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O- C « " o ,-^vo U co cM U cS S * s « > «■• ■a u N co co *-■ PLANKTON 31 Siphonein Siphonoxanthin Fucoxanthin Neofucoxanthin Diatoxanthin Diadinoxanthin Dinoxanthin Neodinoxanthin Peridinin Myxoxanthin Myxoxanthophyll Key Pigments in Algal Photosynthesis CHLOROPHYLLS Types a, b, c, d, e CAROTENOIDS Carotenes Types a> 0> c Xanthophylls Lutein Zeaxanthin Violaxanthin Flavoxanthin Neoxanthin PHYCOBILINS r and c Phycoerythrins r and c Phycocyanins The distribution of these pigments in algae is rather variable. The commonest chlorophyll is the type a, just as in higher plants. Chlorophyll b, also present in higher plants, occurs only in the Chlorophyceae and the Euglenineae. The other classes contain chlorophylls c, d, and e. Of the carotenes, the /?-type is most abundant. The «-carotene, typical Chloro- phyceae and the higher plants, is not found in some algae. In Bacillariophy- ceae, for example, it is supplanted by e-carotene. Each algal class also has its own characteristic xanthophyll, as may be deduced from the num- ber that have been identified. Phycobilins, on the other hand, occur only in the Rhodophyceae and the Myxophyceae, the former containing mostly phycoerythrin, the latter phycocyanin ; the origin of these related pigments in either Rhodophyceae or Myxophyceae is indicated by the designation r or c, respectively. Granick and his associates reported199-200 in 1953 on the protopor- phyrin precursors produced by Chlorella mutants. They have been par- ticularly interested in the relationship between chlorophyll and hemin, seeking information on porphyria and blood formation in human beings. Delia Rosa,201 using C14, demonstrated that Chlorella vulgaris was able to synthesize chlorophyll from acetate and glycine ; he also showed that chlo- rophylls a and b are not interconvertible. Jorgensen41 in Venezuela demon- strated to one of us (M.S.) the red blood wiggler Chironomus growing on the sides of his Chlorella culture containers; he suggested that there might here be a conversion of chlorophyll to heme. French and Young202 indicated that phycocyanin may be an intermediate in the resonance transfer of energy from phycoerythrin to chlorophyll. 32 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE The photosynthetic role of the various pigments can be estimated by comparing the absorption spectrum with the "action spectrum" or photo- synthetic capacity of light at various wave lengths.40'41 Photosynthetic efficiency is usually measured in terms of oxygen production. The chloro- phyll absorption bands are chiefly in the blue, blue-green, red, and infra- red ranges. The yellow-orange color of the carotenoids results from absorption in the blue and green. The chlorophylls and phycobilins are often strongly fluorescent in solution. Tucker203 has even used pigment extraction with acetone as a method of quantitating phytoplankton. Despite all the emphasis on photosynthetic capacities, it must be noted that not all algae contain pigment. Beijerinck180 long ago noted the natural occurrence of yellow and colorless colonies of Chlorella variegata on culture media. Bogorad and Granick200 produced colorless, pale yellow, and light green mutant colonies of Chlorella vulgaris by radiation. Butler204 exposed Chlorella pyrenoidosa to ultraviolet light and obtained colorless mutants without chlorophyll. He also noted that similar perm- anently colorless forms of Prototheca have been observed in nature, and that they were probably mutants from Chlorophyceae. Very recently Tolbert and Zill205 reported photosynthetic studies with protoplasm ex- truded from Chara and Nitella. They found activity 12-15 per cent that of the whole cell, as measured by C14 fixation. Both protoplasm and cut cells reduced carbon dioxide in light to sucrose and hexose phosphate. Dark controls fixed C14 into products associated with plant respiration, but it is not clear whether they represented true photosynthetic pigments. In fact, the whole sequence of biochemical synthesis in the dark or with- out pigment remains rather obscure. This is particularly so in the light of Duysens' claim206 that light energy is convertible only through chloro- phyll a or b (he maintains that in diatoms, for example, carotene must be converted to chlorophyll a.) Radiation and natural mutation are not the only methods of producing non-pigmented algae. Dubie207 produced chlorophyll-deficient Chlorella vulgaris with streptomycin. And even strong radiation does not always effect alterations. Blinks208 reported there was no significant change in composition or function of marine algae one year after the atomic explo- sions in the Bikini area. The only exception was an increase in catalase, possibly due to an increased hydrogen peroxide concentration secondary to radiation. Algal cells generally do not grow old and die; instead, they produce more cells by binary fission. However, not all algae grow by cellular PLANKTON 33 division. It has not been widely realized that sex may play a role even in this remote corner of the cosmos! Sexual union of two gametes to produce a zygote does occur, particularly among the Phaeophyta and Chlorophyta (Chlorella and Scenedesmus are notable exceptions.) Either or both gametes may be flagellated, or mating may be achieved by ameboid motion. Gilbert M. Smith209 notes that algal sexuality was initially de- scribed in 1858 in Oedogonium. In 1926, Jollos demonstrated that the heterothallic green alga Dasycladus excretes sexual substances which in- fluence the movement of gametes toward each other. Since then, much experimentation has been carried on with Chlamydomonas eugametos, a unicellular green alga. Motility is absent in cells grown anaerobically in the dark, but it is revived by filtrates from cells grown in light. The sub- stance responsible is reported to be crocin, an ester of the sugar genti- obiose combined with the carotenoid pigment crocetin. There are also substances affecting maleness and femaleness; these are activated by ex- posure to light, weakened by dilution. The virilizing factor appears to be s of r anal, the feminizing one a gentiobiose of safranal. There are all degrees of sexual activity, depending upon the concentration of activating substances and exposure to light. There are also varying degrees of male- ness and femaleness, even intermediary forms, depending upon the ratio of opposing substances. Similar findings were reported by Sager and Granick211 in 1954. They noted that depleting the nitrogen supply in culture media of Chlamy- domonas reinhardi encourages the differentiation of ordinary cells to gametes. If nitrogen is added, the process is reversed. Light apparently acts only indirectly via photosynthesis, and it is not obligatory for zygote formation of dark-grown nitrogen-depleted cells. Much has been learned about the metabolic characteristics ol algae by observations of cultures. Chlorella has been widely used in photosynthetic experiments especially because it can grow under such varied environ- mental conditions. The rate of algal growth is often measured by "genera- tion time" — the time required to double the number of cells in a culture.41 There are two major phases of growth, one chiefly of cellular multiplica- tion, the other of storage of reserve materials. In the first phase, proteins predominate; in the second, fats and carbohydrates. The main factors influencing algal growth are: available radiant energy, carbon dioxide concentration, temperature, and composition of nutrient substrates. The exact channeling of the various nutrients may depend upon the amount of available nitrogen, or upon the presence of enzyme systems. Chlorella 34 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE extracts, for example, have been shown to contain transaminase systems which aid the transfer of amino groups to either pyruvic or rf-keto-glutaric acid.212 The most active enzyme systems in algae are those of aspartic and glutaric acids, and alanine. Spoehr and Milner41 demonstrated that the composition of Chlorella pyrenoidosa could be varied specifically simply by controlling cultural conditions. In over 300 pure cultures, it has been possible to produce the following wide range of variations in composition and R-value: Protein: 7.3 to 88.0% Carbohydrate: 5.7 to 38.0% Lipid: 4.5 to 86.0% R-value: 37.92 to 63.33 The amount of radiant energy present markedly influences the rate of growth of a culture, either directly or by increasing the nitrogen absorp- tion. The latter is certainly of critical importance, since a Chlorella cell can no longer undergo fission when the nitrogen content falls below 1.6 x 10-13gm. As has already been indicated, the protein content of microscopic algae may vary widely; its level may be grossly estimated from the nitrogen content. There is a definite similarity in the protein content of Chlorella and the leaves of many higher plants. Under optimal cultural conditions, a protein level can be achieved in Chlorella which exceeds that of the best vegetable substances used as animal feed. This includes such materials as Brewer's yeast, torula yeast, soybean meal, dried skimmed milk, and wheat. Not very much is known of the specific nature of algal proteins, except that there is often a resemblance between their amino acid patterns and those of higher plants.213"215 The nutritional value of Chlorella is based not merely on its high protein content, but also on the fact that it con- tains all the essential amino acids. Methionine is present in only small amounts, but this deficiency also occurs in many vegetable proteins. The essential amino acid Biological Index of Chlorella vulgaris has been calcu- lated to be 62 in a pilot plant assay, as compared with an arbitrary value of 100 for whole egg protein. The Index of most animal proteins ranges from 80 to 90, while cereal proteins are usually between 60 and 80. Thus Chlorella is roughly comparable to white flour, corn gluten, and peanut meal. An amino acid assay of dried Chlorella is given in Table IV. Although most algae are able to utilize wide varieties of basic inorganic nutrients and to synthesize from them the more complex metabolites re- quired, some are not so versatile. They may not be able to synthesize all PLANKTON 35 TABLE IV.— Amino Acid Assay of Dried Chlorella Pilot Plant Laboratory Nutrient Sample Sample Torula Yeast Crude protein 44.00% 40.0096 — Arginine 2.06 2.39 3.61% Histidine • 0.62 0.65 1.31 Isoleucine 1.75 1.69 3.75 Leucine 3.79 1.99 3.57 Lysine 2.06 2.43 4.14 Methionine 0.36 0.57 0.84 Phenylalanine 1.81 2.14 2.41 Threonine 2.12 1.91 2.58 Tryptophane 0.80 0.41 0.66 Valine 2.47 2.67 2.98 Glycine — 2.20 0.22 the amino acids from ammonia and the requisite carbon fractions, and they must thus depend at least in part upon an exogenous source of some amino acids. The deficiency is not of the amino acid per se, but of the capacity to form amino groups from ammonia. Thus, for proper growth, Euglena deses requires aspartic acid, while Chlamydomonas cblamydogama needs histidine and aspartic acid.40 Nucleic acid has been demonstrated by staining reactions to be present in algal cells. Jeener216 reported that Polytomella caeca had 6 to 10 per cent of its protein as ribonucleic acid in actively growing cells. He indi- cated that there was great variability in the synthesis of nucleic acid, and that this was unrelated to the rate of general protein synthesis and cellular multiplication. Szafarz and Brachet217 indicated that formation of ribo- nucleic acid can proceed independently of the nucleus in Acetabularia mediterranea. Studies by Goryunova218 have indicated the presence of mucins in Oscillatoria. The assimilation of nitrogen by algae is important not only with respect to amino acids, but also in the synthesis of various carbon compounds. As in the case of certain bacteria, many autotrophic algae are characteris- tically capable of nitrogen fixation. Nitrogen may be absorbed as the element, or as nitrate or ammonia. Frank,219 in 1889, first reported the possible nitrogen-fixing properties of certain algae, but this was not proven until 1928. 