the vessel, the establish- 
ment of diffusion equilibrium between liquid and hydrogen, and 
attainment of constancy in the measured potential, are accelerated. 
This constancy, however, has been obtained by the loss of part of 
the volatile acid (or base) from the liquid to the hydrogen and 

^Sörensen: Bloch. Zeitschr., 21, 1909. 

^Michaelis: Ibid., 18, 1909; 46, 1912. 

'Hasselbaich: Ibid., 30, p. Z17, 1910; 38, p. 77, iQiS; 49, P- 45o, 1913- 



370 Hydrogen Ions in Biological Fluids [April 

would, therefore, indicate a too alkaline (or too acid) reaction of 
the fluid. The liquid is now renewed without changing the gas- 
mixture around the electrode, and shaking is repeated. It is easily 
Seen that electromotive constancy may now be obtained without 
any, or at least without any appreciable, alteration of the tension of 
the volatile acid (or base), i. e., without alteration of the original 
hydrogen-ion concentration of the liquid. 

This procedure may be repeated, if necessary, until the renewal 
of liquid no longer causes any alteration in the measured potential. 
For blood, urine, and probably the majority of biological fluids, a 
Single renewal is sufficient. Sea-water and similar Solutions, which 
are poor in "reaction regulators "^^ (Henderson^'^), are eo ipso far 
more susceptible to the change in carbonic-acid tension resulting 
from the method of measurement and would require three to four 
or, according to circumstances, even a larger number of renewals 
of the liquid before constancy is reached. In such cases it is easier, 
and more correct, to extrapolate graphically from the first three 
measurements (i. e., after two renewals of the liquid) in order to 
get the final value. 

The procedure described here permits one inconsiderable error, 
for which a correction may be made, if necessary. When diffusion 
equilibrium between the hydrogen and the liquid has been obtained, 
none of the components are, strictly speaking, any longer saturated 
with moist hydrogen at the existing barometric pressure but at a 
somewhat lower pressure. The potential changes, however, in ac- 
cord with the logarithm of the hydrogen pressure, so that, e. g., a 
fall in the hydrogen pressure from 760 to 700 (and a greater fall is 
practically inconceivable) would drop the measured potential to a 
value ca. i milli-volt too low. An error like this lies very near the 
limit of error of the whole method but may be eliminated, as already 
mentioned, by analysis of the hydrogen mixture and by calculation. 

Solutions which are poor in "reaction regulators," but whose 
hydrogen-ion concentration (owing to the volatile acid or base they 
contain) must necessarily be measured in the above-mentioned way 
(if it cannot be measured colorimetrically), have been found to 

'" Compounds which, by their presence, diminish the effect on the hydrogen- 
ion concentration of changes in the Proportion of acid or base. 
" Henderson : Ergebnisse der Physiologie, 8, 1909. 



1913] K. A. Hasselbaich 371 

present the difficulty that the potential changes in an unaccountable 
nianner when the liquid in the electrode vessel is at rest. For this 
reason I have proposed, as a normal method in measuring biological 
fluids, that the shaking should be mechanical and permanent, even 
during the reading of the electrometer. The electrode vessel is seen 
in Plate 3, Fig. i. The arrows indicate the direction and extent 
o£ the movement. While the shaking takes place, the electrode is 
constantly immersed in the liquid. 

It is easily seen that this electrode vessel, by another arrange- 
ment of the T-tube, may also be used in measurements which permit 
hydrogen to be led through the liquid. This property of the vessel 
may be of use, e. g., in efforts to control the correctness of the 
electrode by measurement of " Standard Solutions " of a known 
hydrogen-ion concentration. 

Fig. 2 (Plate 3) shows an electrode vessel used by me for 
small quantities of fluid, especially in determinations of the hydro- 
gen-ion concentration in 2-3 c.c. of human blood to which some 
hirudin is added. The blood may be taken f rom the lobe of the ear ; 
it is saturated in a glass syringe by rotation with about 20 c.c. of 
the alveolar air of the individual. The electrode, F, is saturated 
with a current of hydrogen flowing in the direction A— > B -» D -^ E. 
D is a groove in the inner part of a ground glass stopper lubricated 
with Vaseline and, during the flow of the hydrogen, it is turned so 
as to be opposite the hole E in the outer wall of the apparatus. The 
Saturation with hydrogen being completed, D is turned, as shown 
in Fig. 2 (Plate 3), and the cock B, which must be quite free 
from Vaseline, is turned around so that the first portion of the 
liquid from the syringe passes through A and down into the rubber 
tube C. The electrical connection between the liquid in the elec- 
trode vessel and the Solution of potassium chlorid (Fig. i, Plate 3) 
takes place along this route. When cock B is then turned as shown 
in the figure (Fig. 2, Plate 3), and cock H (which is carefully 
lubricated) is opened, the liquid rises in the electrode vessel as high 
as the side-tube ; H is now closed, B turned around, and the syringe 
disconnected. The shaking of the apparatus and the reading of the 
electrometer may now be started, 

When we are dealing with blood or other fluids containing dis- 



372 Hydrogen Ions in Biological Fluids [April 

sociable oxygen Compounds, electromotive constancy is not attained 
until the moment when the layer of liquid into which the electrode 
projects is completely reduced. In such cases it may be useful, by 
saving time, to apply the Suggestion of Michaelis,^^ namely, to let 
the electrode just touch the surface of the liquid. 

I am of the opinion that by following the lines indicated above, 
we shall be able to measure the hydrogen-ion concentration of bio- 
logical fluids in many cases where it has hitherto been considered 
impossible, or where we have had to be satisfied with rough approxi- 
mations. There are undoubtedly numerous questions in biology 
and pathology which these improvements in method may help to 
solve. 

" Michaelis : Loc. cit. 



A METHOD FOR THE DETERMINATION OF 

TRYPTOPHAN DERIVED FROM 

PROTEIN 

JESSE A. SANDERS and CLARENCE E. MAY 
(Chetnical Laboratories of Indiana University, Bloomington, Ind.) 

Introduction. Tryptophan is a protein cleavage prodiict that 
is never obtained abundantly. So far as we know the tryptophan 
yield has been determined quantitatively in the case of but two 
proteins, namely casein^ and wheat ghadin.^ Only traces of tryp- 
tophan can be obtained from other proteins; and some proteins, 
especially gelatin, fail to yield it, if the indications of the usual test 
with glyoxyhc acid and sulfuric acid are rehable. 

Although tryptophan cannot be abundantly obtained from pro- 
teins, considerable importance is attached to it because it is produced 
in the tryptic digestion of protein and, in putrefaction, yields indol. 
The quantity of indican in urine indicates, in a general way, the 
extent of intestinal putrefaction. One usually accepts that con- 
clusion without considering the details of the tryptophan trans- 
formation, which involves the necessary presence of tryptophan 
precursors in the original protein molecules ; the degree of digestion 
of the particular proteins that yield tryptophan; the conversion of 
tryptophan into indol rather than skatol ; followed by the absorption 
of indol, its oxidation to indoxyl, its esterification and its excre- 
tion in the urine in the form of the potassium ethereal sulfate. 

Owing to the evanescent nature of tryptophan, its Isolation from 
tryptic digestion mixtures has been the subject of many investiga- 
tions. Although Hopkins and Cole, Abderhalden, and others, have 

^Abderhalden: Zeit. f. physiol Chem., 190S, xHv, p. 23; Abderhalden and 
Samuely: Ibid., p. 276. (100 gm. of gliadin yield about i.o gm. of tryptophan; 
100 gm. of casein yield 1.5 gm. of tryptophan.) 

^ Osborne and Clapp : Amer. Jour. Physiol., 1906, xvii, p. 231; Osborne and 
Guest : Jour. of Biol. Chem., igii, ix, p. 426. (Hydrolysis of gliadin; revised 
gliadin-tryptophan figures.) 

373 



374 Determination of Tryptophan [April 

used the mercury stilfate-sulfiiric acid method^ in work on casein, 
this treatment has always been reported as giving figures somewhat 
lower than actual valnes. Tryptophan diminishes in quantity after 
a time, and may disappear, during the progress of tryptic digestion. 
In this laboratory we have found that the mercury sulfate-sulfuric 
acid method of Hopkins and Cole does not completely precipitate 
the tryptophan present in the digestion mixture. After precipitating 
the tryptophan-mercury-sulfate product from a casein digestion 
mixture, fikering, neutrahzing with calcium hydroxide and remov- 
ing the calcium sulfate and insoluble calcium salts, we obtained a 
filtrate that gave a characteristic tryptophan test with glyoxylic and 
sulfuric acids. Obviously the Hopkins-Cole method did not com- 
pletely precipitate tryptophan. Because of the smallness of the 
amounts of tryptophan usually derived from proteins, this method is 
necessarily dependent on the use of relatively large quantities of 
protein. The tedious nature of the methods for the purification of 
large amounts of proteins led us to endeavor to devise an accurate 
process involving the use of small amounts of protein. * 

The Solution of the problem seemed to depend on perfecting a 
method for the quantitative determination of small amounts of 
indol. We desired to use i^-napthoquinone mono-sodium sulfonate, 
such as Herter* employed in his work on indol, but could not find 
it on the market. We prepared a substance that reacted with indol, 
giving a deep violet colored Solution such as Herter obtained, but the 
substance formed by the combination of indol with our supposed 
;8-napthoquinone mono-sodium sulfonate was not soluble in Chloro- 
form. Herter used Chloroform to extract the indol-containing Com- 
pound. Further use of our reagent was abandoned. It is probable 
that we had an isomer of Herter's reagent differing mainly from his 
in its reaction with Chloroform. We prepared our reagent by 
cautiously oxidizing " Eikonogen," the Photographie developer, by 
means of concentrated nitric acid. The oxidation was quite satis- 
factory but the_ substance obtained was evidently an isomer, bearing 
the sulfonic acid radical on a benzene nucleus other than the one 
holding the quinone linkages. 

* Hopkins and Cole: Jour. of PhysioL, 1901-2, xxvii, p. 418; Ibid.j 1903, 
xxix, p. 451. (Mercury sulfate-sulfuric acid method; Isolation of tryptophan.) 

* Herter and Foster: Jour. of Bio!. Chent., 1905-6, i, p. 257; Ibid., 1906-7, ii, 
p. 267. (ß-napthoquinone reaction with indol.) 



1913] Jesse A, Sanders and Clarence E. May 375 

General method. We studied the production of indol in the 
tryptic digestion of casein, the tryptophan yield from which is known 
approximately. We used small amounts (i. 0-1.75 gm.) of casein, 
digesting them with strong pancreatin Solutions free from trypto- 
phan, as determined by negative response to the glyoxylic-sulfuric 
acid test. The digestive periods differed in length. At the end 
of each, the mixture was neutralized, reinforced with neutral salts, 
and then made alkaline to one of several degrees of alkalinity. 
After sterilization in an autoclave, the mixtures were inoculated 
with mixed fecal bacteria from the stools of an individual on a 
mixed diet. No bacteria were isolated for purposes of Identifica- 
tion. The organisms were allowed to develop in the digestion 
mixtures at 37° C. for periods of different length. The reaction 
mixtures, after neutralizing and making them alkaline with a known 
amount of alkali, were distilled with steam until about 700 c.c. of 
distillate had been obtained. The distillate was diluted to 1,000 c.c. 
and an aliquot portion was treated with 0.2 per cent. sodium nitrite 
and conc. sulfuric acid solutions. A control Solution containing 
0.25 per cent. of indol (Kahlbaum) was treated in the same manner. 
Each nitroso-indol Solution was then allowed to stand until the 
maximum color developed.^ We used the Wolf colorimeter for 
the tinctorial comparisons, and found that even with the small 
amounts of indol obtained (see figures later) an error of 3 per cent. 
was very easily detected by difference in the intensity of the resulting 
colorations. 

We experienced some difficulty, at first, in mixing definite 
amounts of the indol Solution with the nitrite and sulfuric acid Solu- 
tions, and water, which would give uniform shade and intensity of 
color. Later we obtained very constant results by taking an aliquot 
portion of the indol Solution, adding the nitrite Solution and enough 
water to fill the cylinder of the apparatus almost to the 100 mark, 
then adding the conc. sulfuric acid Solution and sufficient water to 
fill to the mark. On mixing uniformly, a very faint though distinct 

' Moraczewski : Zeit. f. physiol. Chem., 1908, Iv, pp. 42-47; Chem. Ahstr., 
1908, ii, p. 2578. (Colorimetric determination of indol in feces. The abstract 
of the original article contains an error that should be corrected: the sodium 
nitrite Solution has a concentration of 0.2 per cent. instead of 2.0 per cent. 
See also, Levene and Rouiller : Jour. of Biol. Chem., 1906-7, ii, p. 481. A 
bromine-tryptophan colorimetric method for the determination of tryptophan.) 



37^ Determination of Tryptophan [April 

color developed which reached its maximum intensity in about 
half an hour. 

Details of the experiments. About 500 c.c. of skimmed milk 
were diluted in a precipitation jar with five volumes of water. The 
casein was precipitated by the addition of 12.5 c.c. of 10 per cent. 
acetic acid Solution. The casein was repeatedly washed with water 
by decantation and then dissolved in Standard sodium hydroxid Solu- 
tion (enough to dissolve the casein without leaving a large excess of 
alkali). The liquid required about 150 c.c. of n/2 sodium hydroxid 
Solution to produce a permanent alkalinity, using azolitmin paper as 
indicator. After dilution to a definite volume and filtration, two 
nitrogen determinations were made by the Kjeldahl method. It was 
found that each 100 c.c. of the Solution contained 0.8755 ö"^- o^ 
casein. Of the remaining Solution 500 c.c, were neutralized with 
phenolthalein as the indicator and treated with 0.4 gm. of sodium 
carbonate for each 100 c.c. volume of the neutral liquid. Then 
25 c.c. of a saturated pancreatin (commercial) Solution and xylene, 
as a preservative, were added. Incubation was continued at 2)7° C. 
for 24 hours, when an equal portion of the pancreatin Solution was 
added; a third portion was added at the end of 48 hours. The 
incubation was then continued for forty-four days. Steam was 
passed through the alkaline Solution to remove the xylene. The 
digestion was apparently complete ; common tests for tryptophan 
indicated its presence. The mixture was neutralized and reinforced 
with neutral salts, such as Hopkins and Cole used in their work — 5 
gm. Rochelle salt, 0.2 gm. ammonium phosphate and o.i gm. 
magnesium sulfate, per liter. No gelatin was added. The total 
volume was now made up to one liter and divided into four equal 
portions. Each portion contained cleavage products corresponding 
to 1.0944 gm. of casein. 

Portion A was sterilized in an autoclave and inoculated with a 
24 hour slant agar growth of intestinal bacteria.^ Portion B was 

• The method of inoculation was as follows : The bacteria were grown first 
in ordinary broth inoculated from feces. After 24 hours, agar slants were made 
in the usual way. When these were 24 hours old, the organisms were detached 
by means of sterile water and a sterile wire, and the liquid containing them was 
poured directly into the flask to be inoculated. In order to establish a definite 
degree of alkalinity, the digestion mixtures, after steam distillation, were 



I9I3] Jcsse A. Sanders and Clarence E. May 377 

treated with sodium carbonate (0.4 gm. per 100 c.c), sterilized and 
inoculated with some o£ the same 24 hour growth of bacteria. 
Portion C was made alkaline with sodium carbonate to 0.8 per cent. ; 
portion jD to i.o per cent. Both were inoculated as in A and B. 
After four days of incubation, the four flasks were reinoculated 
with fresh 24 hour cuhures of the bacteria and again incubated. 
After eight days' incubation, flask A was removed from the oven. 
Active indol-producing bacteria were present in the mixture. The 
original putrefaction mixture, neutral to litmus and giving a faint 
odor of indol, was made alkaline with sodium carbonate (to 0.4 per 
cent.). With steam distillation, all the indol passed into the first 
300 c.c. of distillate. 