220 Since then, further corroboration has come from Kjeldahl determinations and tracer studies using nitrogen isotopes.40 So far, over 20 species of Myxophyceae have been proven capable of fixing nitrogen. Aspartic acid, glutamic acid, succinamide, asparagine, and glutamine can 36 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE serve as the sole sources of nitrogen for various green algae via deamina- tion or transamination. It is probable that nitrogen enters the general metabolic pool most often in this manner. Nitrogen-fixing algae can contribute to the fertility of the soil, especi- ally in tropical areas. A notable example is the utilization of blue-green algae in the Usar area of northern India to decrease the excessive alkalinity of the soil.15 The algae thrive during the rainy season and decrease the pH of the soil from 9.0 to as low as 7.0, and thus make possible the growing of field crops during the dry season. This resembles somewhat the employment elsewhere of large seaweeds as direct fertilizer. The cultivation of rice is also aided by microalgae. The latter flourish in the wet rice paddies and, by fixing atmospheric nitrogen, enable peren- nial cultivation without additional fertilizer. Nitrogen fixation here may be greatly enhanced if bacteria are also present and contribute their fixation activities. In addition, the algae, by producing oxygen, help to aerate the rice plants. Watanabe221 reported in 1951 that 13 species of 643 blue-green algae from Far East and South Sea area rice fields were capable of atmospheric nitrogen fixation, chiefly in warmer climates. Most of them belonged to the genera Tolypothrix and Nostoc, some to Schizo- thrix, Calothrix, Anabaenopsis, and Plectonema. Aspartic and glutamic acids and alanine were the systems mostly involved. A similar function was reported by Douin,222 who noted a symbiosis between the Cycadaces plant and Anabaena cycadeae, which fixes nitrogen for its host. Much less is known about the carbohydrates of microscopic algae than of the macroscopic.40-41 Whereas the latter may contain large quantities of carbohydrates in the cell walls (chiefly alginic complexes), the micro- algae have relatively scanty structural materials, and their storage products are lipid rather than starchy. Milner has isolated starch and sucrose from Chlorella pyrenoidosa. Broun found 17.8-20.2 per cent (dry weight) total carbohydrate in Scenedesmus obliquus. The substances included mainly insoluble polysaccharide; there were also small amounts of free reducing sugar, sucrose and other oligosaccharides, and water-soluble polysaccharides. Healthy algal cells, particularly in older cultures, liberate various or- ganic substances into their fluid environment. These consist primarily of nitrogenous substances and pentosans. It is probable that more specific knowledge of microalgal carbohydrates will emerge when higher carbo- hydrate contents are achieved by varying cultural environmental condi- tions. PLANKTON 37 The lipids represent one of the most important components of micro- scopic algae.223 Many investigators have confirmed Beijerinck's observa- tion in 1921 that fat droplets accumulate in diatoms as the cultures be- come old and the available fixed nitrogen decreases.40-41 This occurs, as we have indicated earlier, because nitrogen deficiency limits growth and favors the building of reserve materials. The ratio of lipid to carbohydrate is variable; it depends far less on algal genetic traits than it does on environmental factors such as nitrogen supply and enzyme systems. Also noteworthy is the fact that algal fat metabolism is but little reflected in gas exchanges. The lipid content differs not only from one species to another (the blue-greens have very little), but also may vary greatly within the same species with changes in cultural conditions. Chlorella generally contains 20-25 per cent fat, yet nitrogen starvation can increase the amount to as high as 86 per cent. Even during World War II, the Germans, cul- turing Chlorella, NHzschia, and Scenedesmus as potential sources of needed fat, attained lipid yields of 40-70 per cent. These comprised chiefly the triglycerides of stearic, oleic, and linoleic acids. Most of the fat in phytoplankton is unsaturated, with a high propor- tion in the 16 and 18 carbon series.41 Detailed analyses of Chlorella, for example, show 54-67 per cent unsaturated C-18 and 18-29 per cent C-16 acids. Unsaturation ranges from 1.60 to 2.25 double bonds per molecule. Palmitic is the main saturated acid, while stearic constitutes less than 4 per cent of the total fatty acid. As the lipid content of Chlorella rises, the unsaponifiable fraction decreases and the fatty acid portion increases ; at the same time there is a significant fall in the degree of unsaturation of the fatty acids. Chlorella fat resembles other unsaturated plant lipids and falls into the chemical classification of "drying oils." Sterols have been obtained from most of the algal groups, excepting the blue-greens. Klosty and Bergmann224 reported in 1952 that chondril" lasterol is the principal sterol in Scenedesmus obliquus, while ergosterol predominates in Chlorella pyrenoidosa. Chondrillasterol may be remem- bered as having been favorably considered a few years ago as a starting point for cortisone synthesis. Table V offers analyses of the various lipids in several samples of Chlorella. Phytoplankton, like the macroscopic seaweeds, has a high vitamin con- tent. It has been determined that most of the known vitamins are present in Chlorella and related microalgae. The quantities are eminently ade- quate for the algae to be used as food (for man or animals), but not if 38 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE TABLE V.— Analysis of the Lipids of Chlorella Analysis Chlorella lot no. 12 3 4 1. Total lipid {% of Chlorella) 23-37 33.17 62.96 75.51 Composition of total lipid 2. Fatty acids {% of lipid) 28.0 49.5 83-0 86.8 3. Unsaponifiable fraction {% of lipid) 12.0 7.7 3\3 3-3 4. Water-soluble, after saponification (% of lipid) 60.0 42.8 13.7 9-9 5. Calculated fat (% of Chlorella) 6.85 17.2 54.7 68.6 Analysis of total fatty acids 6. Palmitic acid, Cl6 (%) 16.6 10.9 7.9 11.4 7. Stearic acid, C18 (%) 0.4 4.1 3-9 3.5 8. C16 unsaturated acids (%) 29-1 18.3 27.2 18.0 9. C18 unsaturated acids (%) 53.9 66.7 60.9 67.1 10. Total fatty acids (%) 100.0 100.0 99-9 100.0 11. Equivalent weight 269.5 273.6 272.7 274.1 12. Iodine number 163.1 143.8 143.6 125.3 Degree of unsaturation 13. Cl6 plus C18 unsaturated acids -4.4H -3-6H -3-7H -3.2H 14. Cl6 unsaturated acids -4.1H -4.4H 15. C18 unsaturated acids — 4.5H -3.4H they are intended purely as a source of vitamin concentrates. In the latter case, synthetic methods are more economical. Microscopic algae are the ultimate basic source of vitamins in fish, and the finding of large quantities of Vitamin A in fish livers was indeed the stimulus for much of the study of algal vitamins.225 Several decades ago, it was shown that NHzschia closterium and other phytoplankton were good sources of Vitamin A and its carotene precursors.226 In the early 1930's, Jorgensen in Scandinavia considered using algal carotene to feed cows and to fortify margarine.41 Later, in Venezuela, he used dried plankton from Lake Maracaibo as a source of carotene and related unidentified substances as a highly efficacious food supplement in the therapy of lepers. Vitamin B1 is generally necessary for algal growth, and it has been demonstrated in most species in which it has been sought.40 Especially large amounts are found in Chlorella. Young cultures with actively- dividing cells contain more thiamin than do the older fat-storing ones. Thus, von Witsch obtained yields of 1.8 to 18.0 gamma per gram of dried substance from cultures of varying ages. While Chlorella and related algae contain dehydrogenase for direct PLANKTON 39 oxidation of pyruvic acid, there is also some alternative participation in the Krebs tricarboxylic acid cycle, as in vertebrate tissues. For this re- action, the co-carboxylase thiamin pyrophosphate must be present. An example is the great enhancement of pyruvic acid oxidation by the addi- tion of thiamin to the thiamin_deficient Prototheca. If Vitamin Bx is not present as such in the aqueous nutrient, it must be synthesized by the algae. Most are capable of so doing, but some cannot synthesize the thiazole, others the pyrimidine, radicles of thiamin. Thus the thiazole portion must be supplied to Polytomella ocellata, and the pyrimidine component to Euglena gracilis and to a radiation mutant of Chlamydomonas moewusii. The pyrimidine-thiamin requirement of the Euglena can be markedly de- creased by the addition of glutamate. Both thiazole and pyrimidine must be supplied exogenously for the growth of Polytomella caeca, Chilomonas paramoecium, and Prototheca zopfi. Riboflavin, like thiamin, has been demonstrated in practically all algae in which it has been sought. It is a component of the prosthetic group of the flavo-proteins, but its essentiality to algal growth has not been shown. Para-aminobenzoic acid is known to be necessary for the growth of only one alga, a mutant of Chlamydomonas moewusii. With the latter and with Nitzschia, para-a?ninobenzoic acid has the same antagonism to sul- fonamide inhibition of growth as with many bacteria. Aniline may replace para-aminobenzoic acid as a growth factor for Chlamydomonas with an efficiency of one per cent, but it does not neutralize the sulfonamide effect.40'227 Plankton paste contains considerable quantities of cyanocobalamin or Vitamin B12.40,41'228 Traces of this substance are necessary for the growth of various Euglenineae as well as other classes, such as Chlamydomonas chlamydogama of the Chlorophyceae. Vitamin B12 may be concerned in the synthesis of desoxyribosenucleic acid. Ascorbic acid is present in phytoplankton (especially Chlorella) in amounts comparable to those in lemon juice; however most of it is lost in drying and storage. Although little is said about the Vitamin D content of microalagae, feeding experiments have indicated good antirachitic activity. Indirect evidence comes from the potency of fish liver oils, which ultimately derive their Vitamin D from planktonic sources.229 As with seaweeds, the vitamin activity of phytoplankton is based upon ergosterol content. Substances with coagulant effect have been demonstrated by Dam, who determined the Vitamin K content of Chlorella as six gamma per gram of dried material.230 Phytoplankton has a great potential as a source of vitamins in human 40 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE nutrition. Though most of the studies have been with