Determination of indol. A Standard indol Solution was made 
by dissolving 0.25 gm. of the pure substance in a liter of water. 
Twenty-five c.c. of the original distillate from flask A, diluted to 
about 90 c.c, were treated with 10 drops of a 0.2 per cent. sodium 
nitrite Solution, and six drops of conc. sulfuric acid Solution, diluted 
to 100 c.c, and mixed uniformly. The liquid was allowed to stand 
until the maximum rose-red color of the nitroso-indol developed, 
when it showed the same intensity of color as that produced by 1.3 
c.c. of the Standard indol Solution diluted in the same manner to 
100 c.c The total distillate contained 3.9 mg. of indol. In flask 
B, incubation was continued for nine days after the second inocula- 
tion. At the end of that time, indol-producing bacteria were still 
active. The reaction-mixture being distinctly alkaline, no alkali was 
added prior to steam distillation. All the indol appeared in the 
first 650 c.c of distillate, which was diluted to 700 c.c. and thor- 
oughly mixed. Of this Solution 50 c.c. contained as much indol as 
1.2 c.c. of the Standard Solution. The total indol content of flask 
B was 4.2 mg. Flask C was incubated twenty-six days. The 
mixture smelled strongly of indol and was alkaline in reaction. No 
additional alkali was added. A distillate of 1000 c.c. was obtained, 
each 50 c.c. of which contained as much indol as 2.05 c.c. of the 
Standard indol Solution. The total content of indol in this putre- 
faction mixture was 10.25 mg. Flask D was incubated twenty-five 

titrated with n/io hydrochloric acid Solution and then neutralized quantitatively. 
The required weight of sodium carbonate was then added to the neutral Solu- 
tion to give the desired alkalinity. 



378 



Determination of Tryptophan 



[April 



days. It contained indol and was alkaline in reaction. Of i liter 
of distillate obtained by steam distillation, each 50 c.c. contained the 
quantity of indol present in 2.0 c.c. of the Standard Solution, in- 
dicating that the indol in the putrefactive mixture amounted to lo.o 
mg. A summary of the analytic data is appended. 



Flask 


Casein, gm. 


Digestion, days 


Bacterial action, days 


Indol, mg. 


A 
B 
C 

D 


1.0944 
1.0944 
1.0944 
1.0944 


44 
44 

44 
44 


8 

13 
26 

25 


3-9 

4.2 

10.25 

10.00 



The yield of indol could be derived from 1.7872 gm. of trypto- 
phan in the case of sample C and from 1.7436 gm. of tryptophan in 
the case of sample D. The weight of casein corresponding to the 
amount of indol found must have yielded the calculated weight of 
tryptophan. This being the case, 100 gm. of casein yield either 
1-593 g"^- of tryptophan (C) or 1.633 g"^- of tryptophan {D), as 
the minimum amounts. 

Hopkins and Cole claim that intestinal bacteria form small 
amounts of indol-acetic acid and other indol-containing substances, 
but we have not found these in our putrefactive mixtures, although 
they may have been formed in very small amounts, f or which reason 
we give the two results as indicating the minimum amounts of indol- 
yielding radicals in casein. Our results are as high as those ob- 
tained by other investigators. It is likely, of course, that the 
method will be improved by the further study we hope to give it, 
especially in its application to other common proteins. The present 
paper presents only preliminary results. 

The method, as outlined, is slow but it promises to be a satis- 
factory process for the determination of one of the cleavage 
products of protein material that hitherto has been difficult to de- 
termine quantitatively. 



PHYSICAL CHEMISTRY OF MUSCLE PLASMA^ 

FILIPPO BOTTAZZI 

(Physiological Institute, University of Naples, Italy) 

My experiments have been made on striated muscles of oxen, 
dogs, Scyüium stellare and Dentex vulgaris, and on piain muscles 
{M. retractor penis) of oxen. In the case of the dogs, the muscles 
were removed after flushing the blood vessels with 0.9 per cent. 
Solution of sodium chlorid (sometimes cooled to 4-5° C). In 
nearly all cases the muscles were preserved in dry vessels at low 
temperatures. They were freed from fatty and connective tissues, 
then minced, thoroughly pounded with quartz sand and infusorial 
earth, and plasma obtained in a Buchner press, generally at a 
maximum pressure of about 350 atmospheres. In some experi- 
ments the irritability of the animal (Scyllium) was abolished by 
gradually cooling it to about — 2° C, so that on cutting off the 
body musculature no contraction ensued. The muscle plasma 
(about 600-800 c.c.) was collected in dry vessels, centrifuged for 
an hour and preserved in a refrigerator. 

The plasma of striated mammalian muscle was always deep red 
in color and rather turbid; that of fish muscle was less colored. 
The plasma of piain muscle was always opalescent and almost color- 
less. The microscopic examination, made with powerful apochro- 
matic objectives, revealed no trace of morphologic Clements or 
granules in the centrifuged plasma, which always appeared to be 
perfectly homogeneous. But ultramicroscopic examination revealed 
the presence of innumerable very small and highly brilliant granules, 
mixed with a relatively small number of coarse particles, which have 
nothing to do with the granules, being composed of fat, glycogen 
and nuclear or sarcoplasmic f ragments. The existence of the ultra- 

^ Presented at the eighty-first meeting of the British Association for the 
Advancement of Science, in Dundee, September, 1912. In these researches I 
was aided by my assistant Dr. G. Quagliariello. 

379 



380 PJiysical Chemistry of Muscle Plasma [April 

microscopic granules was never observed before; this is the most 
important result of my investigations. 

The number or concentration of the granules in the original 
plasma is so great that the iiltramicroscopic field appears almost 
uniformly luminous — the individual granules cannot be distinctly 
Seen. But when the plasma is diluted with Ringer Solution, the 
granules are separated, and then appear as distinct brilliant cor- 
puscles endowed with lively Brownian movements on a darkish 
homogeneous background. They are not precipitation-particles of 
a dissolved muscle protein, because they do not disappear under the 
action of dilute alkali. Precipitation of such protein might be 
caused by lactic acid produced in the muscles, but in that case the 
particles would be dissolved by alkali — we do not know of any acid- 
precipitated proteins that are not resoluble in alkalies. No ordinary 
reagent causes the granules to disappear at a low temperature. 
Moreover, acid increascs the number of particles, by precipitating a 
special dissolved muscle protein. 

The granules are present in almost equal number in plasmas ex- 
pressed from muscles which have been cooled to a low degree and 
which, therefore, are non-irritant, t. e., from muscles in which acid 
production is greatly diminished. The concentration of the granules 
is greater in the plasma of striated muscle than in that of piain 
muscle. 

Normal muscle plasma is, then, a Suspension of ultramicroscopic 
granules in a liquid zvhich, besides containing mineral salts and ex- 
tractives, certainly holds protein in a state of true Solution. Ac- 
cordingly, the plasma, f reed from the granules, is an optically homo- 
geneous fluid, but on adding to it a weak acid Solution, or on 
heating it at 55° C, additional particles appear — true precipitation- 
particles of a dissolved muscle protein, which may be termed myo- 
protein, while we may give the name myosin to the protein of which 
the plasma granules are made. The existence of other muscle pro- 
teins in the muscle plasma has not been proved. 

The observed plasma granules are apparently a degradation or 
cleavage product of material in the myofibrils, piain or striated, and 
preexist in all muscle plasma. Their preexistence is not only easily 
conceivable, but we are also obliged to assume it, when we reflect 



1913] Filippo Bottassi 381 

that both materlals — fibrillar and sarcoplasmic — exist in muscle 
fibres in two distinct phases, which of course must also remain 
distinct in the plasma. The fibrillar phase being represented in the 
plasma in the form of granules, these are probably constituent ele- 
ments of the fibrils, in harmony with the views of Heidenhain. 

The granulär material tends to flocculate spontaneously ; but 
spontaneous agglutination and Sedimentation of the granules occurs 
very slowly, because of their smallness and the high viscosity of the 
Suspension fluid. Dilution with water, or with neutral, faintly acid 
or alkaline Solutions, dialysis, or heating to about 30° C, accelerates 
the process; but it also occurs, in from about 12 to 24 hours, as I 
have Said, when all accelerating action of physical or chemical 
agents is excluded. This aggregation, followed by precipitation, of 
the granulär material is essentially the so-called "spontaneous 
coagidation" of muscle plasma or extract; it is therefore neither an 
enzymic-coagulation nor a heat-coagulation of dissolved protein. I 
have never observed phenomena like those described by Kühne — of 
nearly instantaneous clotting of cold muscle plasma when raised to 
room temperature. 

Precipitation of the granules is greatly accelerated by heating 
muscle plasma to between 38° and 54° C, when, after a few 
minutes, a heavy precipitate is produced, from which there separates 
a clear yellowish-red fluid, muscle serum. This phenomenon, which 
many authors Interpret as one of heat coagulation of a dissolved 
protein, is, on the contrary, the effect of rapid aggregation and pre- 
cipitation of the suspended granules. When their concentration is 
very great, massive clotting of the plasma occurs. 

The precipitate which appears during the first 24-48 hours of 
dialysis of plasma is composed of the granulär material, and is not 
formed from a dissolved protein. Sometimes the plasma trans- 
forms itself into something like a blood coagulum. 

Heat-coagulation of dissolved myoprotein is a continuous 
process, which does not appear to be complete even at 80° C. As 
we cannot deny that it begins at a temperature as low as 50° C, 
we are bound to admit that the precipitate of granules, formed at 
54°-55° C, probably also contains a little myoprotein. In Opposi- 
tion to von Fürth and others, I have observed that the (dissolved) 



382 



Physical Chemistry of Muscle Plasma 



[April 



myoprotein is totally precipitated by strong and prolonged dialysis. 
As this precipitation process is also a continuous one, I do not 
deny that it begins during the first 24-48 hours; therefore the pre- 
cipitate of granules which forms early during dialysis may also 
contain a little myoprotein. 

A trace of dissolved protein can always be found in plasma 
wiiich has been dialyzed continuously for several months. It is 
probably serumalbumin, which cannot be wholly eliminated. 

Muscle pigments (hemoglobin, MacMunn's myohematin) are 
partly removed from the plasma by the agglutinated granules — ad- 
sorbed by them, and are also precipitated in some degree with myo- 
protein by prolonged dialysis. 

The granules and myoprotein, freed from electrolytes by suffi- 
ciently long dialysis, move toward the anode, when put under the 
influence of a strong electric current; they carry electronegative 
charges. 

For muscle plasma I have determined the quantity of plasma as 
per Cent, of fresh muscle, the total solid and total protein contents 
and ash yield, the specific gravi ty, lowering of the freezing point, 
electric conductivity, viscosity, surface tension and chemical reac- 
tion. Tables 1-3 contain the results of these determinations. 

TABLE I 

General data pertaining to muscle and muscle plasma 





Plasma 
No. 


Muscle 


Plasma 


Muscle 


Weight, 
kg. 


Pressure, 
atm. 


Weight, 
gm. 


Per Cent, of muscle 


Scyllium: Striated 
Ox: Striated 
Dentex: Striated 
Ox: Piain 
Dog: Striated 
Ox: Piain 


7 
8 

9 

5 

II 

12 


1.067 
0.938 
0.894 
I.184 
0.829 
I.190 


350 
350 
50 
350 
350 
50-350 


666 
592 
217 
580 

339 
562 


62 
63 
24 
50 
40 
47 



The plasma varied in quantity from 40 to 63 per cent. (volume), 
for pressures which never exceeded 350 atmospheres. The dry 
residue from piain muscle plasma was less than that from striated 
muscle plasma. The total protein content was relatively low — less 
than that of blood serum. Muscle plasmas are very rieh in their 



I9I3] 



Filippo Bottazzi 



383 



TABLE 2 



Percentage data for water, total solid and total protein contents, and ash yield, 

of muscle plasma 



A. Piain muscle 



Muscle plasma 


Water 


Total solids 


Total protein 


Ash 


Organic solids 
mious protein 


3:Ox 
4: Ox 
5:Ox 
6:Ox 

12: Mixed — 

Fraction a 
Fraction b 
Fraction c 


93-14 
97-57 
93.808 
94.126 

93-27 
92.76 
93-60 
93-99 


6.86 

6-43 
6.192 

5-874 

6.73 

7-24 

6.40 

6.01 


3-63 
3-37 
3-iS 
2.75 
3-37 
4-13 
2.90 
2.67 


1.32 
1-30 
I.148 

1-37 
1.30 


I.91 

1.76 

1.894 

1-754 

2.06 





















B. Striated muscle 






2: Ox 


91.086 


8.914 


4-53 


1-739 


2.64s 


8 


Ox 


92.57 


7-43 


3.65 


0.85 


2.93 


9 


Dentex 


83-90 


16.10 


4.10 


1.82 


3-i8 


7 


Scyllium 
Scyllium 


90.36 


9-64 


3-04 
3-36 


1.80 


4.80 










loa: Scyllium 






2.38 










1 1 : Dog 


87-37 


12.63 


3.85 


1.50 


7.28 



TABLE 3 

Data pertaining to physico-chemical properties of muscle plasma 

A. Piain muscle 



6 






Lower- 






Surface 








Muscle 


Specific 


ing of 
the 


Electric 
conduc- 


Viscos- 


tension 
at25° 


Chem- 
ical 


Remarks regarding 


¥n 


gravity 


freezing 


tivity 


ity 

(P 25°) 


reaction 


the plasma 


CS 






point 
(A) 


(K180) 


(Ch-io7) 




3 


Ox 


1.026 

















Normal, fresh 


4 


Ox 


1.024 


0.806° 


— 


— 


— 


— 


Normal, fresh 


5 


Ox 


1-023 


0.761° 


— 


4-13 


73-15 


7.44 


Normal, fresh 


6 


Ox 


I.02I 


0.730° 


0.0144 


3-04 


76.76 


6.18 


Normal, fresh 


12 


Mixed 


1.024 


0.804° 


— 


4-48 


73-34 


7.30 






Fraction a 


1.026 


0.812° 


— 


5-98 


69-34 


— 


Expressed at 50 atm. 




Fraction b 


1.025 


0.812° 


— 


3-68 


74.67 


— 


Expressed at 200 atm. 




Fraction c 


1.023 


0.810° 


— 


2.96 


76.27 


— 


Expressed at 350 atm. 



Striated muscle 



7 


Scyllium 


1.027 


2.455° 


0.0147 


1-59 





II. I 


Muscles were not cooled 


8 


Ox 


1.027 


0.868° 


0.0107 


1.75 


78.19 


31.4 


Normal conditions 


9 


Dentex 


1.049 


1.196° 


0.0120 


2.82 


76.22 


12.S 


From muscles of cooled 
animal 


10 


Scyllium 
Scyllium: 
Frac. loa 


1.024 
1.024 


2.337° 

2.494° 


0.0149 
0.0150 


3.71 
1.71 


72.9 
73.1 


S.65I 
6.50 j 


From three Scyllia (cooled 
to from -2° to -3° C.) 