Chlorella, other microalgae are comparable. It has been calculated that a quarter pound of Chlorella will supply all the daily minimal human vitamin needs except for ascorbic acid. Table VI, taken from Burlew,41 details the vitamin content of two samples of Chlorella. TABLE VI. — Vitamin Assay of Dried Chlorella Pilot Plant Laboratory Vitamin Sample Sample Carotene (mg./lb.) — 218.0 Thiamin (mg. /lb.) 11.0 4.5 Riboflavin (mg./lb.) 26.2 16.3 Niacin (mg./lb.) 54.0 109.0 Pyridoxine (mg./lb.) — 10.4 Pantothenic acid (mg./lb.) 3.6 9.1 Choline (mg./lb.) — 1370.0 Biotin (meg. /lb.) — 67.0 Vitamin Bi2 (meg. /lb.) 45.0 10.0 Lipoic acid (acetate units/lb.) 1.5 — Algae require basically the same nutrients as higher plants — nitrogen, phosphorus, potassium, sulfur, magnesium, and iron. Unlike the higher plants, all except a few algae can dispense with calcium. Potassium and magnesium (and occasionally calcium) are important in the photosyn- thetic process because their bicarbonates can make available supplemental carbon dioxide. In addition, algae also have need for certain trace elements or micronutrients. Myers41 has specified algal requirements for manganese, zinc, calcium, boron, and copper. The exact metabolic role of these has not been elucidated. Neither has it been for the following substances, which have also been identified in algal cells: antimony, arsenic, beryllium, cobalt, germanium, iodine, lead, manganese, molybdenum, nickel, rubi- dium, silver, strontium, tin, titanium, tungsten, uranium, vanadium, and yttrium. (Does not this listing of elements sound more like a roll-call of our present-day electronics and metallurgical industries, and are not the biologic and the mechanistic worlds drawing closer?) It has been deter- mined that, because of selective absorption, the cellular content of manga- nese, strontium, uranium, and yttrium is above that of the surrounding media. The only known role of molybdenum is that of fixing nitrogen in Anabaena. Even though their functions are obscure, the addition of trace elements to algal culture media has become standard operating procedure. An excellent means for providing continuously adequate micronutrients are PLANKTON 41 the chelating or complexing agents. These are biologically inert cyclic organic compounds ("inner complex salts") resulting from the attach- ment of a group at two points to the same metallic atom. They prevent excessive absorption and precipitation. Ethylenediamine tetraacetic acid (EDTA) is the chelating agent most commonly used. When a given amount is added to a nutrient medium, it will permit the release of enough ions through mass action to provide for cellular needs. By forming easily reversible complexes of trace elements, it provides tonic concentrations at preferred levels throughout the life of a culture. Besides the trace elements, there are other unidentified factors some- times necessary for algal growth. It is common observation that some algae grow with ease in impure culture, yet do not flourish in bacteria-free media. Gymnodinium cannot be continuously subcultured without an organic factor extracted from soil. Similarly Cryptomonas ovata, Synura ovella, and Gloeotrichia ech'mulata demand the presence of certain soil factors. Also, Dity Ilium brightivelli requires, for good growth in artificial seawater, two organic substances from natural seawater. One of these is a specific sulfur grouping, the other a substance also found in yeast or algal extracts.40 The exact nature of these various needed supplementary compounds will doubtless become clearer with more experience in algal culture. Although mass culture of phytoplankton for food and special organic material has long been the subject of considerable discussional enthusiasm, little specific investigation was done until World War II.231 Most of the experiments have been performed with some species of Chlorella, Scene- desmus, or Nitzschia because they grow so fast and tolerate a diversity of cultural conditions. Chlorella pyrenoidosa has been an especial favorite because of its previous wide use in studies on photosynthesis. As listed by Burlew in "Algal Culture, from Laboratory to Pilot Plant,"41 the elements of a plant for growing algae on a large scale are relatively simple: 1 . A container with a transparent upper surface. 2. A means of circulating the culture medium, so that the algae do not settle. 3. A means of controlling temperature. 4. A means of introducing carbon dioxide and other nutrients con- tinuously. 5. A means of harvesting the algae almost continuously. 6. A means of preserving the harvest until used. 42 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE A great variety of apparatus has been employed to achieve the me- chanical requisites. In 1951, Jorgensen and Convit in Venezuela grew freshwater algae in unglazed baked red clay bowls exposed to the open air and hot sun. The nutriment consisted of a commercial fertilizer suspended in water. Evaporation from the porous sides of the bowls kept the tem- perature at 26° C. even in direct tropical sunlight. Von Witsch and Harder in Germany utilized nutrient solutions in glass tubes 3 x 30 cm. and 6 x 150 cm.; these were illuminated by 300-watt water-cooled lamps and supplemented with an air stream containing 0.5 per cent carbon dioxide. At the Carnegie Institution in Washington, greenhouse culture of Chlorella was carried on in five-gallon bottles in 1947; illumination, temperature, and carbon dioxide supply were controlled. This study led the following year to activation of a successful pilot plant for continuous Chlorella culture at the Stanford Research Institute. In 1949, Geoghegan in England tried out a variety of culture vessels: cylinders 15 x 1.75 inches; large tubes 4.5 feet x 2.75 inches; aspirator bottles 18 x 10.5 inches; and a tall outdoor plastic tank 4.5 feet high, 1.5 feet long, and 0.33 feet wide. He concluded that shallow horizontal tubes or troughs were preferable, and that sunlight is more economical than artificial illumination. Wassink and a group at the Agricultural University at Wageningen in Holland experimented with mass Chlorella culture from 1948 to 1950. Initially they used inverted half-liter glass- stoppered bottles with incandescent light. Later they tried outdoor sunlit square meter tanks with 300 liter capacity. The tanks were covered with glass to minimize contamination; carbon dioxide was supplied from a cylinder; and agitation was accomplished by a motorized stirrer. Algae were harvested after 5 to 7 days of growth. Subsequently they set up similar concrete tanks in a dark room and illuminated them with either daylight fluorescent bulbs or high-wattage incandescent lamps. They ob- tained a 12-20 per cent efficiency of light-energy conversion, and there- from concluded that excessive illumination could decrease algal growth under either artificial or natural conditions. Gummert and his colleagues, working in Essen in 1950-51, hoped to demonstrate that local conditions were suitable for large-scale outdoor and greenhouse culture that could take advantage of the huge amounts of waste carbon dioxide in the industrial Ruhr. They succeeded for the most part, but their deliberately nonsterile procedures resulted in troublesome contamination with foreign algae and protozoa. This could be overcome by using protozoa-resistant Scenedesmus, by altering the nutrient media, PLANKTON 43 by increasing the amount of illumination, or, in fact, by doing anything that promoted faster growth of the desired algae. Much was accomplished in America from 1950 to 1953 under the guidance of the Carnegie Institution of Washington. They helped sponsor such investigators as Spoehr, Milner, Davis, Myers, and Krauss, with much of the work done in the Department of Plant Biology at Stanford. Here studies were performed not only with various types of algae and nutri- ments, but also with differing forms of culture equipment. The latter included large bottles, rocking trays, and plastic and glass tubing. The culmination of the Carnegie studies was the establishment of a large pilot plant for Chlorella culture in Cambridge, Massachusetts, by Arthur D. Little, Inc., a consulting research and development organization. Three large-scale culture units were developed and studied. The last, and best, consisted of a massive plastic tubing in U-shape, set up on a roof-top with free exposure to sunlight. It was equipped with a centrifugal pump to circulate the culture, a measured inflow of carbon dioxide (5 per cent in air), a heat exchanger for cooling the culture, a harvesting system, and apparatus to freeze or spray dry the harvested material. From this study, it was concluded that with this type of pilot plant, 20 gm./sq. meter/ day could be realized, equivalent to 17.5 tons Chlorella per acre per year. Pilot plant studies were also carried on in Israel and in Japan. The former were begun in 1951 at the Hebrew University in Jerusalem under government sponsorship. The latter were done in Tokyo in 1952 at the Tokugawa Institute for Biological Research; Tamiya, the director of this group, spent some time as guest worker in the Carnegie Institution labora- tories in Stanford through a fellowship supported by the United States Office of Naval Research. Experiments on mass culture of algae are also being carried on by seg- ments of private industry. An example is the reported work of Pruess and his colleagues232 for a pharmaceutical concern on algal culture in carboys and in deep tank fermentations. Kindred to the latter is the growing of algae in so-called "oxidation ponds," with the double purpose of sewage disposal and formation of algal protein suitable for feeding. Renn233 has estimated that domestic sewage from a community of 10,000 could pro- duce nearly 1,400 pounds of protein daily if properly treated. Oswald and his colleagues234 at the University of California are also enthusiastic proponents of this method. One eager college professor22 has already designed a house with an algal pond on its flat roof. No comment is offered on the monotony of having to stick to one brand of sewage ! 44 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE The enthusiasm for culturing phytoplankton for feeding is based on reports that it is innocuous and that it possesses great nutritional value. These have been well summarized by Fisher and Burlew.41 Mention has already been made of Jorgensen's feeding plankton soups to lepers in 1941, with a resultant improvement in weight and energy. These com- prised cultures of Chroococcus, Homocystinea, Oocystis, Ankistrodesmus, Chlorella, and Scenedesmus. More recently, Jorgensen told one of us (M.S.) that an algal broth, when fed to mice with skin cancer, brought about an appreciable decrease in the size of the lesions; inexplicably, however, test animals maintained on such a diet did not haye a long life span. Currently Jorgensen is feeding his phytoplankton in rather palatable compressed pills. Another early feeding experiment was that in 1949 at the Stanford Research Institute. Rats fed on a diet containing a third Chlorella pyrenoi- dosa had a weight gain only one-third that of a control group. This has been attributed either to the unpalatable flavor or to the mechanical diffi- culty resulting from the hygroscopic nature of the food preparation. Henry, at the National Institute for Research in Dairying, in England, fed freeze- dried Chlorella prepared by Geoghegan to young rats. No adverse effects resulted after four months on a 17 per cent Chlorella diet. In a four- week comparative study, the protein efficiency of Chlorella appeared slightly superior to that of dried brewer's yeast and peanut meal, but inferior to that of skim milk. In 1951, Combs,235 at the University of Maryland, tested New Hamp- shire chickens on Chlorella provided by the Carnegie Institution. A marked increase in growth and an improvement in feed efficiency resulted from adding 10 per cent Chlorella to a basal diet in place of soybean meal. This was attributed to the high vitamin content of the alga. However, a growth depressant effect was noted after extended feeding because of im- pacted beak conditions. This occurred much sooner with 20 per cent Chlorella feedings. Similar results were obtained in 1952 with chicks when vacuum-dried Chlorella was tried by a feed supply concern. Most of the difficulties encountered were apparently due to excessive water absorption by the algae. Other methods of preparation would seem to be indicated. Tamiya22-41 has included substantial amounts of Chlorella in preparing highly acceptable bread, noodles, soup, tea, and ice cream. He has also used hydrolysates of dried algae as a substitute for soy sauce. His students and even his colleagues admitted no ill effects whatever from repeated ingestion of these concoctions. PLANKTON 45 The palatability and acceptability of phytoplankton as a human food are dependent upon many factors. These include especially appearance and taste. These depend, in turn, upon whether the material is fresh or dried; also, whether it is eaten as such, or mixed (and diluted) with more familiar foods. Doubtless much improvement can and will be made in all these respects for a greater acceptability. Certainly these items must be worked out before plankton is really the "food of the future."236 3. Medical Aspects a. Water Supply and Sewage Disposal Despite all the hopeful reports on their merits, it must be emphasized that phytoplanktonic organisms can exert adverse effects upon the waters in which they flourish. These include not only changes in appearance, odor, and taste of the water, but also changes in other aquatic organ- isms.237243 Even more important are the varying degrees of intoxication and reaction in animals and humans who ingest or come in contact with these waters. The potential economic losses are self-evident. Similar to the striking colorations of the seas previously mentioned (e.g., Red Sea, Vermilion Sea) are the alterations in appearance of fresh water bodies. "Water bloom" is present when extensive growths of micro- scopic algae blanket the surface of a lake, pond, or stream. In Europe the phenomenon has been variously called "Wasserbliithe," "fleur d'eau," and "flos aquae"; in some North Central parts of the United States, the ancestral Dutch "vasserbloom" designation is used. Under favorable cir- cumstances, microscopic algae, especially the Myxophyceae, can grow so rapidly that the water assumes the color of the organism. And these colors are indeed variegated. The Oscillatoria and Cylindrospermium produce a green slime and a bluish soup, respectively. Chlamydomonas and Pleuro- coccus are a brilliant green. Ceratium hirundinella presents a rusty color and a powerful stench — perhaps the basis of the Biblical story (Exodus: 7) of the turning of the Nile into blood and the death of the fish therein. "Red water" and "red snow" in Australia are due primarily to Myxophy- ceae, but desmids (Conjugata) may >je a factor. Both "red tide" and "yellow tide" have been reported along the coast of the Gulf of Mexico, while two major outbreaks of "red water" occurred in Gokasho Bay, Japan, in 1933 and 1934, from Gymnodinium mikimotoi. Whereas the colorations are interesting and occasionally even weird, the odors arising from aquatic algal growths are more important, both esthetically and economically. They affect water supply and sewage dispo- 46 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE sal. Odors and tastes may arise from the oils of living algae; from prod- ucts of active photosynthesis; from products of decomposition; and from the death of fish and other aquatic organisms. The oils of living algae are often unpleasantly odorous, each oil being characteristic of the gene- rative organism. The smells may be fishy, aromatic, or grassy. Their disagreeable character may be heightened after planktonic death, when the frail oil globules are more widely dispersed. The odorous photo- synthetic substances liberated during very rapid growth are ill-defined, but the offending materials present after algal decomposition are better known. They include hydrogen sulfide and other sulfurous compounds, phosphorous and nitrogenous substances, methane, ethereal sulfates, and other volatile gases. Fish and other aquatic organisms may die from suffocation, or from toxic substances such as hydroxylamine resulting from the breakdown of algal proteins. These dead animals contribute a liberal stench to the already odoriferous situation. Often enough, the chemical treatments applied to kill off offending algae aggravate the situation even further. The treatment, such as chlorination, kills the organism and liber- ates the taste and odor-producing materials, and chemical combination with chlorine may favor the formation of additional smelly compounds. The most troublesome algae in lakes and reservoirs are the blue-green ones, which grow rapidly and float high in the water, forming surface scums. Greatest growth takes place in waters which are shallow and warm, and in those rich in nitrogen, phosphorus, and loosely-bound carbo- hydrates. The principal causes of bad odors and tastes are the Myxophy- ceae and Diatomales, which are less potent oxygenators than are other algae. When unpleasant odors affect ordinary lakes or ponds they merely detract from the beauty of nature. But when they occur in reservoirs of water intended for daily drinking, they present a trenchant problem. Even mild alterations in smell or taste may be psychologically unacceptable to the average person. As Howard and Berry238 have pointed out, odors caused directly or indirectly by living plankton or by the decomposition of vegetable growths are immediately ascribed by the layman to con- tamination with sewage. This is not too strange, since the decomposing plankton floats on the water, then is thrown onto the shore, where it forms odorous, blackish, discolored masses. It is difficult to convince people that the odor itself signifies no health hazard. When living algae are present in the drinking water, there may be a mild initial taste, but a bitter after- taste ; when the organisms are dead, there is a strong first taste, but practi- PLANKTON 47 cally no aftertaste. Measures to control algal growth244-248 in reservoirs include periodic chemical treatment (with algicides such as copper sul- fate), removal of accumulations from the shores before decomposition occurs, and restricting the food supply of the algae by removal of tree branches, leaves, and other floating debris. While removal or eradication of algae is desirable in reservoirs, quite the opposite is true in sewage disposal units. Oxidation ponds have been widely used for secondary sewage treatment. As summarized by Oswald and his colleagues,234 oxidative sewage treatment is based upon the fact that organic matter decomposes rapidly in water. When in solution, it is readily available to microorganisms and their enzymes. These convert complex organic compounds into simple substances such as carbon dioxide and ammonia. Because oxygen is the ultimate hydrogen acceptor in these rapid conversions, it may become so depleted that aerobic oxidation is halted. Thus the activated sludge and the trickling filter processes are so designed that atmospheric oxygen is forced into the liquid phase at an accelerated rate. Algal photosynthesis offers a completely different method of supplying oxygen. Here the availability of oxygen is independent of the physical laws normally governing aeration from atmospheric sources. Bodies of water containing green algae may attain supersaturation with dissolved oxygen up to three or four times the normal maximum of water in equilibrium with air. The organic waste material is changed by the cyclic activity of algae and aerobic bacteria into more algal cells. These are then disposed into natural streams or allowed to settle and then be digested anaerobically as bottom sludge. The particular merits of this method lie in the minimal odor, nuisance, and operating cost. It is there- fore considered ideal for small communities situated in mild climates. Particularly in Texas and Southern California has there been reliance on the use of sewage lagoons and oxidation ponds for adequate and economi- cal treatment of sewage. Algal purification of drinking water is also some- times employed; on the other hand, uninvited algae often clog standard sand filtration plants. As previously indicated (section on Culture), Renn233 has recom- mended that oxidation ponds be used not merely for sewage disposal, but also to convert nitrogenous wastes into food protein. This can be accom- plished by encouraging the growth of algae (especially Cblorella, Scene- desmus, and Chlamydomonas) which yield a high quality protein. A relatively new problem in sewage disposal concerns atomic wastes.251 Coopey252 demonstrated radioactivity in all forms of life in the Columbia 48 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE River below the Hanford Atomic Energy plant in Washington. He esti- mated that about 570 tons of wet weight plankton passed the plant daily, with a resulting planktonic radioactivity about 0.6 curies per day. Analyses indicated that 5-30 per cent of the plankton's radioactivity came from isotopes with half-lives near to or longer than P32, while the remainder was of shorter half-life. Some radioactivity persisted beyond 600 days' decay. Since algae can transmit their radioactive toxicity right up through the nutritional pyramid to higher food animals, it is clear that more than conventional disposal methods must be devised for atomic wastes. An indirect application of algal attributes relates to mosquito con- trol. Matheson249 noticed that mosquitoes did not breed in certain pools where Chara fragilis thrived; he and Hinman then postulated that the large amount of oxygen from the algae either interferes with the larval food supply or gives the mosquitoes indigestion. However, Biswas,250 over a decade later, insisted that algal flora provide not only food for the mosquito, but also shelter! b. Animal Intoxications The microalgal scums, odors, and tastes described in the last section produce chiefly esthetic problems. Of far greater import are the many harmful reactions reported to have occurred in both animals and humans.253-273 Freshwater phytoplankton has been implicated since 1878 in the intoxication and death of domesticated animals, as well as water- fowl, shorebirds, and their mammalian or avian predators. In Table VII we present a chronological summation of 38 incidents of animal intoxications by phytoplankton as culled from the literature. In most cases attacks occurred after the animals had drunk from lakes or ponds containing heavy algal growth, usually during hot weather. They have variously been suspected of being