II 


Dog 


1.039 


1.088° 


— 


2.24 


68.90 


10.6 


Three hours after expres- 




Dog (i) 


1.037 


1.016° 


— 


— 


70.00 


35.0 


Data obtained the fol- 
lowing day 



384 Physical Chemistry of Miiscle Plasma [April 

yield of ash and content of organic non-protein substances. Gen- 
erally, the dry residue and the protein contents are inversely pro- 
portional to the pressure at which the plasma is expressed. Since 
the specific gravity (1,021-1,027, as a rule) is quite near that of 
blood serum, I believe the salts and extractives, lipoids, glycogen, 
etc., help to accoimt for it. 

The osmotic pressure is always very high (A =0.730-1.088° 
C. for mammals; A =2.337-2.494° C. ior Scyllium; A = 1.196° 
C. for Dcntex), higher than that of the blood. 

The reaction is always acid (Ch- 10'^ = 5.65-12.5 ; but there are 
also higher values : ChIO^=3I-4 f- e.). Acidity is lower in 
plasma of piain muscle and of cooled striated muscle, higher in 
the plasma of striated mammalian muscle. As a rule, the hydrogen- 
ion concentration increases with time; but as this augmentation is 
rather feeble, I believe, with Fletcher, that the maximum production 
of acid substances occurs in muscles soon after their Separation from 
the body. 

The high osmotic pressure of muscle plasma is probably due 
mainly to the substances that determine the acid reaction. 

The low electric conductivity (Kjso = 0.0107-0.0144 for mam- 
mals; 0.0147-0.0150 for Scyllium; 0.0120 for Dentex) and high 
viscosity (p25o= 1.59-2.82 ; 3.04-5.98 for piain mammalian muscle) 
of the plasmas are explained by their corpuscular composition. But 
the very high viscosity of the plasma of piain muscle is probably 
caused by some particular protein derived from the connective 
tissue. 

Zw 

The surface tension (100-^=68.90-78.19) is generally higher 

than that of the blood serum. 

My results are, for the most part, in Opposition to those obtained 
by previous authors. But the new interpretation of phenomena 
like those of spontaneous coagulation and heat coagulation, etc., 
was suggested to me mainly by the granulär Constitution of muscle 
plasma. As I stated above, this is the most important result of my 
investigations, a result which hereafter must be recognized by all 
who study problems pertaining to the chemistry and physical chem- 
istry of the Contents of the muscle fiber. 



1913] Filippo Bottazzi 3^5 

I have endeavored to corroborate my hypothesis, that the 
granules are disintegrated fibrillar material, by trying to stain the 
granules with some pigment which would selectively color the muscle 
fibrils ; and also by attempting to show that the granules possessed 
double refractive power. My attempts have been unsuccessful, 
however, although this was not unexpected, because of the ultra- 
microscopic dimensions of the granules. 



FASTING STUDIES 
II. A note on the composition of muscle from fasting dogs^ 

H. C. BIDDLE AND PAUL E. HOWE 

(Laboratory of Physiological Cheniistry, University of Illinois, 

Urhana, Illinois) 

The variations in the composition of the different forms of 
muscle in the normal individual have received considerable atten- 
tion. A comparison of the nitrogen and moisture contents of heart 
and striated muscle reveals a lower percentage of nitrogen and a 
higher percentage of moisture in the heart muscle than in the skeletal 
muscle. The proportions of these and other constituents in the 
fasting muscle have not been studied extensively. The changes 
which occur in the composition of muscle during fasting are 
significant for the Solution of general problems relating to the 
effects of fasting. 

In a study of the influence of fasting upon the creatine content 
of dog muscle, determinations were made of the proportions of 
nitrogen, moisture, fat and creatine in normal and fasting muscle ; 
and also of the nitrogen and creatine in heart muscle. In these 
preliminary experiments particular attention was paid to the per- 
centages of nitrogen and creatine : an attempt was made to show the 
relation between the nitrogen and creatine contents of muscle, so 
that the ratio of creatine-nitrogen to total nitrogen might be used as 
an index of the changes due to pathological conditions. This factor 
should be more significant than the percentage of creatine in fresh 
muscle, and as accurate as that for the creatine content in muscle 
on a fat- and moisture-free basis. These results, as well as the 
variations in the creatine content of muscle, will be discussed in a 
later paper.^ 

^ Presented before the Columbia University Biochemical Association, 
December 6, 1912; Biochemical Bulletin, 1913, ii, p. 288. 

^Howe and Hawk: Presented before the recent annual meeting of the 
American Physiological Society, but not abstracted in the proceedings. 

386 



1913] H. C. Biddle and Paul E. Howe 387 

The procedure employed in determining some variations due to 
fasting was as follows : muscles (M. sentit endinosus and M. hiceps 
femoris) of normal animals were analyzed for moisture, fat, phos- 
phorus and creatine ; and, in one case, the heart was analyzed for 
nitrogen and creatine. 

Muscles of the same kind were removed aseptically from normal 
dogs under ether anesthesia^ and analyzed for nitrogen and creatine. 
After a prolonged fast the corresponding muscles of the opposite 
legs of the "operated" dogs, as well as the remaining muscles on 
the same side, were analyzed for total nitrogen and creatine, and 
in two cases for moisture and fat. With such a procedure the 
changes which resulted from fasting were studied on the same 
individual, also in fasted animals as compared with different normal 
controls. The "operated" dogs recovered readily and the wounds 
healed rapidly even when the fast was begun immediately after the 
Operation. 

The analytical methods employed were as follows: Total 
nitrogen was determined by the Kjeldahl process ; moisture by dry- 
ing in a vacuum over sulfuric acid at room temperature ; fat by the 
ether-extraction process, and creatine by the Folin procedure as 
modified for meat by Emmett and Grindley.* The creatine was 
extracted according to the methods of Grindley and Woods^ and 
of Mellanby.^ 

The table on page 388 contains the more significant data. 

A consideration of this data shows an increase in the percentage 
of moisture, and a decrease in the percentages of nitrogen and crea- 
tine, in the striated muscle as a result of fasting. From the data 
on a Single normal heart and a single fasting heart there appears 
to have been a decrease in the nitrogen and an increase in the 
creatine content of the fasting heart. We also note that the 
changes which take place in the same fasting individual, as con- 
trasted with the fasting changes when compared with different con- 
trol animals, are approximately the same. 

'We wish to thank Dr. O. O. Stanley, of the University of Illinois, and 
Dr. C. T. Moss, of the Michael Reese Hospital, of Chicago, Illinois, for their 
aid in the removal of the muscles. 

* Emmett and Grindley: Jour. of Biol. Chem., igoy, iii, p. 491. 

^ Grindley and Woods : Ibid., igoö-'o;, ü, p. 309. 

' Mellanby : Jour. of PhysioL, 1908, xxxvi, p. 453. 



388 Fasting Studies [April 

Data pertainlng to the composition of muscle front fasting dogs 



Dog 


Kind of muscle 


Moisture, 
per Cent. 


Nitrogen, 
per Cent. 


Fat, per cent. 


Creatine, 
per Cent. 


E. 
D. 


Normal leg 

Normal leg 
Normal heart 

Normal leg 
Fasting leg 
Fasting heart' 

Fasting legi» 

Normal leg 
Fasting leg" 


73-4 
73-4 


3-42 

3-51 
2-95 

3-34 

2.82 

2.6s 
2.95 

3-99 
3.6l 


2.4^ 
2.28 

0.56 


0.31 
0.33 


B. 




0.23 


81.2 


0-34 
0.31 


C. 

A. 


81.8 


0.49 


0.19 












0-39 



The increase in the moisture content of fasting muscle may be 
associated, in part, with the decrease in the fat content; in normal 
animals there is a decrease in the percentage of moisture associated 
with an increase in the fat content of muscle.^^ The increase in 
the moisture content of fasting muscle may also be due to changes 
in the colloidal State or the molecular condition of the cellular con- 
stituents. This increase in moisture is more significant when we 
consider that there is apparently a greater decrease in the cytoplasm 
than in the nuclei of the cells as a result of a fast;^^ the nucleus 
and the connective tissue, the substances which would then pre- 
dominate, normally contain less water than the cytoplasm. The 
increase in the moisture content of fasting muscle has been noted 
by other investigators. 

The lower absolute nitrogen content of fasting muscle, when 
considered on the basis of fresh muscle, becomes an increased rela- 
tive nitrogen content, when the values for nitrogen are calculated 

^Content of phosphorus = o.i6 per cent. 

'Content of phosphorus = 0.21 per cent. 

'A sixty-four day fast which resulted in 
original weight. The animal received 320 c.c. 

^'A twenty-one day fast. 

" A fifteen day fast, which resulted in a loss of 38 per cent. of the original 
weight. The animal did not receive water. 

" This f act, together with certain other deductions, has been corroborated 
by data in a personal communication from Professor P. F. Trowbridge of the 
University of Missouri. 

"Morgulis: Archiv für Entwicklungsmechanik der Organismen, 1911, xxxii, 
p. 169. 



a loss of 62 per cent. 
of water daily. 



of the 



1913] H. C. Biddle and Paul E. Howe 389 

on a moistiire- and fat-free basis. An alteratlon in the direction of 
increased nitrogen content is more in harmony with the changes 
that actually take place in the muscle. The relatively greater de- 
crease in the volume of the cytoplasm than of the nucleus, and the 
apparent relative increase in the connective tissue, are modifications 
in the direction of a higher nitrogen content of the muscle. The 
proteins which predominate in the connective tissue and in the nuclei 
of muscle contain higher percentages of nitrogen than do the pro- 
teins which make up the major portion of the cytoplasm of the 
cells. 

In addition to changes in the chemical nature of fasting muscle, 
certain physical modifications arise: normal muscle is firm to the 
touch and, when hashed, may be readily handled without sticking to 
the fingers ; fasting muscle, on the other hand, is soft to the touch 
and, when hashed, adheres tenaciously to the fingers. 

While the lowered percentage of nitrogen in the fresh skeletal 
muscle, as a result of fasting, may be due to diminutions in the 
Contents of moisture and fat, the difference in the composition of 
heart and skeletal muscle cannot be explained in this way. If the 
nitrogen content of normal striated and heart muscle be calculated to 
the moisture- and fat-free basis, there still remains a greater per- 
centage of nitrogen in the striated muscle. That there are differ- 
ences in the relative proportions of the soluble protein and the 
stroma in heart, and in striated and smooth muscle, has been shown 
by Saxl,^'* who finds that seven-eighthsof the skeletal muscle consists 
of soluble proteins while but one-third of the heart muscle is of this 
nature. 

A careful differential study of the proteins of fasting muscle 
should throw some light upon the nature of the disintegrative 
processes which take place in the tissues as a result of fasting. 
Such a study is contemplated. 

" Saxl : Beiträge s. ehem. Physiol. u. PathoL, 1907, ix, p. i. 



SOME NOTES ON THE FORM OF THE CURVE OF 

CARBON-DIOXIDE EXCRETION RESULTING 

FROM MUSCULAR WORK FOLLOWING 

FORCED BREATHING^ 

G. O. HIGLEY 
(Ohio Wesleyan University, Delaware, Ohio) 

(WITH PLATE 4) 

In an earlier research^ an attempt was made to determine how 
soon after the beginning of work an increase in the production of 
carbon dioxide begins to show itself in the expired air. This time 
(the latent period) was at first found to vary from three to fourteen 
seconds. Now, clearly, such periods are not long enough to cor- 
respond with the time required for the carbon dioxide formed in the 
muscles at the first muscular contraction to reach the outside air. 
It must first diffuse into the blood from the tissues where it is 
formed, then traverse the venous half of the systemic circulation, 
the right side of the heart, and the arterial half of the pulmonary 
circulation, and finally diffuse into the air of the alveoli before any 
of it can appear in the breath. From the conclusions of Stewart 
and others it appeared that from fifteen to twenty seconds is the 
least possible time required for the blood to traverse this distance, 
to say nothing of the diffusion time. It was finally found that 
the sudden increase in the rate of excretion of carbon dioxide, after 
the beginning of work, was due primarily to a better Ventilation of 
the lungs, while the continuation of the increase was due to the 
Ventilation of the blood and tissues as well. A recognition of this 
fact led to the following modification of the method for the de- 
termination of the latent period of carbon dioxide excretion : 

^ This paper was accepted for publication by the officers of Section VIII, 
d. Eighth International Congress of Applied Chemistry, and was read before 
the Section at a stated meeting on September 11, 1912; Biochemical Bulletin, 
1912, ii, p. 153. 

^Higley and Bowen: American Journal of Physiology, 1904, xii, p. 311. 

390 



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1913] G- O. Higley 391 

After the " normal " rate o£ excretion at rest had been de- 
termined, the subject began forced breathing at a predetermined 
rate, continuing this for a minute or so until the curve of carbon 
dioxide excretion had apparently assumed its permanent direction. 
At this point, at a signal from the experimenter, the subject began 
to drive the bicycle as in the preceding experiments. The effect was 
marked, the new rate of excretion being sharply defined from the 
normal rate preceding it. The further increase in the rate of excre- 
tion, after the beginning of work, was now not so prompt in its 
appearance, and came on more gradually, reaching its maximum 
after a minute or so, depending on the work. 

In these experiments the latent period of increase due to work 
was from seventeen to twenty-two seconds. It is evident that as 
the latent period will vary with the rapidity of the circulation, the 
rapidity of diffusion, and the rate of work, a more definite figure 
was not to be expected. 

Shortly after the publication of these results by Bowen and the 
writer, a communication was received from Prof. N. Zuntz, calling 
attention to the gradual character of the change in rate of excretion 
of carbon dioxide after the beginning of work (as already men- 
tioned) and kindly suggesting a modification of the method of 
carrying out the latent-period experiments. According to Prof. 
Zuntz, if the forced breathing were continued for five minutes 
instead of one minute, as already stated, the blood and tissues 
would become thoroughly ventilated; the direction of the curve of 
carbon dioxide would become parallel to that before forced breath- 
ing began ; and, furthermore, with the beginning of work, the carbon 
dioxide curve, after the latent period of twenty seconds, would 
change much more sharply than it did in the published record. The 
writer accordingly made a series of experiments in which the forced 
breathing was continued for from five to seven minutes before 
bicycle work has begun. 

The results of one of these experiments are seen in Plate 4 in 
which A is the Pneumograph record, Pqrs the carbon dioxide curve, 
T the Chronograph record, and M the bicycle record. The line 
Pq'q^'r, as in the previous paper,^ represents the rate of excretion 

* Higley and Bowen : Loc. cit. 



392 Curve of Carbon-dioxide Excretion [April 

before the beginning of forced breathing; the line Pqq'q"r (broken 
by the arresting of the beam and the addition of foiir gram weights) 
represents the curve of carbon dioxide during forced breathing; r 
is the Position on the curve of the carbon-dioxide-writing lever at 
the instant when work was begun ; and s is the point where the curve 
changes as a residt of the zvork. 

This research was conducted on two subjects. It was found 
very difficult to maintain respiration of uniform depth for five 
minutes, since there is a decided tendency to make the respiration 
shallower. Indeed, notwithstanding the great care on the part of 
the subject, the Pneumograph record indicated, in some cases, a 
lessened depth of respiration toward the end of the forced respira- 
tion period. 

In the case of one subject the curve for rate of excretion of 
carbon dioxide returned, during the period of forced respiration, 
practically to the original value. With the other subject the return 
was less perfect, It would seem that as a result of the additional 
work of the respiratory Organs a return of the rate of excretion to 
the value during normal respiration could not be expected. 