due to botulism, anthrax, or worm infestations; or to poisons such as lead, arsenic, copper, cyanide, or alkali; or, as in South Africa, to gallsickness (carried by ticks), lamsiekte, and poisoning by plants such as senecio, gousiekte, and gifblaar. Also con- fused with algal poisoning has been the African geel dikkop of sheep, produced by photosensitizing toxins. The reported symptomatology of algal intoxications has varied, but the most striking clinically have been the involvements of the neuromuscular and respiratory systems. As described by Francis,253 in the Australian outbreak due to Nodularia spumigeria, "the animals developed stupor and unconsciousness, falling and remaining quiet, as if asleep unless touched, PLANKTON 49 when convulsions came on, with the head and neck drawn back by rigid spasm which subsided before death." Similar manifestations have been reported from Alberta,269 Manitoba,262 North Dakota,266 Minnesota,254 Ontario,268 and South Africa.258"260 In several of these, the animals also exhibited extreme photosensitivity,266 with severe blistering of the skin,258"260 and also jaundice.258 Fitch254 tested laboratory animals on the Microcystis and Anabaena which had killed cattle in Minnesota. The characteristic symptoms in guinea pigs included restlessness, incontinence, deep breathing, sneezing, coughing, salivation, lacrimation, weakness in the hind quarters, clonic spasms, and death. Rabbits presented the same picture plus opisthotonus. Pigeons also developed opisthotonus, as well as rapid blinking and swallowing. Mason and Wheeler270 injected Microcystis aeruginosa extract into mice, rats, guinea pigs, and cats. After a latent period of 20-180 minutes, there appeared pallor, hypotension, tachycardia, hypothermia, hyper- glycemia, respiratory difficulty, and death. Steyn258-260 in South Africa reported general paralysis or strychnine-like convulsions following con- sumption of large quantities of Microcystis by cows; smaller amounts re- sulted in constipation, drop in milk yield, generalized weakness, and severe photosensitivity of the skin. If the animals survived longer, many devel- oped jaundice and ascites. Some of Steyn' s laboratory animals manifested symptoms after simply being drenched with the contaminated water. Smit271 injected or fed Microcystis to rabbits. In acute cases, the animals developed anemia, restlessness, dyspnea, progressive paralysis, coma, and death within one-half to four hours. In chronic cases, the outstanding finding was cirrhosis of the liver, with restlessness, atonia, and ascites. McLeod and Bondar262 in Manitoba found that Microcystis administered to rats and mice caused a loss of equilibrium and progressive paralysis; later there were clonic muscular spasms, dyspnea, and cyanosis, then death within twenty hours. The pathological findings in phytoplanktonic intoxications have been most interesting. Autopsies made in 1930 and 1933 in Minnesota showed "no gross pathology"254; cultures, smears, guinea-pig inoculations, and tests for copper and cyanide were also negative. On the other hand, Mason and Wheeler,270 after injecting Microcystis extracts, observed mark- edly congested livers and dilated right hearts, but little generalized venous congestion. Their terminal clinical findings included severe anemia and reduction in total serum proteins. Small amounts of the toxic extract 50 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE •v. § -ft •ft, ^ * si s .»-> 5 ^ <*> ■ft Si <*> Si •v. V. <: Si Jo Si bo U 5$ SI 0 U 0 u u OJ 55 <*> .« "rt "c3 "rt "c3 "3 3 "3 "rt *-r c .5 *, V C c C c c a <-) c •s ft ~ C ^ o »fi ^ ^ ^ -3 Ul 60 *-* Si S5 -ft ■ft. %> * 1— 60 Ul 60 60 Ul 60 60 •-I to &> ^ Ul & st 1 Ul 60 ■a 1 u 3 <*> -ft ■ft. -ft SJ <4> -ft Si 3 1 3 I 3 s CO Ul Ul p? 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Oxygen consumption of various rat tissue slices also remained unchanged. Steyn258"260 reported marked blistering of photosensitized animal skins, as well as extensive involvement of the liver and the central nervous system. Post-mortem examinations showed the lungs full of blood; the spleen enlarged; and the liver engorged and dark red to black. In chronic cases, the liver was either yellow and soft, or dark and brittle. Ashworth and Mason272 were able to produce general- ized cellular damage by administering Microcystis extract to rats. Especially severe injury was noted in the parenchymal cells of the liver. Succes- sive stages included acute parenchymatous, hydropic, and fatty de- generation, followed by central lobular necrosis. If death did not ensue, complete restoration of normal lobular architecture — without fibrosis — took place in thirty days. The toxic alterations were comparable to those produced by chloroform, phosphorus", and epidemic hepatitis. Similar acute hydropic and necrotic changes were also seen in the heart and kidneys, while microscopic hemorrhages and edema sometimes occurred in the lungs. Ashworth and Mason felt that rapid death was due to shock and circulatory collapse, while delayed demise was consequent to hepatic insufficiency, renal failure, and diffuse cellular damage. Stewart and his colleagues268 reported fresh pulmonary hemorrhages and hepatic and cardiac lesions in Ontario cattle dying after ingestion of Anabaena and Microcystis scum. They found the same material lethal to pigeons and rats. Pathological findings from Alberta were "dilatation of the heart" and "mottled liver." As mentioned earlier, many non-algal poisons have been wrongly suspected. In several cases, botulinus antitoxin was administered, to no avail. Solutions of sodium nitrite and sodium thiosulfate given intraven- ously assertedly produced recovery of some poisoned animals. Although the various outbreaks are now known to have been algal in origin, the specific nature of the toxins involved has certainly not been clear. Fitch254 determined that the substances from Microcystis and Anabaena are non- volatile, heat-stable when dry, stable in solution at ordinary temperatures, but unstable when heated to 100°C. They are not electrolytes, alkaloids, or toxalbumins. They are organic componds of low molecular weight, some- what resistant to acid and alkaline solutions. The substances apparently disappear when the algae putrefy. Mason and Wheeler270 were able to inactivate Microcystis extracts partially by heating with acid or alkali. Steyn260 clamied that growing algae are poisonous, that they discharge their poisons into the water when they die, that the decomposition of PLANKTON 53 masses of dead algae decreases the toxicity, and that boiling the con- taminated water does not reduce its toxicity. He thought that Microcystis contains two active principles : a potent f ucoin-type hepato-neurotoxin, and a fluorescent photosensitizing pigment, phycocyan. Wheeler, Lackey, and Schott273 determined that Microcystis is less poisonous when fresh than when frozen or dried. They found the toxin dialyzable, soluble in alcohol, and able to withstand autoclaving in neutral solution. It also survives, unfortunately, the laboratory equivalent of a water purification process — alum coagulation, chlorination, and filtration. It is adsorbable on carbon, but only when this is used in comparatively large amounts. According to McLeod and Bondar,262 in a Manitoba outbreak associated chiefly with Aphanizomenon (and some Microcystis and Anabaena), the toxic material was in the plants and not in the water. They found the toxin stable in the plant, not destroyed by air drying at 37 °C, by freezing, or by ultraviolet irradiation. Shelubsky257 claimed in 1951 that the Microcystis toxin is alkali-labile. In contrast with other investigators, Louw263 has reported isolation of two alkaloids from Microcystis. The first is an in- active one with a probable formula of C10H19NO2. The other, without stated molecular structure, was isolated as a picrate with a melting point of 165 °C. It is supposedly the active hepatotoxin, and it has been effective in experimental rats. Somewhat similar findings with marine algae from the South Pacific have recently been described by Habekost,274 who iso- lated substances toxic to mice. c. Human Intoxications Algal poisoning has not been limited to animals. During the past 25 years, phytoplankton has also been incriminated in human reactions. These have presented as three general clinical pictures: dysenterial disorders, systemic allergic reactions, and local allergic eruptions. Table VIII sum- marizes the reported human cases.275*281 (Poisonings from ingestion of plankton-nourished fish will be discussed separately later.) The epidemic intestinal disorders of the early 1930's involved many thousands of people, in areas as widely separated as the National Parks in the Northwestern United States275 and Charleston, West Virginia.276 Symptomatology consisted of nausea, vomiting, diarrhea, and sometimes abdominal pain. It was self -limiting and usually resolved in a few days. These outbreaks were of no known specific etiology, but, significantly enough, were always concomitant with the presence of extensive algal blooms in the local water supply. Doubtless many other similar outbreaks have been neither studied nor reported. 54 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE §3 6 6 B 1 1 C<1 Si «5 o o o o 5 | 2 so "a> •5 4* til _Q 3 3 c <4i w ^ "os "os "c3 >-> -o d *3 co w c o _o 4-» ; a, > TD c a. "c3 a. iu C U co >> Q 60 c u l-H — * rrt -T? 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U9 « o oj — 4h o 2 u ^ o c o CO rO ^r (N CN M-s NO , >% >^ >> >* oj oj oj oj oj 2 22 22 do* $$ 0^ <^S °& Ul Ul Ul Ul Ul Oj Oj oj Oj oj e e e e e g o o o o o ;=; CO CO CO CO CO nJ OJ OJ 2 OJ OJ 2 OJ OJ s G G G 0 0 CO CO CN 00 ON O H M fN (N <-Ci rr, rO rO OJ OJ OJ >% >^ >N OJ OJ OJ 222 % G G G -« G G G 5 000^ CO CO CO ^ ir\ no r- 00 fO rd rO rO T3 TO OJ u* CO TO ON m PLANKTON 59 < n xr ™ (N (N ^2 ^S u 6 — M o xr SO OS 6 o xr CO , to OS 3 u "a3 co c co 1 TJ as CO t-l co ■i -o JC u pq s co c co > w CQ W aj S.2 v ,V CO cS 3 ffl S *J ^> cu s 6 3 co a C U N ■ CO 1) 3 ■aits U a? 3 O 'C c^ ^J u cd 2 ?2z u • ^^ u rt (j u f*< >-C <-C PL, O, co C rt r r co co co 6 C 60 c c c co co" Ph 3 O CQ c >-} 1-1 3 CO O ,. CO C ^ co O >H U hJ U C u « > > U3 >n aS u u Pm u as Ph CO U3 - « CO I— H CO t-H u a Ph C _, <1) aS u ^ o aS CQ c co J3 co a, a co M as 4-1 > CO o S^ t4 C7\ o o eft fft ^r Cs Cs CTs I ■-1 Tf xf ^f ^ I1 M1 tf O Cs C\ On so XT r^ ^r ift \r\ tr\ so *0 xr •* xr ^ ^r C\ Cs Cs Cs Cs 00 00 00 CS CS CS CS O fft Xf xf v\ \r\ its Cs CS Cs CS "T3 aJ M c o U aS co G St N o v. O XT G C O O CO CO ^ o 5 xf xr -a ai T3 as tJ •*■> CO <_< CO T! T-" tn 1 ffi as DC h-j h-j u • •4 Ul c aS u. |X| G O o u c c as -o U G •V JG O >Hu CO co o Cr5 U3 O u aS - s 4J O J5 o rft Xf \r\ SO XT xr XT XT N 00 C\ O h xf XT xf irs i/-s (N eft xr ^s V "O . "^) *-> aS — i aS G V .5 4> aS *-• J3 *-* e J2 C 2 5 42 ^ 3 8 DC < X H tft SO h» 00 trs irs crs vrs 60 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE mouse on injection, while a few milligrams orally can be fatal to a human being, especially when taken on an empty stomach. It has been estimated that 4,000 algal organisms are required to produce human toxicity. The shellfish store the poison chiefly in their "livers" or digestive organs; thus the "dark meat" of the infested bivalves is much more toxic than the "white" portions. The fish themselves are unaffected by the toxin, and they will in fact excrete it in several weeks if Gonyaulax disappears from the marine diet.297-298.302-303 Post-mortem examinations of animals and human beings who have died of acute paralytic shellfish poisoning have, surprisingly enough, revealed no structural changes. However, in chronically poisoned experimental animals, distinct alterations are noted in the medullary ganglion cells and in the large cells of the ventral horn of the spinal cord.302 The Golgi apparatus of small and medium-sized spinal ganglionic cells is also dam- aged. Neural mitochondria remain normal, but those in the convoluted tubules of the kidney show definite damage. In concert with the latter finding are the clinical nephropathies reported by the French in human shellfish poisoning.304'305 It is probable that other algae besides Gonyaulax contain toxins. Some have been associated with Ceratium and Prorocentrum, but no direct ex- traction or isolation has yet been accomplished. It is also quite logical to expect that fish other than mussels and clams could have improper plank- tonic relations and thus transmit their toxins to predators. This is indeed so, and — except for the puffer fish and possibly barracuda — it is likely that poisonousness is directly dependent upon periodic ingestion of noxious plankton, rather than being due to inherent toxicity. Planktogenic fish toxicity can occur during any time of the year, while puffer and barracuda may be most lethal during the reproductive season. Epidemics have been unrelated to spoilage or bacterial infection. Poisonous fish are found through the world. They are most numerous in the warm seas (especially in the Caribbean and Central and South Pacific), but even certain arctic sharks have been reported to be toxic. Dur- ing World War II, fish poisoning represented quite a problem, since epidemics were encountered wherever American forces were stationed in the Pacific theater. The Japanese, too, estimated that over 400 of their military men died in Micronesia from eating toxic fish. Halstead and Lively300 have made a partial listing of fish most often found to be poison- ous: surgeonfish, triggerfish, pompano, porcupine fish, wrasse, snapper, filefish, surmullet or goatfish, moray eel, parrot fish, sea bass or grouper, PLANKTON 61 barracuda, and puffer. They have divided fish poisoning into four clinical groups, the last three of which are probably planktogenic: 1. Tetraodon (puffer) poisoning 3. Ciguatera 2. Gymnotborax (Moray eel) poisoning 4. Scombroid poisoning Tetraodontoxin is the only ichthyosarcotoxin about which anything specific is known. Japanese studies indicate it is a white hygroscopic powder, soluble in water but not in the ordinary organic solvents. Although it is thought to have a formula of C16H31N016, little else is known except that it is not an alkaloid or a protein or a protamine. It is a most potent poison, with symptoms occurring within thirty minutes of ingestion. Initially there are numbness and tingling of the face and extremities, then nausea, vomiting, headache, dizziness and overwhelming weakness. Next come speech impairment, dyspnea, and generalized muscular paralysis. Death from respiratory paralysis may occur in 1-24 hours. The mortality rate is about 60 per cent. In Gymnotborax poisoning, similar neurotoxic manifestations are seen at first, but then the patient develops motor incoordination and violent convulsions, laryngeal spasm, coma, and respiratory paralysis. Here the mortality rate is only about 10 per cent, and, in the non-fatal cases, major symtomatology disappears in about ten days. With Ciguatera (originally considered only Caribbean), the attacks are milder than with Gymnotborax and usually come on more slowly. The chief symptoms include sensory disturbances, myalgias, arthralgias, and severe weakness. Only 2-3 per cent of the victims die, but recovery may take many months. Scombroid poisoning results from eating various tropical tuna-type fish. The mani- festations are mostly histamine-like: headache, flushing of the face, con- junctivitis, giant hives, erythema, and gastric upset. Recovery usually takes place in 8-12 hours. Haffkrankbeit — erroneously designated "Haff's Disease" by some enthusiastic eponymist — was first described in 1924295 among fishermen in Koenigsberg HafT ("harbor") in East Prussia. It follows contact or ingestion of fish (primarily eels) and fish livers. It is characterized by a sudden onset of weakness, severe myalgia, myoglobinuria, and some digestive upset. Pathological findings include fatty degeneration of the liver and kidneys, and damage to the anterior horn cells of the spinal cord. Over 1,000 people have been reported afflicted, some fatally. Similar episodes have been recorded at Lake Ysmen in Sweden. Originally Haff- 62 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE krankheit was considered to be caused by arsines in sewage. However, the correlation of outbreaks with climatic changes, and the predominantly neurotoxic nature of the manifestations, suggests that this disease, like the several other categories of fish poisoning, is dependent upon periodic planktbnic intoxication. No method of testing suspected fish is known, other than feeding it to animals (kittens have been a favorite). Neither does any specific antidote exist. Treatment has consisted primarily of rapidly emptying the gastro- intestinal tract by mechanical means, apomorphine, and laxatives. Adminis- tration of charcoal as an adsorbent may be helpful. Also important are supportive measures such as stimulants, parenteral fluids, oxygen, and artificial respiration. Intubation and tracheotomy may be necessary with respiratory distress. Anticonvulsants and opiates may have their indica- tions, while some assign a special merit to intravenous calcium gluconate. To us it would seem logical also to employ the adrenal steroids, either to combat anaphylactoid reactions or to diminish end-organ response to toxins. Anti-histamine agents, too, might play a part in treating the Scombroid type of poisoning. Doubtless the wisest management of fish poisoning consists of avoiding those fish with a known toxic potential; and, in the case of shellfish, avoiding them when the Gonyaulax or other suspected algae are flourishing. V. DISCUSSION After combing thousands of articles on algae and plankton, we are in firm agreement with Conway Zirkle,306 who laments the fact that our age of specialization has produced a "splintered learning." He points out that many important developments are hope- lessly buried because of the compartmentalization of knowledge, the spe- cialized jargons of the various sciences, and the overwhelming volume of new publications. Because of a limited lateral diffusion of information, many workers remain unaware of pertinent findings even in kindred fields. This is especially true in the realm of medicine. With the tremendous mass of studies that physicians must continually digest, their meager knowl- edge of algae and plankton is readily understandable. The rare medical DISCUSSION 63 articles on these subjects have usually dealt only with some special point of interest — or a courtesy reference has appeared in a botanical, nutri- tional, or public health journal. The present assemblage of data brings into focus much diversified material, a good portion of which deserves closer integration into the field of medicine. Interest in algae and plankton has varied over the years. From ancient times, the predominant emphasis was on the seaweeds ; at the turn of this century, it shifted to zooplankton ; and, since World War II, it has centered chiefly about phytoplankton. The traditional medicinal use of seaweed has declined sharply, as has its role as a human nutrient. Its value today resides primarily in the contained minerals and vitamins, and its greatest utility is as fertilizer and animal provender. There is also much merit in the alginates, and a good potential exists for other seaweed derivatives such as the antibiotics and anticoagulants. Consideration of plankton has for the most part been focused on its use as food.307-309 Zooplankton is nutritionally far richer than seaweed, but it does not possess the signifi- cance its more enthusiastic proponents confer upon it. Despite the fact that the Thais21 have reportedly collected 5,000 tons in one year, it should not be forgotten that there are tremendous variations in quantity and quality of yield in different oceans and at different seasons. While zooplankton may be considered an excellent emergency ration and food supplement, its extensive use as a food staple awaits the development of far more efficient collection techniques. Since it is still much too early to envision extensive in vitro photo- synthesis197 without the intermediary of plants, it is the phytoplanktonic organisms which offer the most exciting prospect for nutritional gain. They not only supply controllable proportions of protein, fat, vitamins, minerals, and calories — they can also be cultured selectively on a large- scale basis. The marvelous adaptability of algae like Chlorella gives man another round in the crucial Malthusian battle of population versus food supply. In the past three centuries, the world population has risen from a half billion to two and one-half billion.25 If fission and fusion do not permanently put an end to both mankind and its problems, the population will be four billion in 1980, and eight billion in 2050. Selective fresh- water mass culture of organisms (we suggest the term "aquaculture" as more descriptive than "hydroponics" and less restrictive than "mari- culture"29) like Chlorella in specially built plants hold great promise for large quantities of valuable foodstuffs. It is more practical than "manuring" the sea with salts and minerals — "three-dimensional farming" — to increase 64 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE the growth of phytoplankton, whether one hopes to harvest the phytoplank- ton as such, or whether it is planned to support a greater fish population. The latter is, in fact, a wasteful process, since it has been calculated that a pound of codfish ultimately represents 100,000 pounds of marine phyto- plankton!7 Before phytoplanktonic feeding becomes commonplace proced- ure, there will have to be long-term human experiments, with careful observation of possible toxicities and biochemical changes, especially with respect to electrolytes, hemic elements, and proteins. It is also a certainty that a re-education of food tastes will have to take place before the accept- ability of planktonic foods reaches a high level.310 Some other applications of phytoplankton reach almost into the realm of science fiction. Bassham,311 at the University of California, has been working for the U.S. Navy on adapting microscopic algal cultures to con- trol the oxygen and carbon dioxide in atomic submarines which would be submerged for long periods. It has been calculated that one kg. fresh weight of Chlorella (equal to 100 liters of growing algal suspension) can easily supply the 25 liters per hour oxygen required by a 70 kg. man, as well as utilize the carbon dioxide exhaled by him. In a similar fashion, the Department of Space Medicine of the Air Force22 is considering using algae in space ships, with the intriguing extra notion of complete recycling of biological elements : the algae are also to be used as food by the