While, therefore, the writer is able to confirm Prof. Zuntz's pre- 
diction regarding the sharpness of the change, as a result of work, in 
the curve of carbon dioxide after continued forced respiration, he 
can confirm only in part Prof. Zuntz's prediction on the return of 
the curve, during forced respiration, to the direction which it had 
before forced respiration was begun. 

This work was done in the physiological laboratory of the Uni- 
versity of Michigan, Ann Arbor, Michigan. 



THE INFLUENCE OF BAROMETRIC PRESSURE ON 
CARBON-DIOXIDE EXCRETION IN MAN^ 

G. O. HIGLEY 
(Ohio Wesleyan University, Delaware, Ohio) 

(WITH PLATE 5) 

Introduction. This work was suggested by that of Lombard^ 
on " Some of the influences which affect the power of muscular 
contraction." In that research, which was made with the ergo- 
graph, Lombard found that, in general, there was a fall of muscular 
power during the day, this result being noted on eighteen out of a 
series of twenty-three days. However, on certain days, the fall in 
power due to fatigue was slight and on five days the power was 
greater at the last experiment than at the first. These exceptions 
led to the suspicion that barometric changes had an influence on 
muscular endurance. When, later, a comparison was made between 
Lombard's endurance curve and the curve of barometric height, it 
was found that, while no constant relationship existed between the 
two variables, they varied in the same sense on twenty out of twenty- 
three days ; i. e., in general " when the barometer rose during the 
day, or feil less than on the preceding day, the muscular endurance 
either rose, or feil less than on the preceding day." 

It has been shown, furthermore, that while a diminution of 
barometric pressure increases both the respiration rate and the 
volume of air respired, after allowance is made for the increase of 
volume due to the lower pressure the volume respired is less 
(Speck). 

Now, the effect of increasing barometric pressure upon the power 
of the muscular System might possibly be due to some influence 

^ This paper was accepted for publication by the officers of Section VIII, 
d, Eighth International Congress of Applied Chemistry, and was read before the 
Section at a stated meeting on September 11, 1912; Biochemical Bulletin, 
1912, ii, p. 153. 

^ Lombard : Journal of Physiology, 1892, xiii, p. i. 

393 



394 



Carhon-dioxide Excretion in Man 



[April 



exerted through the nervous and circulatory Systems tending to 
increase the readiness of metabolism; if such were the case then a 
Variation in barometric height should be accompanied by a Variation, 
in the same sense, in the rate of excretion of carbon dioxide. 

Plan of the experiments. It seemed that a series of experi- 
ments carried out for a month on three healthy subjects might throw 
light on this question, and also give interesting results as regards the 
effect of other conditions on the rate of carbon dioxide excretion. 
A series of respiration experiments was planned, accordingly, for 
three subjects, A and B, students in the University of Michigan, 
and the writer, C. A and B were 24 and 22 years of age re- 
spectively, and weighed, without clothing, 158 and 159/^ pounds. 
C was 46 years of age and weighed, exclusive of clothing, 148 
pounds. Each subject was to live his regulär daily life except that 
no vigorous muscular exercise was to be engaged in immediately 
preceding any experiment and that nothing whatever was to be 
eaten between meals. The plan of work is indicated in the ap- 
pended summary, where the data for the third part of each experi- 
ment are placed below those for the first and second : 



Subject 


Hour 

of 
rising 


Reclined 


First 
experi- 
ment 


Breakfast 


Time until next 
experiment 


Reclined 


Second 

experiment 

begun 


Dinner 


A 
B 
C 


6 
6 
6 


6:4s 
7:05 
7:25 


7:00 
7:20 
7:40 


7:40-8:00 
8:00-8:20 
8:00-8:20 


4 hr. 
4 hr. 
4ihr. 


11:4s 
12:05 
12:25 


12:00 
12:20 
12:40 


12:40 
1:00 
1:00 




Subject 


Time until next 
experiment 


Reclined 


Third experiment 
begun 


Lunch 


Experi- 
menter 


A 
B 
C 


4 hrs. 
4 hrs. 
43 hrs. 


4:45 
5:00 

5:2s 


5:00 
5:20 
5:40 


5:40 
6:00 
6:00 


c 

A 
B 



The routine of work was as follows : The subjects rose at 6 
o'clock, reaching the laboratory at about 6:35. A reclined upon a 
couch at 6 :45 in preparation for the first experiment. B and C 
prepared all the apparatus, making the initial calibration of the 
balance, weighing the guard tubes, reading and recording the 
barometric height, the outdoor and room temperature, etc. In order 
to enable the experimenter to judge the better as to the physical 



I9I3] 



G. O. Higley 



395 



condition of each subject, mouth temperature and pulse were also 
taken and recorded. This routine at the laboratory was followed 
at 12 M. and 5 P. M. 



TABLE I 



Data showing the excretion of carbon dioxide by subjects A, B and C, 

in milligrams per minute. 



Date 


S 


ubject A 




Subject ß. 


Subject C. 


Dec. 


7 A. M. 


12 M. 


5 P. M. 


7 A. M. 


12 M. 


5 P.M. 


7 A. M. 


12 M. 


5 P. M. 


23 


406 


422 




381 


498 


567 


406 


390 


447 


24 


438 


460 


447 


489 


422 


541 


419 


409 


409 


26 


442 


448 


466 


SI4 


429 


(743) 


422 


403 


428 


27 


422 


453 


466 


535 


434 


548 


390 


403 


390 


28 


403 


448 


(635) 


488 


507 


498 


397 


375 


419 


29 


407 


422 


381 


520 


553 


546 


382 


387 


456 


30 


438 


483 


405 


529 


495 


518 


419 


381 


374 


31 


42 s 


470 


444 


(647) 


489 


476 


390 


362 


438 


Jan. 




















2 


433 


487 


480 


438 


(611) 


570 


393 


422 


473 


3 


470 


436 


422 


416 


462 


508 


394 


377 


422 


4 


507 


435 


480 


537 


442 


553 




386 


448 


S 


469 


473 


442 


528 


466 


515 


410 




396 


6 


458 


466 


442 




525 


531 


449 


403 


476 


7 


46s 


442 


432 


439 


560 


466 


406 


386 


411 


9 


416 


436 


436 


410 


442 


462 


390 


380 


448 


10 


416 


459 


448 


455 


506 




383 


42s 


473 


II 


456 


439 


426 


453 


422 


526 


363 


402 


337 


12 

T "7 


446 







543 






388 






13 
14 


405 


462 


445 




476 


504 


398 


427 




16 


436 


496 


449 


469 


531 


440 


402 


396 


437 


17 


412 


453 




418 


460 






459 




18 


377 


407 


462 




459 


469 


364 


442 


435 


19 


472 


487 


422 


445 


474 


409 


403 


438 


429 


20 


469 


476 


436 


402 


455 


432 


402 


474 


419 


21 


462 


436 


493 


399 


442 


493 


406 


448 


434 


23 


428 


481 


429 


509 


442 


436 


396 


449 


429 


24 


422 


517 


495 


500 


459 


537 


409 


460 


429 


25 


422 


475 


402 




528 


486 


422 


468 


428 


26 


495 


561 










422 


422 




Averages 


438 


462 


443 


472 


476 


501 


401 


414 


427 



Results. The results of this series of experiments are shown 
in Table i, in milligrams of carbon-dioxide excretion per minute. It 
will be noted that A's average for the midday experiments is con- 
siderbly higher than that for the morning and evening experiments. 



39^ Carbon-dioxide Excretion in Man [April 

This is due, in part at least, to the fact that this subject took his 
heartiest meal in the morning. The excretion of carbon dioxide 
for B and C, on the other band, was greatest in the evening, since 
these subjects took their dinner at i P. M. 

The remarkably high excretion shown for A at the evening ex- 
periment of December 28 (635 mg., while the average for that 
hour for this subject is only 443 mg.) is explained as follows: 
This subject went skating in the afternoon of that day and at about 
2 130 o'clock had the misfortune to break through the ice, becoming 
wet to the neck. On being rescued, he walked about two miles in 
his frozen clothing, exposed meanwhile to a strong wind at a 
temperature of about — 6° C. On reaching his room he took a 
thorough rubdown, made a change of clothing, rested for one and 
one half hours, and appeared at the laboratory at the usual hour for 
the experiments, with the result stated above. It will be noted that 
all of this subject's values for the following day, especially that of the 
evening, were much below the average, indicating a reaction from 
the exposure and excitement of the preceding day. The high ex- 
cretion of the morning of January 4 is supposed to be due to lunch 
eaten late on the preceding evening; that of 12 o'clock, January 26, 
to an exceptionally heavy morning meal ; and the low result of the 
evening of January 19 to an especially light midday meal. 

The irregularity of results obtained from B are somewhat diffi- 
cult to explain. Those of the morning of December 27, 30 and 31, 
were due to lunch eaten late the preceding evening and in the case 
of the two latter results, also in part to excessive haste to reach the 
laboratory in time for the regulär experiment. Other high results, 
especially those of 5 P. M., December 26, and of 12 M., January 
2, were undoubtedly due to Indigestion. 

Passing now to a study of the relation of carbon dioxide excre- 
tion to barometric changes, Plate 5 will be found to embody, in the 
form of curves, the results already given in Table i, with time as 
abscissae, and milligrams of carbon dioxide per minute as ordinates; 
it presents curves for A, B and C, together with that for the 
barometer in millimeters of mercury and of the outdoor temperature 
in degrees centigrade. The temperature of the room was practically 
constant throughout the series of experiments. Three curves are 



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I9I3] G. 0. Higley 397 

given for each subject where the necessary data were at band. In 
each case the morning, midday and evening ciirves are represented, 
respectively, by solid, long-dash and short-dash lines. 

Analysis of the results. Comparison of the data for haro- 
metric pressure and carhon dioxide excretion. Before proceeding 
to a rigorous mathematical investigation of the relationship between 
barometric change and carbon dioxide excretion, it seemed desirable 
to make a comparison of these two variables at a number of the 
dates on which especially marked barometric fluctuations took place, 
since in such cases the effect would be more pronounced and less 
likely to be masked by other varying conditions, such as amount 
and character of the preceding meal, character of muscular exercise, 
etc. To f acilitate such a comparison Table 2 was prepared ; it 
indicates experiment number; dates between which the comparison 
is made ; barometric height, rise or fall ; subject ; carbon dioxide for 
the two days between which comparison is made; rise or fall of 
excretion; and relation between barometric change and carbon 
dioxide excretion, whether direct or inverse. Taking first the 
morning values, it was found that the barometer rose between 7 

A. M., December 23, and 7 A. M., December 24, from 739 to 746, 
or 7 mm. During the same period the excretion of carbon dioxide 
of the three subjects changed as follows : That of A from 406 to 
438 mg. per minute, an increase of 32 mg.; that of B from 381 to 
489, an increase of 108 mg., and that of C from 406 to 419, an in- 
crease of 13 mg. per minute. Thus with rising barometer there 
was an increase in the rate of excretion of carbon dioxide in the 
case of each subject. A similar result is obtained in four other 
morning experiments (two subjects). In three morning experi- 
ments there are two direct results each. One experiment shows two 
indirect results, i. e., there is a change in carbon dioxide excretion 
which is opposite in sign to that in the barometer. 

Summing up the results of the morning experiments we have 
the following : Eleven experiments were carried out on A, seven on 

B, and eleven on C. The degree of correspondence of barometric 
change with carbon dioxide excretion was : 



A, 7 cases out of 11, or 63.6 per cent. 

B, 6 cases out of 7, or 85.7 per cent. 

C, 6 cases out of 11, or 55.5 per cent. 



398 



Carhoiirdioxide Excretion in Man 



[April 



TABLE 2 

Data obtained at 7 a.tn. 





















Relation between 




















barometric 






Barometer 






Excretion of carbon dioxide 


change and 


No. 


Date. 
Dec. 








Subject 








carbon dioxide 
excretion. 






Rise, 


Fall, 


Per minute, 


Rise, 


Fall, 










Heights, mm. 


mm. 


mm. 




mg. 


mg. 


mg. 


Direct 


Inverse 


I 


23-24 


739 -746 


7. 


— 


A 
B 
C 


406-438 

381-489 
406-419 


32 

108 

13 


— 


32 

108 

13 




2 


26-28 


745-I-72I 




24.1 


A 
B 
C 


442-403 
514-488 

422-397 


— 


39 
26 

25 


39 
26 

25 




3 


28-29 


721 -742.1 


21. 1 


' 


A 
B 


403-407 
488-520 


4 
32 


Z 


4 
32 














C 


397-382 


— 


15 


— 


15 


4 


29-31 


742.5-736 





6.1 


A 
B 
C 


407-425 


18 


— 


— 


18 












382-390 


8 


— 


— 


8 


5 


Jan. 






















2-4 


737-I-743-5 


6.4 




A 
B 
C 


433-507 
438-537 
393-394 


74 
99 

I 


— 


74 
99 

I 




6 


5-7 


743-5-732.5 




II. 


A 
B 
C 


469-465 
528-439 
4x0-406 


— 


4 

89 

4 


4 

89 

4 




7 


11-12 


753.9-737-1 





16.8 


A 


456-446 


— 


10 


10 














B 


453-543 


90 


— 


— 


90 












C 


363-388 


25 


— 


— 


25 


8 


12-14 


737.1-751-2 


I4.I 


— 


A 
B 
C 


446-405 


— 


41 


— 


41 












388-398 


10 


— 


10 




9 


18-19 


748.2-739.3 





8.9 


A 
B 
C 


377-472 


95 


— 


— 


95 












364-403 


39 


— 


— 


39 


10 


23-24 


749 -739.8 


'~^ 


9-2 


A 
B 


428-422 
509-500 





6 
9 


6 
9 














C 


396-409 


13 


— 


— 


13 


11 


24-26 


739.8-758.8 


19. 


— 


A 
B 
C 


422-495 


73 


— 


73 














409-422 


13 


— 


13 





The direct results from the midday experiments were as foUows 

A, 3 cases out of 7, or 42.8 per cent. 

B, 3 cases out of 9, or 33.3 per cent. 

C, 6 cases out of 9, or 66.6 per cent. 



From the evening experiments the direct results 



were 



A, 3 cases out of 6, or 50 per cent. 

B, 4 cases out of 9, or 44.4 per cent. 

C, 4 cases out of 8, or 50 per cent. 



I9I3] 



G. O. Higley 



399 



TABLE 3 

Data showing the relation of carhon dioxide excretion to barotnetric change, 

at noon for subject A. 



Barometer 
reading, mm. 



739 
746 
742 
722 
726 
742 
740 
736 
738 
742 

745 
743 
739 
732 
745 
753 
749 
753 
747 
748 
747 
740 
744 
743 
749 
738 
752 
758 



Carbon dioxide 
excretion, mg. 