space- men, while their excreta will serve as nutriment for the algae ! From the strictly medical standpoint, the number of disease syndromes attributable to algae comes rather as a surprise and raises some interesting questions. The most obvious afflictions are the allergic dermatitides re- sulting from bathing in water contaminated with algae. Also under- standable are the respiratory irritations from water-borne or inhaled algae ; in fact, many cases of allergic rhinitis (and its corollary, chronic sinusitis) in coastal areas might be traceable to algae instead of being blamed on "damp climate" or that handy favorite of allergists, "dust." Then there are the nephropathies, and the striking epidemic gastroenteritides, so blandly ascribed to the "24-hour or 48-hour virus." By all odds, the most dramatic algal intoxications are the acute neuro- toxic and myotoxic ones typified by paralytic shellfish poisoning and some other ichthyosarcotoxicoses. Although pitifully meager long-range studies have been made, it seems incredible that such potent toxins should not produce related chronic sequelae in victims who have survived acute attacks, or who have ingested smaller amounts of toxin over prolonged periods. One cannot help but wonder whether chronic algal intoxication DISCUSSION 65 may not play a part in causing some of our idiopathic neuropathies such as multiple sclerosis, the muscular dystrophies, and amyotrophic lateral sclerosis. Interestingly enough, very recently a U.S. Public Health Service team has reported312 an extensive survey of amyotrophic lateral sclerosis among the Chamorros of Guam and other islands of the Mariana group. They made the study because the incidence of the disease there is 420 per 100,000 population, a rate 100 times higher than in the rest of the world; at any one time, one per cent of the adults are affected, and eight to ten per cent of adult deaths are produced by it. No cause was discovered, except perhaps some tenuous genetic relationship to the Chamorro tribe. The investigators have apparently given little consideration to the water contacts or fish-eating habits of these people. It may be more than co- incidence that epidemics of fish poisoning — likely due to toxic phyto- plankton — have been particularly plentiful in the Marianas. That algae have not been implicated in the nerve afflictions may be due to the difficulty in demonstrating them, or simply to their not even having been considered. It is interesting, if hazardous, to speculate also on a possible algal role in another neurologic disease — acute poliomyelitis. The viral etiology is certainly proven beyond any doubt, yet the epidemiology is anything but clear. Is it possible that a symbiotic virus-alga state exists, favoring viral transport or even viral multiplaction ? The thought is suggested by the fact that the highest incidence of poliomyelitis is during the warm summer months, when algal blooms are most prolific; also, that infection occurs frequently following swimming. 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M., and Savory, M.: Exenteration of the orbit and use of alginate mould for applying skin grafts, Brit. J. Ophth. 35:39, 1954. 132. Gosset, J., and Martin, J.: Un nouvel hemostatique chirurgical: 1'alginate de calcium, Mem. Acad, de chir. 73:273 (March 16) 1949. 133. Oliver, L. C, and Blaine, G.: Haemostasis with absorbable alginates in neurosurgical practice, Brit. J. Surg. 37:307 (Jan.) 1950. 134. Mullard, K. S.: The control of haemorrhage in extrapleural pneumothorax, Thorax 3:233, 1948. 135. Trimble, J. R., and McShane, W. P.: Alginate as a potential decontaminating and therapeutic material against mustard burns, Chemical Corps Medical 72 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE Laboratories, Array Chemical Center, Md. CMLRE-ML-52 Medical Labora- tories Research Report no. 238, Jan., 1954, p. 1. 136. Cain, R. M., and Steele, H.: The use of calcium alginate soluble wool for the examination of cleansed eating utensils, Canad. J. Pub. Health 44:464, 1953. 137. 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Soc. chim. biol. 33:312, 1951. 143. Roche, J., and Yagi, Y.: Sur la fixation de l'iode radioactif par les algues et sur les constituants iodes des laminaires, Compt. rend. Soc. de biol. 1 46: 642 (May 10) 1952. 144. Foran, R. R.: The mineral and vegetable resources of the sea, Am. J. Pharm. 97:249, 1925. 145. Sutton, H.: Watching the herring marinate, Saturday Review Aug. 21, 1954, p. 32. 146. Chilson, F.: Modern cosmetics, 1938, Drug and Cosmetics Industry, New York. 564 p. 147. Cabrero Gomez, F.: Estudio de las algas marinas espariolas desde el punto de vista de su approvechamiento industrial, 1951, Consejo superior de in- vestigaciones cientificas, Madrid. 110 p. 148. Herdman, W. A.: Founders of oceanography and their work, 1923, E. Arnold & Co., London. 340 p. 149. Crowder,.W.: Living jewels of the sea, Natl. Geographic Magazine .52:290, 1927. 150. Moore, H. B.: Strange babies of the sea; scientists are slowly solving the mysteries of plankton, the ocean's vast underwater pasturage, Natl. Geographic Magazine 102:41, 1952. 151. Conseil Permanent International pour l'Exploration de la Mer: Bulletin planktonique pour les annees 1908-1911, H0st, Copenhague, 1912-1915. 152. Rylov, W. M.: Das Zooplankton der Binnengewasser, 1935, E. Schweizerbart, Stuttgart. 272 p. (Die Binnengewasser, hrsg. v. A. Thienemann, vol. 15). 153. Conseil Permanent International pour l'Exploration de la Mer: Zooplankton sheets. Chaetognatha I, [by F. S. Russell], 1938, [Copenhague] 4 p. BIBLIOGRAPHY 73 154. Jespersen, P., and Russell, F. S., ed.: Fiches d'identification du zooplankton, no. 1, 1949 , H0st, Copenhague. 155. Wautier, J.: Introduction a l'etude des biocoenoses, 1952, P. Ferreol, Lyon. 124 p. 156. Jenkins, T.: Sea-fishing, 1909, Asiatic Society of Bengal, Calcutta. 9 p. 157. Harvey, H. 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A.: Plankticheskaia produktivnost severnykh morey SSSR. Plankton productivity of the northern seas of the USSR, 1940, Moskovskoe obshchestvo ispytateley prirody, Moscow. 84 p. (In Russian with English summary). 166. Borutskii, E. V.: Sezonnoe vertikalnoe raspredelenie planktonykh copepoda v tolshche vodnoi massy Belogo ozera v Kosine p otdelnym stadiiam razvitiia. [Seasonal vertical distribution of plankton copepods in waters of the White Lake, according to various states of development] (In Russian), Zoologiche- skii Zhurnal (Moscow) 29:120, 1950. 167. Wiborg, K. F.: The production of zooplankton in the Oslo Fjord in 1933- 1934 with special reference to copepods, Hvalradets skrifter, Scientific Re- sults of Marine Biological Research, Norske Videnskaps-Akademi, Oslo, no. 21, 1940. 168. Kerr, J. G.: Plankton as food? [Letter] to the editor of the Times, Times (London) May 6, 1941, p. 5. 169. Gardiner, A. C: Plankton as a source of food, Nature (London) .748:115 (July 26) 1941. 170. Knowlton, G. C, and Irving, L.: Evaluation of the effect of eating plankton upon the water requirement of men in life rafts, Mem. Rep. Aero M. Lab. (TSEAL 3-696-76) p. 1-4 (Aug. 16) 1945. 171. Heyerdahl, T.: Kon-Tiki, across the Pacific by raft, 1953, Garden City Books, Garden City, N. Y. 218 p. 172. Bombard, A.: The voyage of the Heretique, 1953, Simon and Schuster, New York. 214 p. 173. Lone New Yorker drifts to Samoa in 115 days across Pacific on raft, New York Times, Oct. 15, 1954. 74 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE 174. Carson, R. L.: The sea around us, 1951, Oxford University Press, New York. 230 p. 175. Priestley, J.: Experiments and observations relating to various branches of natural philosophy; with a continuation of the observations on air, 1779-1781, J. Johnson, London, v. 1-2. 176. Ingen-Housz, J.: Experiments upon vegetables, discovering their great power of purifying the common air in the sun-shine, and of injuring it in the shade at night . . . 1779, Elmsly and Payne, London. 177. Cohn, F.: Untersuchungen iiber die Entwicklungsgeschichte der mikros- kopischen Algen und Pilze, Verhandlungen (Nova acta), Kais. Leopoldinisch- Carolinische Akademie der Naturforscher (Breslau & Bonn) 24(1) :103, 1854. 178. Faminitzin, A.: Beitrag zur Symbiose von Algen und Thieren, Memoires Academie imperiale des sciences, St.-Petersbourg, ser. 7, v. 36, no. 16, 1888. 179. Engelmann, T. W.: Farbe und Assimilation, Bot. Zeitung 41:1; 17, 1883. 180. Beijerinck, M. W.: Culturversuche mit Zoochlorellen, Lichenengonidien und anderen niederen Algen, Bot. Zeitung 45:725 ; 741 ; 757, 1890. 181. Molisch, H.: Die Ernahrung der Algen, Sitzungsberichte Kais. Akad. Wis- sensch. Wien. Math.-Naturw. Classe. Abt. 1, 1 04: 783, 1895 (Abhandl. I) ; 20^:633, 1896 (Abhandl. II). 182. Willstatter, R., and Stoll, A.: Untersuchungen iiber das Chlorophyll, 1913, Springer, Berlin. 424 p. 183. Warburg, O.: Ueber die Geschwindigkeit der photochemischen Kohlen- saurezersetzung in lebenden Zellen, Biochem. Ztschr. 700:230, 1919. 184. Roach, B. M. B.: On the relation of certain soil algae to some soluble carbon compounds, Ann. Botany (London) 40:149, 1926. 185. Barker, H. A.: The oxidative metabolism of the colorless alga, Prototheca zopfii, J. Cell & Comp. Physiol. 8:231, 1936. 186. Pearsall, W. H., and Loose, L.: The growth of Chlorella vulgaris in pure culture. Proc. Roy. Soc. London, s.B. 121:451, 1937. 187. Pringsheim, E. G.: Pure cultures of algae, their preparation and maintenance, 1946, University Press, Cambridge [Eng.] 119 p. 188. Myers, J.: Physiology of the algae, Ann. Rev. Microbiol. .5:157, 1951. 189. Schussnig, B.: Handbuch der Protophytenkunde, 1953, G. Fischer, Jena. v. 1. 190. Growth of protozoa (Conference on Growth of Protozoa, Sect, of Biol., New York Acad. Sc, Oct. 24 and 25, 1953). Ann. New York Acad. Sc. ;56(5):815, (Oct. 14) 1953. 191. American Geographical Society (New York): Study in human starvation. Atlas of distribution of diseases, no. 8, 1953; no. 9, 1953- 192. May, J. M.: The mapping of human starvation, diets and diseases, Geographi- cal Review 43:403, 1953. 193. Kuckuck, P.: Beitrage zur Kenntnis der Meeresalgen, Wissenschaftliche Meeresuntersuchungen, Biologische Anstalt (Helgoland, Kiel) N. F. 2:325, 1897. 194. Naumann, E. C. L.. Grundlinien der experimentellen Planktonforschung, 1929, E. Schweizerbart, Stuttgart. (Die Binnengewasser, hrsg. v. A. Thiene- mann, vol. 6). 195. Mangenot, G.: Recherches sur les constituants morphologiques du cytoplasma des algues, Arch, de morphologie, no. 9, 1922. 