422 
460 
448 
453 
448 
422 

483 
470 

487 
436 
435 
473 
466 
442 
436 
459 
439 
462 
496 

453 
407 
487 
476 
436 
481 
517 
475 
561 



X 



■ 4,2 
2.9 

- 0.8 
■21.2 
-16.4 

- 0,8 

■ 3-2 

- 7-2 

■ 4-4 

- 0.4 
1,8 
0.2 

- 3-4 
-10.4 

2-5 

9.9 
6.6 
10.3 
4.2 
4.8 
4.6 

- 3-2 
0.8 
0,6 
6.5 

- 4-5 
9.6 

I5-I 



-40 

- 2 
-14 

- 9 

-14 

-40 

21 

8 

25 
-26 

-27 
II 

4 
-20 
-26 

- 3 
-23 

o 

34 

- 9 
-55 

25 

14 

-26 

19 
55 
13 
99 



^•2 



17.64 

8,41 

0.64 

449.44 

268.96 

0.64 

10,24 

51,84 

19.36 

0,16 

3-24 

0,04 

11,56 

108.16 

6.25 

98.01 

36.00 

106.09 

17.64 

23.04 

21.16 

10.24 

0.64 

0.36 

42.25 

20.25 

92.16 

228.01 



1,642,43 



1-2 



1,600 

4 

196 

81 

196 

1,600 

441 

64 

625 

676 

729 

121 

16 

400 

676 

9 

529 

o 

1,156 

81 

3.025 

625 

196 

676 

361 

3.025 

169 

9,801 



Products {XV) 



27.078 



Negative Positive 



5-8 



67.2 
57-6 

HO. 

48,6 

13-6 

65- 

29,7 

151,8 



43-2 

253-0 

80. 

15.6 

247-5 



1,174-6 



168 

II. 2 
190,8 
229,6 

32 



10,4 



2.2 



208 



142.8 



II. 2 



123-S 

I24,& 

1,494,9 



2,749,4 
1,174,8 



1.574-6 



| i>642.43 ^., 127^078 



^{xy)= 1,574-6 
Coefficient of correlation : 



Na-^a-^ 28x7.65x31.1 



= +0.236 



^ . 0.2365 
Regression = — ^ = 0.95 



400 Carhon-dioxide Excretion in Man [April 

Or, out of a total of seventy-seven experiments, there was direct 
relationship between barometric change and carbon dioxide excre- 
tion in forty-two experiments, or 54.5 per cent. 

It will be Seen from these results that the apparent degree of 
correspondence, so far as it is revealed by this method of analysis, 
is greater in the morning experiments than in those carried out at 
midday or in the evening. This is probably due to the fact that in 
the morning not merely the digestive organs, but the whole System, 
is in a more uniform condition than at any other time during the 
twenty-four hours. 

Application of the method of least Squares. It now seemed 
desirable to subject the results obtained in this series of experiments 
to a more rigorous analysis than that just described, with a view of 
discovering what is the degree of correlation between the two vari- 
ables, the barometric height and the rate of excretion of carbon 
dioxide, during muscular rest. The data obtained in the experi- 
ments were, therefore, examined by the method of least Squares, 
which was applied separately to the three sets of data from each 
subject in order that the effect of different times of day might be 
determined separately. 

In Table 3 are given the barometric height and the correspond- 
ing carbon dioxide excretion f the problem is to find the correlation 
between these two quantities, and also the regression of carbon 
dioxide on barometric height, i. e., the amount of change in excre- 
tion of carbon dioxide for a millimeter change in barometric height. 
The means of columns i and 2 are obtained in the usual manner, by 
dividing the total in each column by the number of experiments 
{N). Having obtained these means, two additional columns are 
formed, giving the deviation of each Observation from the mean of 
its column. In columns 5 and 6 are entered the Squares of the 
deviations {X^ and F^). The Standard deviation {a^) is now ob- 
tained by dividing the sum of the Squares in the fifth column by the 
number of experiments, N, and extracting the square root of the 
quotient; the Standard deviation for 3; is, of course, found in the 
same manner. 



p;ir2 I1642.4 12/ 127078 

"^ = A'^ = >i-2F- = 7.65 and ., = ^^=^-g^ = 3i.i 

* The data are those obtained from experiments on subject A at noon. 



I9I3] 



G. 0. Higley 



401 



The products XY are now collected, the negative in column 7 
and the positive in column 8, and the totals determined. We have, 
then, 2(XF) =2,749.4—1,174.8 = 1,574.6. 

From this the coefficient of correlation (f ) is obtained : 






^xy) 



1,574.6 



Na.a^- 



= + 0.236 



28 X7.65 X 31. 1 

The positive sign of this coefficient indicates, of course, that the 
relationship between barometric change and carbon dioxide excre- 
tion in this case is direct or that the two variables change in the 
same sense. Since a coefficient of correlation of i indicates perfect 
correlation, the result obtained in the series of experiments repre- 
sented in Table i indicates a slight degree of correlation. The 
probable error of a correlation coefficient of this value for a series 
of 25 observations is at least 0.13 so that the value of r is 
0.16 ±0.13. 

The results of the whole series of experiments are summed up in 
Table 4. 

TABLE 4 

General Summary 



Sub- 
ject 


Hour 


xi 


y2 


N 


Ol 


<ra 


S(.xy) 


Coefficient 
of corre- 
lation, r 


Probable 
error 


Regres- 
sion 


A 


7 A.M. 


1,769.7 


24.951 


29 


7.8 


293 


828. 


+0.12 


±0.13 


0.45 


A 


12 M. 


1,642.4 


27.078 


28 


7.6s 


3I.I 


1.574-6 


+0.236 


±0.126 


0.95 


A 


5 P.M. 


1.315.71 


17.620 


24 


7-4 


27.1 


589-9 


— 0.12 


=tO.I26 


0-44 


B 


7 A.M. 


1,324.02 


59,721 


24 


7-4 


49.8 


1.787-7 


-0.2 


=fcO.I29 


0.13 


B 


12 M. 


1,31302 


40,919 


26 


6.9 


44.2 


818.3 


— 0.04 


±0.124 


0.09 


B 


5 P.M. 


1,158.7 


102,631 


25 


6.5 


64.0 


1,926.1 


-0.23 


±0.13 


0.17 


C 


7 A.M. 


1,228.2 


8,693 


27 


8.0 


17.9 


1,228.2 


+0.316 


±0.12 


0.7 


C 


12 M. 


1,652.0 


26,484 


27 


7.8 


31.2 


2,627.6 


+0.39 


±0.11 


1-5 


C 


5 P.M. 


1,232.6 


23,076 


25 


7.02 


30.38 


1,325-1 


+0.248 


±0.125 


1.07 



Conclusions. There were indications in this work of an in- 
fluence of barometric change on carbon dioxide excretion in the 
case of one subject, C, since there were three positive coefficients of 
correlation having the value of 0.316, 0.39, and 0.248, for morning, 
noon, and evening experiments (perfect correlation would be in- 
dicated by a coefficient of i ) ; a slight direct influence is also indi- 
cated in the case of A, whose coefficients were 0.12, 0.236 and 
— 0.12. In the case of B, whose values of carbon dioxide excre- 



402 Carhon-dioxide Excretion in Man [April 

tion tliroughout tlie work were qiiite irregulär, there were three 
negative coefficients with values — 0.2, — 0.04, and — 0.18. 

These results are, perhaps, what might have been expected. 
The barometric change is evidently a minor influence and its effect 
is there fore liable to be masked by other influences, such as exercise, 
amount and character of meals, etc. Moreover, the effect upon the 
muscular endurance noted by Prof. Lombard in bis own case has 
not been verified in the case of other subjects. The writer is of the 
opinion that if a series of parallel ergographic and respiration ex- 
periments were made on a number of subjects, it would be found 
that positive effects of barometric changes on muscular endurance 
are accompanied in general by positive coefficients of correlation of 
barometric change with rate of excretion of carbon dioxide. 

This work was done in the Physiological Laboratory of the 
University of Michigan, with the apparatus described by Higley 
and Bowen {American Journal of Physiology, i904-'o5, xii, p. 31 1 ). 



THE RELATION OF ACAPNIA TO SHOCK, AND A 
CONSIDERATION OF THE MECHANICAL 
EFFECTS OF ARTIFICIAL HYPER- 
RESPIRATION UPON THE 
CIRCULATION 

HENRY H. JANEWAY and EPHRAIM M. EWING 

(Laboratories of Experimental Surgery and Physiology of the New York 
University and Bellevue Hospital Medical School^) 

It has been claimed that the most important factor in the causa- 
tion of shock is diminution of the carbon-dioxide content in the 
blood and that this diminution is a regulär consequence of all influ- 
ences resulting in shock. That carbon dioxide exercises significant 
physiological f unctions cannot be denied ; determination, theref ore, 
of the true significance of the diminution of its normal proportion 
in the blood is important and bears a special relation to the various 
methods of artificial respiration utilized in thoracic surgery. 

This study was undertaken for the purpose of investigating the 
relation of acapnia to shock. All experiments were performed on 
dogs. 

The first series of experiments was conducted for the purpose 
of studying the effect of Variation- in intrapulmonic air-pressure 
upon the blood-pressure. The thorax was opened laterally, a T-tube 
connected with a water-manometer was tied in a small bronchus, and 
the heart enclosed in a Henderson cardimeter connected with a 
recording tambour. The blood-pressure was recorded from the 
carotid artery, The thorax was then closed and intratracheal insuf- 
flation was given from an apparatus provided with an exhaust valve, 
which reduced the pressure to approximately zero from four to 
twelve times per minute. When the machine was running at a pres- 
sure of 6 mm. of Hg there was an average rise of blood pressure 
of 15 mm., each time the exhaust valve operated. 

* The work presented in this paper was begun in the Laboratory of Biological 
Chemistry of Columbia University, at the College of Physicians and Surgeons; 
BiocHEMicAL Bulletin, 1912, ii, p. 175. 

403 



404 Relation of Acapnia to Shock [April 

In one experiment with an increase in intrabronchial pressure 
of from 8 to 30 mm. Hg, the blood pressure feil from 122 to 55 
mm. Hg, and the Volumetrie tracing indicated that the Output from 
the heart had diminished about 44 per cent. These variations in 
blood-pressure were completed within a few seconds after the 
change in intrabronchial pressure, and could be duplicated at will. 
A rise of intrabronchial pressure above 8 to 10 mm. Hg always 
caused a fall in blood-pressure and it was concluded that the Varia- 
tion in pressure was the result of a diminution of the venous return 
to the heart, resulting from compression of the veins in the thorax. 
In view of the marked changes in the blood-pressure and Output of 
the heart resulting from small variations in intrapulmonic pressure, 
it is evident that, in any experiments planned for the purpose of 
estimating the part played in the production of shock by a diminu- 
tion of carbon-dioxide content, induced by artificial hyper-respira- 
tion, the effects of the increase of intrapulmonic pressure upon the 
return flow of blood to the heart must be considered. 

With the second series of animals, Henderson's experiments 
were duplicated, the dogs being artificially respired by means of 
a force-and-suction pump, working about seventy times per minute. 
The animals were given morphin, and ether was administered only 
when necessary. In these experiments, blood-pressure feil about 
40 per cent. within one minute after artificial respiration was begun, 
and then decreased more slowly throughout the experiment to 
between 40 and 50 mm. Hg. At the end of the experiment, when 
the artificial respiration was stopped, the blood pressure increased 
60 to 90 per cent. within a few seconds. In all experiments the 
blood analysis showed that the carbon-dioxide content, at the end, 
was only 40 to 50 per cent. of the original amount. These animals, 
at the end of two to three hours of artificial respiration, were all 
in a condition of deep shock. This degree of shock was indicated 
by a rapid pulse, a low blood-pressure, and a marked degree of 
insensibility to sensory Stimulation. Three of the animals so 
treated lived three days (dying of secondary effects of the experi- 
ment), and one lived twenty-four hours. None of them died from 
the immediate effects of the experiment. During these experiments, 
when the artificial respiration was interrupted or permanently 



1913] Henry H. Janeway and Ephraim M. Ezving 405 

stopped at the end of the experiment, the period of apnea lasted 
only one or two minutes, so that no death resulted directly from 
asphyxia dependent in turn upon acapnia. The absence of a pro- 
longed period of apnea is explained by the fact that the effect of ether 
was not added to that of morphin. 

With a tJiird series of animals the experiments just described 
were duplicated, with the exception that the carbon-dioxide content 
of the blood was maintained at its normal level, or slightly above it. 
The conservation of the carbon dioxide was accomplished by insert- 
ing a large rubber bag, to act as a reservoir, between the suction 
pump and the force pump, thus creating an almost perfectly closed 
circuit ; the dog thus rebreathed expired air. To replace the small 
amount of air and carbon dioxide lost from the animal's trachea, 
carbon dioxide was administered from a tank into the rubber bag, 
where it mixed with air drawn in from the trachea. In these ex- 
periments the animals went into the same degree of shock in two 
or three hours as those of the second series, in which the carbon- 
dioxide content of the blood was diminished to 40 per cent. of the 
original volume. One animal died on the table just before the com- 
pletion of the experiment, the others lived for from one to three 
days. Blood-pressure changes in the two series were similar but a 
characteristic of the experiments, in which the carbon-dioxide con- 
tent was kept at or a little above the normal, was a less rapid and 
weaker heart-beat than that observed when the carbon-dioxide con- 
tent was diminished. 

No other conclusions can be drawn from the experiments of 
Series i and 2 than that the reduction in the carbon-dioxide content 
of the blood was not an important factor in the causation of shock 
produced by hyper-respiration, and that in shock so produced, the 
essential factor was an interference with the venous return to the 
heart. 

In the fourth series of experiments the effects of aerating and 
handling the intestines were studied. A celluloid window was 
placed in the abdominal wall, and a stream of warm moistened air 
was passed over the intestines for a period of three hours. During 
this procedure the animals breathed normally, the blood-pressure 
was 163 mm. Hg, the content of carbon dioxide was slightly 



4o6 Relation of Acapnia to Shock [April 

diminished, and there was no evidence o£ shock. Beneath the cellu- 
loid the absence of peristalsis could be observed as well as the effi- 
ciency of the aeration and failure of the intestines to become dry. 
The celkiloid was then removed, the intestines spread out, and the 
aeration continued. After 45 minutes the carbon-dioxide determi- 
nation indicated a content of 38.8 vol. per cent., and blood-pressure 
was 1 53 mm. Hg. The intestines were then handled ; in ten minutes 
blood-pressure had fallen to 98 mm., in twenty minutes to 56 mm. 
Hg, and in forty minutes there was still 31.6 vol. per cent. of carbon 
dioxide in the arterial blood. 

In another experiment the intestines were exposed and aerated 
(not handled). The carbon-dioxide content of the blood was main- 
tained by connecting a long tube with the trachea. After one hour 
and a half, blood-pressure had changed but i mm. Hg, and the 
animal was in good condition. The intestines were then handled 
and in ten minutes the blood-pressure feil from 122 to 60 mm. Hg. 
The carbon-dioxide content w^as 45.1 vol. per cent. In twenty-five 
minutes the blood-pressure was 46 mm. Hg, the carbon-dioxide 
content normal, and the dog in a severe degree of shock. 

In these abdominal experiments the primary factor concerned is 
unquestionably the manipulation of the intestines and not any 
diminution of carbon-dioxide content caused thereby. It will be 
remembered that in the similar experiments with aeration of the 
intestines reported by Henderson, the intestines were handled gently. 
We have been unable to find any mention in his paper of aeration of 
the abdominal cavity with air alone beneath a celluloid membrane 
as a control. 

Henderson's control experiment, in which he did not secure 
shock, included aeration (with astreamof air plus carbon dioxide) 
of the abdominal cavity beneath a celluloid window in the abdominal 
wall. Our own experiments show that aeration of the intestines 
without the addition of carbon dioxide does not produce shock. 



CLEAVAGE OF PYROMUCURIC ACID BY MOLD 

ENZYMES 

ARTHUR W. DOX and RAY E. NEIDIG 
(Chemical Section of the Iowa State College, Arnes, Iowa) 

In a recent paper^ we have presented data showing that the 
formol-titrimetric method of Sörensen can be applied to the de- 
termination of the cleavage of hippuric acid by enzymes. The six 
saprophytic fungi studied were found to produce an enzyme capable 
of hydrolyzing as miich as 90 to 100 per cent. of a Solution of 
sodium hippurate in the presence of toluene as an antiseptic. The 
production of enzyme was independent of the presence of the cor- 
responding Substrate in the nutrient medium upon which the fungus 
was cultivated, and the age of the culture, within the limits studied, 
had little influenae upon this enzymic activity. 

If we assume that the synthesis in the animal organism of hip- 
puric acid from benzoic acid is an enzymic process, the synthesis of 
the corresponding derivatives from substituted benzoic acids might 
well be attributed to the same cause. It is known, for example, 
that o-brombenzoic, salicylic, toluic and other substituted benzoic 
acids, when administered orally, are conjugated with glycocoll and 
excreted through the kidneys as o-brombenzoylglycocoU, salicyluric, 
toluric, etc., acids. Likewise, a homologue of benzoic acid, e. g., 
phenylacetic, may be conjugated in the same way and eliminated 
as phenylaceturic acid. It is quite unlikely that these substituted 
benzoic acids all require separate enzymes for their conjugation 
with glycocoll, and it is equally improbable that the hydrolysis of 
the substituted hippuric acids would require specific enzymes. 

This reasoning may be extended, also, to analogous Compounds 
where the benzene nucleus is replaced by a heterocycle. For 
example, ct-pyridine carboxylic acid is united in exactly the same 
manner with glycocoll and excreted as a-pyridinuric acid. The 

* Dox and Neidig : Zeitschr. f. physiol. Chem., 1913, Ixxxv, p. 68. 

407 



4o8 



Cleavage of Pyromucuric Acid 



[April 



striking analogy between benzene and the two heterocycles, fur- 
furane and thiophene, led Jaffe^ to a study of the behavior of the 
corresponding derivatives of these substances in the animal organ- 
ism. As was anticipated, fiirfurol behaved exactly as did benzal- 
dehyde, undergoing oxidation to the acid and then conjugation with 
glycocoll, and was ehminated principally as pyromucuric acid. 
Similarly, thiophenic acid was excreted as thiophenuric acid. 

None of these heterocychc analogues of hippuric acid have, to 
OUT knowledge, been studied with reference to their cleavage. 
Knowing from previous work that the lower fungi produce an 
enzyme capable of hydrolyzing hippuric acid, we thought it would 
be of interest to test their activity toward one of these heterocyclic 
Compounds. 

With this object in view the following experiments were under- 
taken. Cultures of seven molds were grown for two weeks on the 
nutrient medium previously described.^ The extraction of enzyme 
was efYected by the following method : The mycelium was washed 
with distilled water, ground in a mortar with fragments of glass 
and the juice obtained at a pressure of 350 kg. per sq. cm. In each 
case about 20 c.c. of extract were obtained, 10 c.c. of which were 
used in the enzyme experiment and 10 c.c. in the control. In the 
enzyme experiment, 25 c.c. of a i per cent. Solution of pyromucuric 
acid,* previously neutralized with sodium hydroxide, were added 



TABLE I 

Data pertaining to the cleavage of pyromucuric acid 



Source of enzyme 


Titration 

«/lo Ba(OH)2 

c.c. 


Control 

«/ioBa(OH)j 

c.c. 


Difference 


Cleavage 


Aspergillus fumigalus 


10.9 
19.1 
20.4 
7.0 
13-1 

II. 

13.8 
4-9 
0.6 


5-6 
12.0 

13-3 

2.8 

6.3 
5-0 
10.5 
0.2 
0.4 


5-3 
7-1 
7-1 
4.2 
6.8 
6.0 

4.7 
0.2 


35.8 
48.0 
48.0 
28.4 
45-9 
40.S 
22.3 


Aspergillus niger 


Aspergillus clavatus 


Penicillium roqueforti 

Penicillium camemberti 

Penicillium expansum 


Fusarium oxysporium 


Emulsin (Kahlbaum ) 


31.8 


Taka-diastase (Parke-Davis) . . 


1.4 



^Jaffe and Cohn : Berichte, 1887, xx, p. 2311; Jaffe and Levy: Berichte, 
1888, xxi, p. 3458. 

'Dox and Neidig: Loc. cit. 

* The pyromucuric acid was prepared by the method of Jaffe and Cohn. It 



I9I3] 



Arthur W. Dox and Ray E. Neidig 



409 



and to the control, 25 c.c. of water. Toluene was used throughout 
as an antiseptic. After two weeks, during which time the flasks 
were shaken at frequent intervals, the formol-titration was made, 
with the results given in Table i. 

Comparing the foregoing results with those obtained with hip- 
puric acid,^ it will be noted that the extent of the cleavage of the acid 
into its components is in this case considerably less. This can hardly 
be taken as evidence in support of any assumption that we are 
dealing with two separate enzymes. Quantitative differences ob- 
served in work of this nature have little significance, unless the 
same enzyme preparation has been employed and a uniformity of 
all other conditions maintained. 

TABLE 2 

Data pertaining to the formation of ammonia. 



Organism 


Titration 

n/io H2SO4 

c.c. 


Control 
K.'IO H.iS04 

c.c. 


Ammonia 

«/lO H2SO4 

c.c. 


A.S'bereillus fumisatus 


0.90 
I.OO 
I.OO 

0.35 
0.90 
1.6s 
2.00 


0.53 
0.68 
0.78 

0.33 
0.63 
0.88 
1.68 


0.17 


Aspergillus niger 


0.32 


Aspergillus clavatus 


0.22 


Penicillium roqueforti 


0.02 


Penicillium camemberti 


0.27 


Penicillium expansum 


0.77 


Fusarium oxysporium 


0.32 



As in our previous work with hippuric acid, the contents of the 
flasks after this titration were distilled with magnesium oxide for 
the determination of ammonia. 



Ammonia can hardly be regarded as a direct product of the 
cleavage of pyromucuric acid. The small amount found probably 
results from further decomposition of the glycocoU by another 
enzyme. This cleavage of glycocoll is, however, so slight as to be 
practically negligible. 

Thiophenuric acid was not available at the time these experi- 
ments were carried out. We propose to test the activity of mold 
enzymes toward this substance at a future date. 

melted at 163° C. ; 0.25 gm. required 14.8 c.c. of n/10 Ba(0H)2 Solution for 
neutralization ; theory, 14.8 c.c. 
'Dox and Neidig: Loc. cit. 



ANALYSIS OF THE ASH OF THE CASTOR BEAN 

MARSTON LOVELL HAMLIN 
(Harriman Research Lahoratory, Roosevelt Hospital, New York) 

In the discussion, following the presentation at the February 
meeting of the Columbia University Biochemical Association, of the 
results of investigations conducted in this laboratory on the effect of 
manganous sulfate on the action of lipase in the castor bean, 
Ricinus communis,'^ Professor Gies suggested that the effect pro- 
duced in vitro by comparatively large amounts of manganese was 
very possibly induced in the plant by much smaller amounts, and 
that it would be of particular interest to test the ash of the seed for 
its presence.^ The ash of the seed was therefore tested for man- 
ganese ; and, at the same time, silica, magnesia, lime and phosphoric 
acid were determined. 

A sample of the kerneis of the seed, cold-pressed, ground, ex- 
haustively extracted with ether, as for use in lipolytic experiments,^ 
and dried in vacuo over sulfuric acid, was slowly ignited in a 
platinum dish. The black residue was treated several times with 
nitric acid and re-ignited tili free from carbon. The residue was 
weighed, taken up with water and nitric acid, and the Solution 
filtered. Calcium sulfate was precipitated in the filtrate by sulfuric 
acid and alcohol; and, in the filtrate from this precipitate, mag- 
nesium and phosphorus were determined in separate aliquot parts, 
each as magnesium pyrophosphate.* 

Of the powdered kerneis, 4.7698 grams gave 0.3483 gram of 
ash, or 7.3 per cent. Of this, 0.0018 gram was insoluble and 

*Falk and Hamlin : Jour. Amer. Chem. Soc., 1913, xxxv, p. 210. An abstract 
of this paper appears in this issue of the Biochemical Bulletin (p. 455). 

' The importance of infinitesimal amounts of manganese in plant growth has 
been repeatedly pointed out by G. Bertrand. For a recent presentation of his 
views, see his general lecture delivered before the Eighth International Congress 
of Applied Chemistry, New York, September, 1912. 

* Falk and Hamlin : Loc. cit. 

* Abderhalden : Handbuch der bioch. Arbeitsmeth., 1912, vi, p. 381. 

410 



I9I3] 



Marston Lovell H amiin 



411 



taken as silica; this amounted to 0.5 per cent. of the ash, The 
calcium sulfate weighed 0.0326 gram, which represented 0.0134 
gram of calcium oxide, er 3.8 per cent. of the ash. Of the filtrate 
diluted to 500 c.c, two portions of 200 c.c. each were taken for the 
determination of magnesium and phosphorus. In one, 0.1052 gram 
of magnesium pyrophosphate represented 0.0671 gram of phos- 
phorus pentoxide, or 48.2 per cent. of the total ash. In the other 
0.0798 gram of magnesium pyrophosphate represented 0.0289 grarn 
of magnesium oxide or 20.7 per cent. of the total ash. 

To test for manganese^ 5.000 grams of the oil-free powdered 
kerneis were ignited as above and the ash taken up with 4 c.c. of 
nitric acid Solution (sp. gr. 1.20) and water, and this liquid filtered. 
It was diluted to 20 c.c. and a 10 c.c. portion was boiled with 0.5 
gram of lead peroxide for several minutes, the precipitate allowed 
to settle, and the liquid decanted into a test tube. Next a Solution 
containing 20 milligrams of manganous sulfate per 5 c.c. was 
diluted with nitric acid Solution and water to fifty times its original 
volume, and treated in the same way. It was found that the Solu- 
tion of the ash was matched in color by a Solution 1/700 as concen- 
trated as the original manganous sulfate Solution; therefore 5 grams 
of the kernel powder contain 4 X 20 X 1/700 milligrams of man- 
ganous sulfate, or 0.000028 gram of manganese, which is 0.00056 
per cent. The results are summarized below : 





SiO, 


CaO 


MgO 


P2O, 


Mn 


Total ash 


Per cent. in dry, oil-free kernel 

Per cent. in ash 


0.04 
o.S 


0.28 
3-9 


i-Si 
20.7 


3-52 

48.2 


0.00056 
0.0076 


7-3 


Schulze and Godet, who analyzed the ash of this seed,^ obtained 
the following data for dry but not oil-free kerneis : 




CaO 


MgO 


PjOs 


Total ash 


Per cent. in drv substance 


o.i"; 


0.72 


1.16 
31-9 


3-64 


Per cent. in ash 


4.0 






19.8 





These results indicate that the cold-pressing and ether extraction 
in my own work removed substance amounting to about half the 
weight of the kernel. 

'Noyes, Bray and Spear: Tech. Quart., 1908, xxi, p. 116. 
'Schulze and Godet: Z. f. physiol. Chem., 1908, Iviii, pp. 156-61. 



NOTES ON THE CHEMICAL NATURE OF THE 

"TANNIN MASSES" IN THE FRUIT OF 

THE PERSIMMON 

ERNEST D. CLARK 

(Biochemical Laboratory of Columbia University, at the College of Physicians 

and Surgeons, New York) 

Introduction. The researches of Lloyd upon the nature of the 
ripening process in persimmons and dates led him to believe that the 
loss of astringency at maturity is due to the combinatlon of tannin 
with a colloidal substance of carbohydrate nature in the "tannin 
mass." This combination takes place ordinarily at the time of 
ripening, probably under the influence of enzymes ; but it may be 
hastened by artificial means, such as exposure to the vapors of acetic 
acid and ethyl nitrite, and to carbon dioxid under normal and supra- 
normal pressures. Lloyd^ defines his tannin mass as "the tannin 
idioblast containing tannin associated with a second coUoid." After 
the Union of tannin and this colloidal substance has taken place no 
more tannin can be extracted nor do alkaloids indicate its presence. 
It is evident, then, that a knowledge of the chemical substances in the 
tannin mass would facilitate further investigation of the mechanism 
of the ripening process. 

Preparation of tannin masses. The tannin masses used in 
cur experiments were prepared by Prof. F. E. Lloyd^ as follows : 
Fully ripe persimmons (Diospyros) were shaken and macerated 
with water until they formed a thick paste, from which the heavy 
tannin masses settled out. This process was repeated until the sepa- 
rated masses were thoroughly washed by decantation and also 
purified from all debris. The resulting thick Suspension of tannin 

''Lloyd: Biochemical Bulletin, 1911, i, p. 7. See also Plant World, 191 1, 
xiv, p. i. 

^ Lloyd: Zeitsch. f. Chem. und Ind. d. Kolloide, 1911, ix, pp. 65-73. The 
material was shipped to this laboratory from Prof. Lloyd's laboratory at 
Auburn, Ala. 

412 



1913] Ernest D. Clark 4^3 

masses was kept under a layer of ether. This material had the 
appearance of a multitude of minute cylindrical particles which were 
colorless and transparent. Upon exposure to the air these particles 
soon became brovvn, probably f rom oxidation. 

General properties of the tannin masses before hydrolysis. 
In both the Millon and xanthoproteic tests the tannin masses 
gradually assumed a dark brown color ; a change apparently similar 
to that resulting from slow oxidation by the air. With Fehling or 
Fehling-Benedict Solution the masses turned black at once, but no 
reduction was observed. Tests for pentoses with conc. hydro- 
chloric acid Solution and phloroglucinol caused the particles to be- 
come bright red, but this acid ahne caused the same change. lodin 
in potassium iodid Solution produced no coloration. Repeated 
fusions with metallic sodium failed to indicate the presence of 
nitrogen. This Observation was confirmed by subjecting 2 gm. of 
the material to the Kjeldähl process for nitrogen, with negative 
results. 

The presence of phloroglucinol in the masses was shown by 
adding to them a little vanillin in hydrochloric acid Solution when 
the particles were stained a beautiful magenta shade, a result similar 
to that obtained when pure phloroglucinol and vanillin react in the 
presence of traces of hydrochloric acid. Various other phenolic 
substances, however, form brightly colored condensation products 
with vanillin under such conditions.^ 

All tests for pentose by boiling with hydrochloric acid Solution 
and allowing the vapors to act on anilin acetate paper, and also by 
boiling the particles with conc. hydrochloric acid Solution plus orcin, 
were negative. No pentose is present, apparently, in spite of the 
red color given by the phloroglucinol-hydrochloric acid test, a result 
that may have been caused by the interaction of these substances 
with a phenolic substance (like vanillin) rather than by pentoses. 
With the Molisch reagent a very strong positive result was obtained 
and, furthermore, we found that exactly the same purplish ring was 
formed by the tannin masses with pure sulfuric acid alone. 

'Hartwick and Winckel (Archiv d. Pharm., 1904, ccxlii, p. 471), showed the 
presence of phloroglucinol in the tannin masses of Ceratonia siliqua and Phoenix 
dactylifera. Tichomirow (Bull. Soc. Imp. d. Nat. Sei., Moskau, 1905) obtained 
the same reactions in the tannin masses of the persimmon, indicating the presence 
of a phenol. 