330 p. BIBLIOGRAPHY 75 196. Huber-Pestallozzi, G.: Das Phytoplankton des Siisswassers, Systematik und Biologie, 1938 (Pt. 1); 1941-1942 Blastocystis 55 Blood formation 31 Blood plasma substitute 21 Blue-green algae 36, 37, 46, 50, 51 Boron 40 Bread 6, 7, 12 Bromine 9, 12, 20 Burns 14, 19 Calanus 23, 24 Calcareous algae 16 Calcium 8, 9, 11, 18, 19, 40 Calothrix 36 Cancer 13, 14 Carbohydrate 2, 7, 8, 25, 26, 28, 30, 33, 34, 36, 37, 46 Carbon 28, 29, 30, 35, 37, 53 Carbon dioxide 2, 32, 33, 40, 41, 42, 43, 47, 64 Carotene 3, 29, 31, 32, 38, 40 Carotenoids 29, 31, 32, 33 Carragheen 6, 16, 17 Carragheenin 8, 17, 20 Catalase 32 Cathartics 14, also see Laxative Cation exchange resins 19 Cell wall components 2, 3, 7, 8, 36 Cellular plasma 17 Cellular respiration 28. Also see Photosynthesis Cellulose 2, 3, 8, 29 Centroceros 13 Ceratium 45, 60 Char a 32, 48 Chelating agents 41 Chemistry 23, 25, 28, 30,-38 Chewing material 21, 22 Chilomonas 39 Chironomus 31 Chitin 8, 25, 26 Chlamydomonas 3, 30, 33, 35, 39, 45, 47 Chlorella 3, 27, 28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 47, 55, 56, 63, 64, 65 "Chlorellin" 55, 56 Chlorination 46 Chlorine 9, 25, 46 Chlorophyceae 14, 31, 32 Chlorophyll 3, 29, 31, 32 Chlorophyta 2, 3, 27, 33 Chloroplast 29 Choline 40 Chondria 16 Chondrillasterol 37 Chondrus 3, 6, 7, 16, 20 Chordaria 15, 20 Chroococcus 3, 44 Chrysophyta 3, 27 Ciguatera 57, 58, 61 81 82 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE Cirrhosis 49, 65 Cladophora 3 Clams 56, 58, 60 Classes of algae 2, 3 Classification 2, 3 Coagulant 39 Cobalt 39, 40. Also see Cyano- cobalamin Co-carboxylase 39 Codium 9 Coelosphaerium 50 Color classification of algae 2 Colorless algae 28, 32 Complexing agent, see Chelat- ing agent Confervae 14 Conjugata 45 Constipation 13, 19, 49 Copepods 4, 22, 23, 24, 25, 26 Copper 12, 13, 40, 47, 65 Corralina 3, 14 Cortisone synthesis 37 Cosmetics 17, 22 Cough 16 Crannogh 6 Crocetin 33 Crocin 33 Crustaceae 4, 23, 26 Cryptomonas 41 Culture 17, 28, 29, 32, 33, 34, 36, 37, 38, 41, 42, 43, 44, 47, 63, 64 Culture apparatus 41, 42, 43 Custard powder 7 Cyanocobalamin 10, 39, 40. Also see Cobalt Cyanophyta 3, 27 Cycadaces 36 Cylindrospermium 45 Dasycladus 33 Deamination 36 Decomposition of algae in water 46 Definitions 2, 4, 22 Dehydrated algae 7 Dehydrogenase 38 Delesseria 20 Demulcent 15, 16 Dermatologic disorders, see Skin disorders Desmarestia 11 Desmid 45 Desoxyribosenucleic acid 39 Diarrhea 14, 15, 16, 53. 54 Diatomales 46 Diatoms 3, 4, 22, 25, 27, 32, 37 Dictyopteris 16 Digenea 15 Digestibility 7, 25, 29 Dinoflagellate 4, 22, 25, 27, 56 Dinophyceae 56 Dietary shortage 1 Dietary supplement 1, 11, 26, 44, 63, 64 Dillisi 6 Dity Ilium 41 Divisions of algae 2, 3 Dorset weed 6 Drinking water contamination 46, 47 Dropsy 13 Drowning, diagnosis of 56 Drying oil 37 "Duckweed" 5 Dulcitol 8 Durvillea 14, 16 Dulse 6, 10, 16, 21, 22 Dysentery 15, 53, 54 Earache 14 EDTA 41 Eels 58, 59, 60, 61 Egregia 30 Enteromorpha 15 Enzyme systems 33, 34, 37, 47 Ergosterol 10, 37, 39 Euglena 3, 35, 39 Euglenineae 31, 39 Euglenophyta 2, 3, 27 Euphasiids 23. Also see Krill Families of algae 2, 3 Fats 7, 9, 25, 26, 33, 37, 38, 63 Fatty acids 9, 37, 38, 56 Feeding experiments 10, 11, 12, 18, 25, 39, 44, 49, 64, 65 Femaleness 33. Also see Sex- uality Fertilizer 11, 36, 42, 63 Fish 4. 15, 23, 26, 38, 45, 46, 53, 56, 57, 58, 59, 60, 61, 62, 64 Fish poisoning 53, 56, 57, 58, 59, 60, 61, 62, 65 Flavoprotein 39 "Fleur d' eau" 45 Floridean starch 8 Florid ee 16 Floridoside 8 Fluorescence, see Phosphores- cence "Flos aquae" 45 Folic acid 10 Folinic acid 10 Food, algae and plankton as 1, 4, 5, 7, 13, 24, 25, 27, 29, 37, 38, 41, 44, 45, 47, 48, 63, 64 Freshwater algae 5, 27, 30, 42 Freshwater phytoplankton 3, 48 Freshwater zooplankton 23, 24 Fried seaweed 7 Fucoidin 8, 20 Fucus 3, 9, 12, 15, 16, 17, 22 Fugu poison 58 Fungi 55 "Funori" 22 Furcellaria 9 Galactosides 8 Gametes 33 Gastrointestinal disorders 14, 15, 53, 64 Gelidiaceae 6, 17 Gelidium 3, 6, 8, 15, 16, 20 Genera of algae 2, 3, 36 "Generation time" 33 Gentiobiose 33 Germanium 40 Gigartina 3, 17, 30 Gloeotrichia 41, 50 Gloiopeltis 22 Glucopyranoside 8 Glucose 7, 8 Glutamate 39 Glutamic acid 8, 35, 36 Glutamine 35 Glutaric acid 34 Glycine 8, 35 Goiter 13, 14, 15 Gonyaulax 3, 56, 60, 62 Gout 14 Gracilaria 6, 13, 15 Gristle moss 7 Growth of algae and plankton 4, 33, 37, 38, 39 Gymnodinium 3, 41, 45 Gymnothorax poisoning 61 Haffkrankbeit 61, 62 Hansen's disease 38, 44 Harvesting 41, 43 INDEX 83 Heartburn 14 "Hekistoplankton" 4 "Helminol" 15 Heme 31 Hemicellulose 8 Hemin 31 Hepatitis 14, 52, 65 Hepatotoxins 49, 52, 53 Heretique 26 Hexoses 32 Histidine 35 Homocystinea AA Human toxicity, see Toxicity Hydrocarbons 9, 29 Hydrocolloids 19 Hydrogen 30, 47 Hydrogen peroxide 32 Hydrophylic colloids 17 "Hydroponics" 63 Hydroxylamine 46 Hypnea 13, 16 Hypovitaminosis 1 Ice cream stabilizer 20 Ichthyosarcotoxin 61, 64. Also see Fish poisoning Illumination of cultures 42, 43 Inflammation treatment of 14 Ingestion of algae and plank- ton 4, 5, 6, 7, 25, 44, 48, 49, 52 Injection of algae and plank- ton 18, 49, 50, 52, 54, 55 "Inner complex salts" 41 Intestinal disorders 14, 15, 17, 53 Intracellular storage matter 7, 8, 33, 36, 37 Iodine 9, 11, 12, 15, 16, 20, 21, 38, 40 hide a 20 Irish moss 6, 12, 15, 16 Iron 40 Isoleucine 35 Isotopes 35, 48 Jaundice 49 Jelly moss 1 Kan ten 6 Karengo 21 Kelp 2, 6, 8, 9, 10, 14, 15, 16, 17, 22 Kombu 6 Kon-Tiki 26 Krebs tricarboxylic acid cycle 39 Krill 4, 22, 23 Kwashiorkor 1 Labor 16 Laminaria 3, 6, 7, 8, 9, 10, 12, 13, 15, 16, 17, 18, 20 Laminarin 8, 20, 21 Land plants 29, 30, 34, 40 Laver 6, 10. 13, 14 Laxative 15, 19. Also see Cathartic Lead 40 Leprosy, see Hansen's disease Leucine 8, 35 "Limnetic" growth 5 Linoleic acid 37 Lipids 2, 9, 28, 29, 30, 34, 36, 37, 38 Lipoic acid 40 Lipoid substances 9 "Littoral" growth 5 Lysine 8, 35 Macrocystis 3, 17, 30 Macroscopic algae 3, 5, 7, 8, 9, 10, 12, 17, 36. Also see Seaweed Magnesium 8, 9, 29, 40 Maleness 33. Also see Sex- uality Manganese 40 Mannitan 7 Mannitol 7, 8 Mannose 7 "Manuring" of the sea 63 Manuronic acid 8, 17 "Mariculture" 63 Marine algae 2, 3, 15, 16, 17, 20, 30, 54 Marine plankton 2, 3, 4, 23, 24, 25 Marine sauce 6 Marine zooplankton 23, 24 Medicinal usage 13, 14, 15, 16, 17, 38, 44, 45 Menstrual disturbances 13 Metabolism 2, 28, 29, 32, 37 Methionine 34, 35- Microcystis 3, 18, 49, 50, 51, 52, 53, 54, 55 Micronutrients, see Trace ele- ments Microscopic algae 3, 4, 7, 11, 27, 28, 29, 34, 36, 37, 38, 39, 40, 45, 48, 55. Also see Phytoplankton Minerals 2, 17, 27, 28, 29, 63 Molluscs 4 Molybdenum 40 Mosquito control 48 Mucin 35 Muscus 14 Mussels 56, 57, 58, 59. 60 Mutants 31, 32. 39 Myotoxic manifestations 61, 64 Mysids 23 Myxophyceae 31, 35, 45, 46, 55 Nannoplankton 4 Navicula 30 Nephropathies 60, 64 Nereocystis 3, 21 "Neritic" plankton 27 Neurologic manifestations 12, 14, 48, 49, 56, 60, 61, 64, 65 Neurotoxins 49, 52, 53, 61, 62 Niacin 10, 40 Nickel 40 Nit el la 3, 32 Nitrogen 2, 8, 18, 25, 28, 30, 33, 34, 35, 36, 37, 40, 46, 47 Nitrogen fixation 35, 36, 40 Nitzschia 3, 37, 38, 39, 41 Nodularia 48, 50 Nostoc 36 Nucleic acid 9, 35 Nutrients 12, 23. 27, 33, 34, 40, 41, 42, 43, 63, 64 Nutritional uses 2, 5, 10, 11, 29, 39, 44, 63, 64 Oarweed 15 Obesity, treatment of 16 Odour 45, 46, 47, 48 Oedogonium 3, 33 Oikomonas 30 Oils 46 Oleic acid 37 Oligosaccharide 36 Oocystis AA Orders of algae 2, 3 Oscillatoraceae 54, 55 Osctllatoria 35, 45, 54, 55 "Oxidation ponds" 43, A6, 47 Oxidative processes 28 Oxygen 29, 32, 36, A6, 47. 48, 52, 64 Oxyuriasis 15 84 THE ROLE OF ALGAE AND PLANKTON IN MEDICINE Palmitic acid 9, 37, 38 Pantothenic acid 40 Para-aminobenzoic acid 39 Passerine birds 13 Pearl nwss 7 Pectins 2, 3, 8 Pentoses 7 Pentosans 36 Peptide 8, 9 Phaeophyceae 7, 8 Phaeophyta 3, 10, 33 Phenylalanine 8, 35 Phosphorescence 26, 27, 32, 56 Phosphorus 40, 46, 48, 52 Photosensitivity 48, 49, 52, 53 Photosynthesis 2, 28, 29, 31, 32, 33, 40, 46, 47, 63 "Phyceine" 56 Phycobilin 5, 29, 31, 32 Phycocolloids 17, 20 Phycocyanin 3, 31, 53, 54, 55 Phycoerythrin 3, 31 Phycology 1 Phyllogigon 15 Physiology of algae 28 Phytoplankton 3, 4, 23, 24, 27, 28, 29, 32, 37, 38, 39, 41, 44, 45, 48, 50, 51, 54, 63, 64. Also see Micro- scopic algae Phytoplankton poisoning, see Toxicity Pigments 2, 9, 28, 29, 31, 33, 53, 55 Pilot plant studies 34, 35, 40, 41, 42, 43 Plankton 1, 3, 4, 5, 22, 24, 25, 26, 27, 39, 45, 46, 48, 56, 62, 63. Also see Phyto- plankton, Zooplankton, Mi- croscopic algae foods 26, 44, 45, 63, 64 poisoning, see Toxicity Plant plankton, . see Phyto- plankton Plectonema 16 Pleurococcus 45 Poisoning, see Toxicity Polysaccharides 8, 20, 36 Poliomyelitis 65 Polytomella 3, 35. 39 "Pondweed" 5 Porphyra 3, 6, 10, 13, 21 Porphyria 31 Potassium 9, 12, 17, 19, 40 Poultices 13, 16 Prawns 22, 23 Proline 8 Prorocentrum 3, 60 Protein 1, 2, 7, 8, 25, 26, 29, 30, 33, 34, 35, 43, 44, 46, 47, 63 Protoporphyrin precursors 31 Protosyphon 56 Prototheca 3, 28, 32, 39, 55 Protozoa 4, 29, 42 Puffer fish 57, 58, 51 Pulmonary disorders, treatment of 16 Pyrexia 15, 16 Pyridoxine 40 Pyrrophyta 3, 27 Pyrimidine 39 Pyruvic acid 34, 38, 39 R-value 30, 34 Radiant energy 29, 33, 34 Radiation 32, 39, 47, 48 Radioactivity 21, 47, 48 Rashes 14. Also see Skin disorders Red Sea 27, 45 "Red snow" 45 "Red tide" 45, 54, 55 "Red water" 27, 45, 56 Reducing sugars 7, 36 Renal disorders 14, 52, 60, 64. Also see Urinary disorders Reproduction 2, 32, 33, 35. Also see Sexuality Respiratory manifestations 48, 49, 56, 61, 64 "Reversible hydrocolloid tech- nic" 19 Rheumatism 14 Rhizoclonum 16 Rhodomela 20 Rhodophyceae 8, 31 Rhodophyta 3, 6 Rhodymenia 3, 6, 8, 10 Riboflavin 10, 39, 40 Ribonucleic acid 35 Rice cultivation 36 Rickets 9, 10, 29 Rock moss 1 Rotifera 4 Roughage 15 Rubidium 40 Saccharina 9 Safranal 33 Salt 12, 14, 25, 27, 63 Sargassum 14 Sausage casings 7 Scabies 14 Scenedesmus 3, 28, 33, 36, 37, 41, 42, 44, 47 Scbizothrix 36 Scombroid poisoning 61, 62 Scrofula 14, 15, 16 Scurvy 14 Sea, color of 27, 45 Sea kale 6 "Sea lettuce" 6, 9, 14 "Seabathers' eruption" 54, 55 Seaweed 2, 3, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 16, 17, 20, 21, 22, 36, 37, 39, 63, 65. Also see Macroscopic algae bread 7, 12 dye 22 glue 22 "gums" 17 isenglass 6 Sensitivity to algae 18. Also see Allergy Serine 8 Seven Little Sisters 26 Sewage disposal 43, 45, 46, 47 Sewage lagoons 47 Sexuality 33. Also see Repro- duction Shellfish poisoning 56, 60, 62, 64 Shipwreck survivors 23, 25, 26, 63 Shock 21 Shrimp 4, 25, 26 Silica 3 Silver 40 Skin disorders 14, 17, 19, 44, 49, 52, 53, 54, 55, 64 "Slimming teas" 16 Sodium 9, 17, 19 Soil, alkalinity of 36 Soil conservation 11, 36 "Solar engines" 29 Sorbitol 8 Space ships 64 Species of algae 2, 27 Sprains 14 Starch 8, 36 Stearic acid 37, 38 Sterols 2, 9, 37 Stichococcus 3, 30, 56 INDEX 85 Stilophora 16 Stipites 16 Stipes 16 Strontium 40 Succinamide 35 Sucrose 32, 36 Sugar wrack 13 Sulfates 9, 46 Sulfur 40, 41 Surgical powder 18 "Swayback" 12, 65 Sweating, prevention of 16 Synura 41 Syphilis 13 Taste of algae and plankton 45, 46, 47, 48 Temperature, effects on cul- tures 41, 42 Tetraodon 57, 61 Tetraodontoxin 61 Therapeutic value of algae 13, 14, 38, 44 Thiamin 10, 38, 39, 40 Thiazole 39 "Three dimensional farming" 63 Throat disorders 13, 16 Threonine 8, 35 Thyroid gland 15, 16. Also see Goiter, Iodine Tin 40 Titanium 40 Tocopherol 10 Tolypothrix 36 Toxicity 11, 18, 45, 48, 49, 50, 51, 52, 53, 54, 56, 57, 58, 59, 60, 61, 62, 64, 65 clinical findings 48, 49, 52, 54, 61, 64 differential diagnosis 48, 52, 54, 64, 65 pathologic findings 49, 52, 60, 61 symptomatology 54, 64 treatment 62 Toxins, chemistry of 52, 56, 60, 61 Trace elements 17, 40, 41 Transaminase systems 34 Transamination 36 Trehalose 8 Trichomonal vaginitis, treat- ment of 17 Triglycerides 37 Tryptophane 35 Tungsten 40 "Ultraplankton" 4 Viva 3, 6, 9, 10, 30 Unsaturated fatty acids 37, 38 Uranium 40 Urinary disorders 15, 16. Also see Renal disorders Valine 8, 35 Vanadium 40 "Vasserbloom" 45 Vermifuges 16 Vermilion Sea 27, 45 Vitamins 2, 7, 9, 10, 29, 37, 38, 39, 40, 44, 63 Water, coloration of 27, 45 purification of 53 supply 45, 53 taste 45, 46, 48 "Waterbloom" 45, 65 "Wasserbluthe" 45 Whales, food of 4, 23 World population 1, 27, 29, 63 Wracks 15, 22 Xylan 8 Xylose 8 "Yellow tide" Yttrium 40 Zinc 40 45 Zooplankton 3, 4, 22, 23, 24, 27, 63