414 "Tannin Masses" in Fruit of the Persimmon [April 

Finally, the tannin masses stained deep blue with ferric chlorid 
Solution and, as Lloyd found, this color was quickly destroyed by 
nitric acid. These properties of the tannin masses show that the 
latter contain neither reducing sugars nor protein; they also suggest 
that phloroglucinol occurs with tannin and cellulose material. 

Hydrolysis of tannin masses with 0.2 per cent. and 2 per 
Cent, hydrochloric acid Solutions. The addition of 0.2 per cent. 
hydrochloric acid Solution to tannin masses, with subsequent heating, 
caused them to disintegrate, giving the whole liquid a bright cherry 
red color. The tannin masses as such disappeared and a white 
flocculent residue remained suspended in the red liquid. The mix- 
ture was filtered and to the filtrate we added 0.5 per cent. sodium 
hydroxid Solution until the acid was neutralized. Beyond this 
neutral point the red color disappeared, but on standing or aftcr 
further treatment with alkali, a gradually increasing brownish 
coloration took its place. 

The neutral filtrate was evaporated to dryness on the steam 
bath and an aqueous Solution of this reddish residue was subjected 
to the f ollowing tests, with the results indicated : With ferric chlorid 
Solution, a purplish black coloration was given ; with Fehling-Bene- 
dict Solution, a dark brownish precipitate was formed at once but 
it soon changed to a characteristic reduction when heated; the 
Molisch test was a typical positive one; with vanillin-hydrochloric 
acid Solution a red color appeared; and a peculiar non-typical pre- 
cipitate was produced when we attempted to form an osazone with 
Phenylhydrazine hydrochlorid-sodium acetate mixture. 

We filtered off the white amorphous residue and washed it free 
from chlorid. Upon testing it we found there was little if any 
reduction of Fehling-Benedict Solution; with the vanillin-hydro- 
chloric acid reagent there was no red coloration except in a few 
deeply stained particles (stone-cells?) f with ferric chlorid Solution 
a brownish color appeared but no bluish shade. Finally, the residue, 

* " Undoubtedly they were stone cells, as I observed the same thing. To get 
this reaction all one needs to do is to add hydrochloric acid to the mucilaginous 
pulp which includes stone cells, and these become colored. When I observed this 
I did not refer the reaction to the presence in the tannin masses of the phloro- 
glucinol. This I later satisfied myself to be the case." (Lloyd : Personal com- 
munication.) 



1913] Ernest D. Clark 415 

when suspended in the " biuret reagent," was slowly colored blue as 
the cellulose material absorbed copper f rom the alkaline Solution.^ 

Next, the tannin masses were hydrolyzed with a 2 per cent. 
hydrochloric acid Solution. The hydrolytic products, when treated 
in exactly the same manner, gave results identical with those just 
described. The more concentrated acid seemed to favor the forma- 
tion of dark-colored products. 

From the foregoing results it appears probable that tannin 
masses contain tannin and phloroglucinol combined with a third sub- 
stance, from which union they are released when hydrolysis takes 
place. The acid gelatinizes that part (colloidal) of the masses 
which appears to be cellulose or some related substance. 

Hydrolysis of tannin masses with 0.5 per cent. and 5 per 
Cent, sodium hydroxid Solutions. Treated with 0.5 per cent. or 
5 per cent. Solution of sodium hydroxid, "tannin masses" gave 
purplish brown mixtures containing a bulky gelatinous residue, 
which we removed by filtration. The filtrate was carefully neutral- 
ized with dilute hydrochloric acid Solution and the reddish brown 
precipitate that formed was filtered off, washed, and dissolved in 
water, This liquid was tested for reducing power with Fehllng- 
Benedict and ammoniacal silver Solutions, both reagents showing 
weakly positive results. The Molisch reagent, and also ferric 
chlorid Solution, gave dark non-typical colorations. With vanillin- 
sulfuric acid Solution a typical red color was produced, when the 
liquid was evaporated. 

The Solution of the material not precipitated from alkaline Solu- 
tion by acid was now tested in the usual way for reducing power, 
presence of phloroglucinol, etc., with uncertain results, due to the 
dark color of the Solution. When alkaline Solutions of the hydro- 
lytic products are exposed to the air, oxidation seems to occur and 
dark complex substances are formed. The gelatinous material 
which resisted hydrolysis was filtered and washed. It appeared to 
consist of the collapsed cell-walls of tannin masses. When dried it 
formed a light-colored scaly mass composed of small particles. On 
the whole, the results of alkaline treatment of the tannin masses 

^Kantor and Gies: Biochemical Bulletin, 191 i, i, p. 269. 



41 6 "Tannin Masses" in Fruit of the Persimmon [April 

are similar to those obtained with acids, except that alkali accelerates 
oxidation by atmospheric oxygen. 

Properties of mixed Solutions of pure tannin and phloro- 
glucinol. Mixtures in varying proportions of Solutions of pure 
tannic acid and phloroglucinol were tested and found to show many 
similarities to the hydrolytic products of the tannin masses. The 
Molisch test was strongly positive. With ferric chlorid such mixed 
Solutions gave a purplish black color which was accompanied by the 
gradual formation of a precipitate of the same shade. The typical 
influence on Fehling-Benedict Solution was observed ; namely, heavy 
greenish precipitation at first but, upon heating, this changed to a 
characteristic red reduction. When some of the mixed Solution was 
evaporated with the vanillin-hydrochloric acid reagent, a definitely 
positive result was obtained but the tannin seemed to cause a 
brownish tint not given by phloroglucinol alone. 

Pure phloroglucinol Solutions were negative to the Molisch test 
and also to the Fehling-Benedict test. Ferric chlorid Solution pro- 
duced a clear blue color with pure phloroglucinol but did not give 
a precipitate, thus differing f rom tannin, which forms a blackish pre- 
cipitate at once under these conditions. Solutions of pure tannin 
yield a typical Molisch test and reduce Fehling-Benedict Solution in 
the characteristic manner just described. It is evident that, in the 
presence of phloroglucinol, one cannot conclude that a purple color 
with ferric chlorid indicates tannin. The addition of nitric acid to 
the mixture already turned purple by ferric chlorid caused an in- 
stantaneous change to a brownish tint. 

It is obvious, then, that, in the tests indicated, mixtures of tannin 
and phloroglucinol differ in no essential way from the hydrolytic 
products of the tannin masses. 

Sources of the tannin and phloroglucinol in the tannin 
masses. The material in the tannin masses from which tannin and 
phloroglucinol are derived by hydrolysis is probably one of the so- 
called phloroglucin-tannoids, which were found by Graebe® to yield 
these hydrolytic products by treatment with acids, etc. WeinzierF 
showed that phloroglucinol is widely distributed among plants but 

* Graebe : Ber. d. d. ehem. Ges., 1903, xxxvi, p. 212. 

'Weinzierl: Oesterr. bot. Zisch., 1874, xxvi, p. 285. See also Nickel: Bot. 
Centralblatt., 1891, xlv, p. 394. 



1913] Ernest D. Clark 417 

he was unable to say whether it occurs as a waste product or one 
useful in the synthetic processes. 

Waage^ made some interesting observations in regard to phloro- 
glucinol in plants. He confirmed the Statement that the substance is 
widely distributed in plants and suggested that it might arise during 
photosynthetic processes, just as glucose probably does. He feit 
that, because of the dark blue color given by phloroglucinol and 
ferric chlorid, and because of the bleaching of methylene blue by 
the former substance, one cannot rely on the previous deductions in 
regard to the presence of tannin in cell vacuoles based on these 
reactions for tannin. In fact, he criticized, on the same ground, the 
work of af Klercker^ on tannin in vacuoles. Schiff^*^ found that 
under suitable conditions, phloroglucinol and carbon dioxid combine 
to form phloro-tannic acid which, upon heating, yields a red sub- 
stance like phlobaphene. Evidently, tannin is present along with 
phloroglucinol in many plants, but the physiological röle of these 
substances is as little known as is the nature of the combination 
between them in cases like that of the tannin masses of the per- 
simmon, Much more work on persimmons will be necessary to 
explain the part played by phloroglucinol in the loss of astringency 
in the mature fruit. 

Summary. Tannin masses from the fruit of the persimmon, 
by hydrolysis with weak acid or alkali, yield tannin, phloroglucinol, 
and considerable insoluble colloidal residue. Hydrolysis of such 
tannin masses does not produce hexose or pentose. 

The nature of the union between the tannin and phloroglucinol 
is unknown but it is probably similar to that of the phloroglucin- 
tannoids in various plants. 

The colloidal residue that resists hydrolysis seems to be a cellu- 
lose-like substance, which readily forms gelatinous masses with 
water or alkaline Solutions. Quantitative studies on large amounts 
of this third substance are desirable. 

In the presence of phloroglucinol, the ferric chlorid test for 
tannin is unreliable. 

* Waage: Ber. d. d. bot. Ges., 1890, viii, p. 250. 

'af Klercker: Bihang tili K. Svenska Vet.-Akad., 1888, xiii, p. 18 (repaged?). 

*• Schiff : Ann. d. Chem., 1888, ccxlv, p. 36; 1889, cclii, p. 87. 



41 8 "Tannin Masses" in Fruit of the Persimmon [April 

A study of the conditions necessary for the formation, and also 
the hydrolysis, of the phloroglucinol-tannin combination might help 
to explain the nature of the ripening- process in persimmons. 

This work resulted from a Suggestion by Prof. F. E. Lloyd to 
Prof. Wm, J. Gies, that material would be supplied in abundance 
for a study of the "tannin mass." I am indebted to Professor 
Lloyd not only for the material but also for many suggestions. To 
Professor Gies I am likewise indebted for helpful advice. 



HISTON AND ITS PREPARATION 

WALTER H. EDDY 

(Laboratory of Biological Chemistry of Columbia University, at the College of 
Physicians and Surgeons, New York) 

Contents. — Introduction, 419; historical review of histon preparations, 420; 
properties of histons, 425; thymus histon, 428; experimental, 430; summary of 
conclusions, 438; selected bibliography, 439. 

I. INTRODUCTION 

Several years ago I began a study of some artificial protein Com- 
pounds, as an introduction to an inquiry into the nature of protein 
conditions in cells {y)} In preparing histon from calf thymus 
glands by the Huiskamp (10), LiUenfeld (19) and Bang (2) 
processes, I found that the ammonia-precipitated product from 
neutral histon-hydrochlorid Solution was insoluble in water and 
hence not available for the intended study of histon Compounds. 
This result led me to reject the ammonia-precipitated histon and 
to use, instead, a Solution of histon hydrochlorid (made neutral by 
dialysis), Lately, in furtherance of this study, I have endeavored 
to prepare water-soluble basic histon, free from admixture with 
histon Salt of any kind. Various methods of preparation have been 
proposed, but I find that thymus histon is almost invariably pre- 
cipitated by the addition of an excess of ammonia to an acid Solu- 
tion of the substance, in spite of the fact that solubility in water is 
cited again and again as a property of histon. The only exceptions 
to this Statement were found in Kos&el's original paper (12) on 
goose-blood histon and in the description of Lota histon by Ehr- 
ström (8). I have spent much time in determining by various 
methods of preparation that the product precipitated by ammonia 
(thymus histon) is invariably insoluble in water. Recently, in re- 
viewing Fleroff's paper (9) on para-histon, I found a definite State- 
ment that ammonia-precipitated thymus histon is insoluble in both 
hot and cold water, and may be washed with water for purification. 

^ Figures in the text enclosed in parenthesis refer to the numbered iteras 
in the bibliography at the end of this paper. 

419 



420 Histon and its Preparation [April 

II. HISTORICAL REVIEW OF HISTON PREPARATIONS 

Before proceeding to the description of my experiments the 
reader may find it interesting to examine a brief summary of the 
main points in this confusing Situation as brought out in the litera- 
ture of the subject. 

I. Histons already prepared. The types of histons already 
prepared, with indications of sources and names of the original 
investigators, are shown in the appended summary : 



Name 


Source 


Prepared by 


Goose-blood 


Corpuscles 


Kossei (12) 


Thymus 


Calf thymus 


Lilienfeld (19) 


Salmin albumose 


Unripe sperm of the 


Miescher and 




salmon 


Schmiedeberg (22) 


Arbacin 


Testes of sea-urchin 


Matthews (21) 


Globin 


Hemoglobin 


Schulz (23) 


Scombron 


Mackerei sperm 


Bang (2) 


Para-histon 


Thymus 


Fleroff (9) 


Gadus 


Codfish sperm 


Kossei and 
Kutscher (16) 


Lota 


Burbot sperm 


Ehrström (8) 


Hen blood 


Blood corpuscles 


Ackerman (i) 


Centrophorus 


Centrophorus sperm 


Kossei (15) 



2. Methods of preparation. The methods of preparation are 
indicated in the f ollowing summaries : 

A. GoosE-BLOOD histon: Kossel, 1884 (12), Preparation I. 
Kossei centrifuged blood corpuscles free from plasma and then 
dissolved them in water and ether. The residue was extracted with 
water and ether until free from color and the final colorless mass 
extracted with hydrochloric acid Solution, The acid liquid was 
then saturated with sodium chlorid, the precipitated histon was 
filtered off, and purified by washing with acidified sodium chlorid 
Solution and then dialyzing against distilled water until free from 
chlorid. The resulting Solution was then treated with an excess of 
alcohol-ether and the precipitate dried to constant weight at 105° C. 
Preparation IL In the second preparation the hydrochloric acid 
Solution was precipitated with ammonia instead of sodium chlorid. 

B. Thymus HISTON : Lilienfeld, 1893 (19). Thymus glands 
were freed from fat with a knife, minced and the hash pressed in a 



1913] Walter H. Eddy 421 

hempen bag to remove the juice, which contained lymphocytes that 
were separated in a centrifuge. The lymphocytes were then ex- 
tracted with water to extract nucleohiston. (In a modification of 
this process, the minced glands were extracted directly with water.) 
The nucleohiston was precipitated from the extract with acetic acid, 
and purified by re-solution in water to which a little sodium car- 
bonate had been added and reprecipitating with acetic acid. The 
precipitate was then treated with 0.8 per cent. hydrochloric acid 
Solution. From this Solution of histon-hydrochloride, histon was 
precipitated with ammonia (added either before or after dialysing 
free from free acid). The ammonia-precipitated product was 
finally purified by washing with alcohol and ether, and dried to 
constant weight. 

C. Salmin histon: Miescher, 1896 {22). The nuclei of 
unripe salmon sperm were extracted with 0.25 per cent. hydro- 
chloric acid Solution, the extract filtered and (after dialysing to 
neutrality) the filtrate precipitated by Saturation with ammonium 
Sulfate or sodium chlorid. 

D. Arbacin histon: Mathews, 1897 (21). Preparation I. 
Dried sperm heads were extracted with 1-2 per cent. sulfuric acid 
Solution and the acid extract poured into a large volume of alcohol 
to precipitate the histon-sulfuric acid complex. The precipitaie was 
purified by washing with alcohol-ether and dried to constant weight. 
Preparation II. The alcohol-precipitated product was dissolved in 
water, the Solution made ammoniacal and filtered (no precipitate 
at this point), the filtrate poured into alcohol, and the resulting 
precipitate washed free from ammonia and redissolved in a small 
volume of water. Ammonia added to this concentrated Solution 
failed to completely precipitate the histon, which showed a strong 
tendency to remain dissolved in ammoniacal Solutions. Matthews' 
studies were based on the use of alcohol-precipitate from acid ex- 
tract, alcohol-precipitate from an ammoniacal water-solution, and 
ammoniacal water-solution neutralized with sulfuric acid. 

E. Globin histon: Schulz, 1898 (23). A Solution of hemo- 
globin was treated with dilute hydrochloric acid Solution and a 
brown precipitate obtained, soluble in the presence of a very slight 
excess of acid. When this precipitate was dissolved in acid and 



422 Htston and its Preparation [April 

one-fifth its volume of alcohol and ether added, the color passed 
into the ether leaving a clear water-alcohol Solution, from which 
ammonia precipitated a yellowish flocculent mass of globin, that, 
when washed free from ammonia, began to dissolve, a few drops 
of acetic acid Solution completing the process. Dialysed free from 
acid, a clear, neutral, slightly colored, odorless and tasteless Solution 
of globin resulted. Schulz' tests were based on this Solution and on 
the ammonia-precipitated product. The analyses were made on the 
latter, washed with alcohol and ether, and dried in vaciio to constant 
weight at a temperature of ioo° C. 

F. ScoMBRON histon: Bang, 1899 (2). Unripe mackerei 
sperm was heated with alcohol and the residue dried. This dried 
residue was then extracted with dilute hydrochloric acid Solution 
and the histon precipitated from the filtrate with caustic soda, 
ammonia, or by Saturation with sodium chlorid. The product was 
purified by re-solution in water containing a trace of acid, reprecipi- 
tation with the desired reagent, washing with alcohol and ether, and 
drying to constant weight. Bang's Statement of " characteristic 
properties " of histon was based upon the water-solution of the 
products precipitated by sodium hydroxid or sodium chlorid. His 
analyses were based on the ammonia-precipitated product. (See 
pages 424 and 426. ) 

G. Para-histon: Fleroff, 1899 (9), Preparation I. 
Minced thymus glands were treated with alcohol and ether, and the 
residue extracted with 2 per cent. sulfuric acid Solution (100 gm. of 
thymus to each 1,000 c.c. of acid Solution). The filtered acid ex- 
tract was then precipitated with three volumes of alcohol, the 
precipitate dissolved in hot water, and the Solution heated with 
sodium picrate. The histon picrate was then reconverted into the 
Sulfate by treatment with 2 per cent. sulfuric acid Solution and 
ether, and reprecipitation with alcohol. This process was repeated 
twice. The final precipitate was then dissolved in water, f reed from 
Sulfate with barium hydroxid, and excess of barium precipitated 
with carbon dioxid. To this turbid, viscid liquid was then added 
an equal volume of alcohol, and ammonia, and the liquid filtered. 
Excess of alcohol added to the filtrate precipitated para-histon. 
This was still further purified by dissolving in water and repre- 



1913] Walter H. Eddy 423 

cipitating with alcohol-ether. Preparation II. {Levene method). 
After transformation to picrate, and removal of picric acid with 
sulfuric acid as detailed above, the histon was precipitated from the 
sulfuric acid Solution with ammonia. The filtrate containing the 
para-histon was then precipitated with alcohol, the precipitate puri- 
fied by Solution in hot water, and reprecipitation with alcohol-ether. 
The product was then dried to constant weight. Fleroff determined 
the properties of his material in studies of the alcohol precipitate 
and its water-solution. 

H. Gadus histon: Kossel and Kutscher, 1900 (16). Dry 
codfish sperm was extracted many times with hydrochloric acid 
(20 c.c. conc. hydrochloric acid Solution to each liter of water), 
the combined acid Solutions then filtered, histon precipitated by 
Saturation with sodium chlorid, and purified by dialyzing against 
running water until free from sodium chlorid and a clear water- 
solution obtained, which was then precipitated with ammonia, and 
the histon washed with ammonia-water, alcohol, and ether, and 
dried to constant weight, 

I. LoTA histon: Ehrström, 1901 (8). Dry sperm of the 
burbot was extracted with conc. hydrochloric acid Solution at room 
temperature for an hour, the acid extract treated with 3-4 volumes 
of water, the filtrate neutralized with sodium hydroxid and diluted 
with five volumes of water to precipitate the histon. This product 
was purified by digesting the precipitate on a water bath with 0.5 
per Cent, hydrochloric acid Solution, precipitating the histon with 
ammonia, washing the material with water, alcohol, ether, and 
drying to constant weight. Ehrström used a hydrochloric acid 
Solution neutralized with sodium hydroxid for the determination of 
properties. His analyses were based on the ammonia-precipitated 
product. 

J. Hen blood histon: Ackerman, 1904 (i). {Plenge's 
method). The hen blood was centrifuged with 0.9 per cent. sodium 
chlorid Solution for the Isolation of the corpuscles, which were 
extracted with alcohol for the removal of pigment and again centri- 
fuged free from alcohol, and dried with alcohol and ether. Histon 
was obtained by extracting the dry material with i per cent. hydro- 
chloric acid Solution, precipitating with ammonia, and purifying by 



424 



Histon and its Preparation 



[April 



washing with alcohol and ether, and drying to constant weight. 
Ackerman used the ammonia-precipitated product for his analyses. 

K. Centrophorus histon: Kossel and Kutscher, 1906-7 
(15). Report was given of the preparation but no details as to 
method or properties. 

L. Reviews. The available reviews merely summarize the 
methods of preparation already in the literature. Oppenheimer 
(1909) recommends a method of preparing histon which is essen- 
tially that of Lihenfeld (19).^ Abderhalden (1909-10) recom- 
mends the method of Kossel and Kutscher (16).^ In our experi- 
ence both of these methods result invariably in a product that is 
practically insoluble in water. The only other method reported in 

TABLE I 

Data pertaining to percentage elementary composition of histons 



Kind 

1. Goose-blood (12) 

2. Goose-blood (12) 

3. Thymus (19, 2) 

4. Salmin-albumose (22) 

5. Globin (23. 2)^ 

6. Arbacin (21) 

7. Scombron (2) 

8. Para-histon (9) 

9. Gadus (16) 

10. Lota (8) 

11. Hen-blood (i) 

12. Centrophorus and 

Spharechinus (15) 



Precipitated by 



NaCl 
NH4OH 
NH4OH 
NaCl or 

(NH4)2S04 

NH40H 

Alcohol 
NH4OH 
Alcohol 
NH4OH 

NH4OH 

NH4OH 



C 



50.88 
50.90 
52.31 
52.14 
52.34 
52.31 

51.21 
59-47 



49.86 
51-91 



H 



7-05 
7.16 
7.06 
7.20 

7-31 

7.60 
7.20 



7-23 
7-31 



O 



23-44 



20.32 



N 



17.77 



8.46 
7-42 
8.35' 

7.64 
6.80 
6.89 
5-91 
9-79 
7-97 
8.65 
8.64 
6.46 
6.49 
8.31 



No data given by Kossel. 



0.42 
0.79 



0.00 



Ash 



0.52 
0.6s 
0.66 



0.52 
0.84 



^ Extracted minced, fat-f reed, glands with water and precipitated with acetic 
acid. Histon was extracted with 0.8 per cent. hydrochloric acid Solution and 
the histon precipitated with ammonia. 

*The fat-f ree glands were extracted with water and sufficient hydrochloric 
acid was added to make the strength of the acid 0.8 per cent. The filtered 
extract contained the histon, which was precipitated with ammonia. 

* Second figure for N is that of Bang and Fleroff. 

* All determinations by Bang except second N. 



I9I31 



Walter H. Eddy 



425 



the literature is that of Lawrow (18). Ammonia is used to precipi- 
tate the histon and purification is secured by redissolving- in liydro- 
chloric acid Solution and reprecipitating with ammonia. It intro- 
duces no new features. 

III. PROPERTIES OF HISTONS 

1. Results of elementary analysis. A summary of the avail- 
able analytic data is presented in Table i. 

2. Solubilities. The solubilities of the histon products are in- 
dicated by the data in Table 2. 

TABLE 2 

Data pertaining to the solubilities of histons 



Kind 



1. Goose-blood (12) 

2. Goose-blood (12) 

3. Goose-blood (2) 

4. Thymus 

5. Thymus 

6. Salmin (22) 

7. Arbacin (21) 

8. Globin (23) 

9. Scombron (2) 

10. Gadus (16) 

11. Lota (8) 

12. Para-histon (9) 

13. Hen-blood (i) 

14. Centrophorus and Sphäre- 

chinits (15) 



Precipitated by 



NaCl 

NH4OH 

NaOH 

NH4OH 

NaCl 

NaCl 

Alcohol 

NH4OH 

NaOH 

NaCl 

NH4OH 

Alcohol 

NH4OH 



Reagents 



S* 
I 

s 
I 

s 
s 
s 
s 
s 

S5 

I 

s 



I* 

I 

I 

I 

I 

I 

I 

II 

I 

I 

I 

I 

I 



ffio 
f~) '' 



I 
I 

s 

Si 
Si 

Sä 
81 
I 
I 

Si 

s 






X ° 

2: 



I 
I 
I 

I 
I 
I 

S3 

I 

I 

I 

I 

s 
I 



t/1 ü 
CO 



Si 
Si 

s» 
s» 

SI 
SI 
SI 

s 

SI 

SI 
SI 
S 



(No data given by Kossei). 



S 
S 
S 

S2 

S 
S 

s 
s 
s 
s 
s 
s 
s 



S Ol 

3 



I 
I 
I 
I 
I 
I 

I 
I 
I 
I 

s 



* S = soluble ; I := insoluble. 

* Soluble in excess of the reagent. 
"Becomes insoluble if allowed to stand. 

' Precipitate dissolves in slight excess of ammonia. 

* Alkali precipitates (30 per cent.) water Solution. 

" Kossei States properties are same as those of ordinary histon. 
properties are assumed to be identical with those of i. 



Hence the 



426 



Histon and its Preparation 



[April 



3. Characteristic precipitation reactions (Bang). Table 3 
presents a summary of the characteristic precipitation tendencies of 
histons, as specified by Bang. 

TABLE 3 

Data pertaining to the precipitation of histons (Bang) 



Kind 



1. Goose-blood (12) . . 

2. Goose-blood (2) . . . 

3. Thymus 

4. Thymus 

5. Salmin (22) 

6. Arbacin (21) 

7. Arbacin (21) 

8. Globin (23) 

9. Scombron (2) 

10. Para-histon (9) . . . . 

11. Gadus (16) 

12. Lota (8) 

13. Hen-blood (i) 

14. Centrophorus and 

Spharechinus (15) 



Nature of product 
in aqueous Solution 



NaCl ppt. 
NaOH ppt. 
Histon-HCl 
NaCl ppt. 
Histon-HCI. 
Histon-H2S04 
Ale. ppt. (NH4OH 

sol.) 
NH4OH ppt. 
NaOH ppt. 
Ale. ppt. 
NaCl ppt. 
Histon-HCI 
Histon-HCl 



Reagents 



B 

ÄS 
X 



. c 



Pi 
Pi 

p4 
p4 
p2 

N6 

N6 

pl 

p2 

N 
P 
Pi 



p2 

p2 
p2 
p2 
p2 

N5 

N6 
p2 
p2 

N 

p2 
p2 

P 



p* 

P 

P 

P 

P 



P 
P 

N 
P 

N 



N* 

N 

N 

N 

N 



N 
N 
N 
N 

N 



No data given. 



N 



2 n 

e s 



p3 

P 

P 

pe 
P« 

N6 
p6 



o « 

E- 

5 o 
2 E 



p3 

P 

P 



p3 
P« 



p3 



ps 

P 
P 
P 
P 



P 

P« 

P 



o o 






-■ 

_ 3 



p3 



p3 



pe 



P 
P 
P 
P 



P 
P 
P 
P 
P 
P 



4. General precipitation reactions. Additional general data 
on the precipitation of histons are given in Table 4. 

5. Color reactions. Available data pertaining to the responses 
of histons to protein color tests are indicated in Table 5. 

* P = precipitated ; N = not precipitated. 
^ Soluble in excess of the reagent. 
''Insoluble in excess of the reagent. 

* From neutral but not f rem alkaline Solutions. 

* Thymus is apparently insoluble in ammonia in the absence of salts pro- 
vided only a small amount of ammonia is used to precipitate it. Once formed 
a large excess of ammonia can be added without dissolving the precipitate. 

" No precipitate with ammonia unless the Solution is very concentrated er 
alcohol is present. 

' In neutral or weak alkaline Solution. 



I9I3] 



Walter H. Eddy 



427 





V 


piOE Dinn-EX 


Qh Oh a, 1 1 d, Ol 1 Hl 




'0 

u 

0, 



c 

II 


^-» 

CS 
'Qi 

"D 
u 
u 

a. 
II 

Ph 
* 


^ Complete on Saturation. 

"Soluble in excess of reagent. 

'Ammonia Solution of globin is less easily precipitated by alcohol. 

*In cold, not in hot. 







pp-B DUSOBJOiqOUX 


Cli Oh &< 1 1 H, d, 1 0« 1 


•0 

1 




pp-B OUOIJ 


p ■ p . p ■ 1 Q. Q. [^ 1 p. Q. 




piOB oipqAiouioqdsoq<j 


Ol d dl 1 1 d dl 1 d 1 


vi 




ppB onsSutiqoqdsoqj 


dddj dddddj 


>1 




ppB oqaov 


222 1 1 2 1 1 22 


rt 

E 




♦OS'H 


222 1 1 d 1 222 


ü 

(3 




DH 


222 1 1 d i 1 22 


V 

•0 

C 


*0 


31E533B tnnipog 


c^ 12: 1 1 1 1 1 1 1 




'(HO)«D 


d d d 1 1 d d 1 d d 


CO 




HO^M 


dddd |dd2dd 


2 


^O^D'^I 


1 1 1 1 (^ 1 12 1 1 





Ö 


^ONSh 


1 12 1 Miliz 


••* 


*OSn3 


1 1 Z 1 i Z II 1 2 




5J 


^ID^J 


1 |Z 1 12 1 1 12 


.- ^ 




(q) 3;e33Dv qd 


222 1 i 1 1 1 2a. 


■^ s 

rt 'Ö 




(u) 3jE5aov qd 


222 1 1 2 1 1 22 


^ ■§ 


W Ö5 


'ID^H 


222fo i 1 0.22 1 


.2 S 




"ID^O 


2^2 1 1 2 1 1 1 1 




ID'-HN 


dd2 1 1 22 1 1 1 


C J3 


5 


^OD'^'^N 


d d 1 1 1 d 1 d d 1 






*OS''(^HM) 


dddd Iddldd 


rt ^ 
«3 3 




*ossk 


d d d 1 1 d 1 1 d 1 s 


-4-* .IM 




ID^N 


dddd |ddi<dd ^ 


, fc- 


•"*-» 
Q 


J3qw-Ioqo3iY 


dd2| dd|ddd 'S 


Hl« 
•*- 


loqooiv 


* d2 1 dd 1 O-Ad Z 




^ 
^ 


s; C.2 

— S 1/ *-• 
Z-c - " 
rt er 


-^ „ 

.-s.au ^ g-g. .u 

rt rt .2 .2 .23 ffi rt u Ifl .22 
2 2 K K < ffi 2 2 < 2 a 


-"Sä 
c i <u 

— tn J3 

"Sb <+- rt 






•0 

s 

Ü 


88 

2 3 2 

> 

x: 

w pj f» 


: : :^3 : 

NN f^ - 

ö 1 .S ^ 2 » 5 


'S ° 

M 
M 


U] Ui W 

rt a.^ 
-^-' (-1 
(u t; _g 

<u JJ 

> 

et 



428 Histon and its Preparation 

TABLE 5 

Data pert