NOAA TR NMFS SSRF-644 A UNITED STATES DEPARTMENT OF COMMERCE PUBLICATION NOAA Technical Report NMFS SSRF-644 89 U.S. DEPARTMENT OF COMMERCE National Oceanic and Atmospheric Administration National Marine Fisheries Service SEATTLE, WA February 1972 Inhibition of Flesh Browning and Skin Color Fading in Frozen Fillets of Yelloweye Snapper [Lutjanus vivanus) HAROLD C. THOMPSON, JR. and MARY H. THOMPSON Wltfint blotofteal laboratory LIBRARY seputttt Woodt Hole, lYiass. NOAA TECHNICAL REPORTS National Marine Fisheries Service, Special Scientific Report-Fisheries Series The major responsibilities of the National Marine Fisheries Service (NMFS) are to monitor and assess the abundance and geographic distribution of fishery resources, to understand and predict fluctuations in the quantity and distribution of these resources, and to establish levels for optimum use of the resources. 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Laboratory tests of an electrical barrier for con- trolling predation by northern squawfish. By Galen II. Maxfield, Robert H. Lander, and Charles D. Volz. July 1970, iii + 8 pp., 4 figs., 5 tables. 612. The Trade Wind Zone Oceanography Pilot Study. Part VIII: Sea-level meteorological properties and heat exchange processes, July 1963 to June 1965. By Gunter R. Seckel. June 1970, iv + 129 pp., 6 figs., 8 tables. 613. Sea-bottom photographs and macrobenthos col- lections from the Continental Shelf off Massa- chusetts. By Roland L. Wigley and Roger B. Theroux. August 1970, iii + 12 pp., 8 figs., 2 tables. 614. A sled-mounted suction sampler for benthic or- ganisms. By Donald M. Allen and J. Harold Hudson. August 1970, iii -f 5 pp., 5 figs., 1 table. 615. Distribution of fishing effort and catches of skip- jack tuna, Katsuwonus pelamis, in Hawaiian waters, by quarters of the year, 1948-65. By Richard N. Uchida. June 1970, iv + 37 pp., 6 figs., 22 tables. 616. 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Stans, Secretary NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION Robert M. White, Administrator NATIONAL MARINE FISHERIES SERVICE Philip M. Roedel, Director NOAA Technical Report NMFS SSRF-644 Inhibition of Flesh Browning and Skin Color Fading in Frozen Fillets of Yelloweye Snapper (Lutjanus vivanus) HAROLD C. THOMPSON, JR. and MARY H. THOMPSON Marine Biological Laboratory LIBRARY SFP 1 3 1972 Woods Hole, Mass. SEATTLE, WA February 1972 For sale by the Superintendent of Documents, U.S. Government Printing Office Washington, D.C., 20402 - Price 25 cents The National Marine Fisheries Service (NMFS) does not approve, rec- ommend or endorse any proprietary product or proprietary material mentioned in this publication. No reference shall be made to NMFS, or to this publication furnished by NMFS, in any advertising or sales pro- motion which would indicate or imply that NMFS approves, recommends or endorses any proprietary product or proprietary material mentioned herein, or which has as its purpose an intent to cause directly or indirectly the advertised product to be used or purchased because of this NMFS publication. CONTENTS Page Introduction 1 Materials and methods 2 Preparation of samples 2 Yelloweye snapper (Lutjanus vivanus) 2 Chemicals 2 Method of chemical application 2 Packaging 3 Storage 3 Organoleptic evaluation of fillets 3 Chemical methods 3 Total ribose determination 3 Free ribose determination 3 Results 3 Discussion 4 Literature cited 5 TABLES Page 1. Organoleptic scores for cooked snapper fillets stored at — 10° F . . . . 3 2. Appearance scores for cooked snapper fillets stored at — 10° F 3 3. Free and total ribose content of snapper flesh stored at — 10° F . . . . 3 in Inhibition of Flesh Browning and Skin Color Fading in Frozen Fillets of Yelloweye Snapper (Luijanus vivanus) By HAROLD C. THOMPSON, JR. and MARY H. THOMPSON1 National Marine Fisheries Service Fishery Products Technology Laboratory Pascagoula, Mississippi 39567 ABSTRACT Fresh yelloweye snapper (Lutjanus vivanus) which had been scaled and evis- cerated were treated with the chemicals 3,3'-thiodipropionic acid, glutathione, di- sodium ethylenediaminetetraacetate dihydrate in combination with propyl gallate, and monotertiary butylhydroquinone. The inhibitory effects that these chemicals had on flesh browning (Maillard reaction) were studied over a 12-month frozen storage period. Also, the effects of vacuum packaging on snapper skin discolor- ation were studied. INTRODUCTION The Gulf of Mexico red snapper fishery ori- ginated off the northwest Florida coast 15 to 20 years prior to the Civil War (Carpenter, 19(55). Catches at that time were made with the use of handlines from sailing vessels con- taining live wells. At that time, ice for use on commercial vessels was too expensive owing to cost of shipping from the point of manu- facture. The snapper fishery did not start on a large scale until after the end of the Civil War ( War- z-en, 1898). The fishery really began to bloom when ice manufacturing plants began produc- ing ice at a reasonable cost. With ice to cool their catches, the crews of the snapper vessels were able to go out farther and stay longer. 1 Authors: Harold C. Thompson, Jr., Chemist, and Mary H. Thompson, Research Chemist, National Ma- rine Fisheries Service, formerly Fishery Products Technology Laboratory, Pascagoula, Miss. 39567, pre- sently Southeast Fisheries Center, Miami Laboratory, Miami, Fla. 33149. Another boost was given to the fishery around 1920 when sail-rigged vessels became diesel powered. The majority of the snapper fleet still ices the fish in the hold; however, a few new freezer vessels have entered the fleet. The total landings of snapper in the United States for 1968 were 11,500,000 lb. valued at $3,700,000 while the totals for 1969 were 9,500,000 lb. valued at $4,000,000 (Riley, 1970) . The present methods used in commercial pro- cessing of snapper are limited to (1) eviscer- ation and freezing the whole fish and (2) fil- leting or steaking the fish and then freezing them. The majority of the snapper are either sold in a fresh iced condition or as eviscerated frozen fish. The flesh of the fillets and steaks turns from white to brown in color during ex- tended periods of frozen storage, thus limiting the market owing to lack of consumer eye appeal. The red skin on the fillets and steaks, one of the snapper's main selling points, also fades in color to a beige or bronze during frozen storage. Skin coloration is used by most large firms as a quality indicator. There are three broad types of browning re- actions recognized in foods. The first is car- amelization which occurs when polyhydroxy- carbonyl compounds are heated to relatively high temperatures in the absence of amino acids. The second is a group of oxidative re- actions which convert ascorbic acid and poly- phenols into di- or polycarbonyl compounds. The third type, which is the most common, in- volves the reactions of aldehydes, ketones, and reducing sugars with amines, amino acids, pep- tides, and proteins and is known as the Maillard reaction. Liston et al. (1963) state that the free amino groups of fish proteins are available for re- actions with reducing sugars or with some of the products of oxidation of lipid material. The end result of a complicated series of re- actions is the formation of brown colors as well as changes in flavor. According to Von Tigerstrom and Tarr (1965), the browning which occurs in muscles of many species of fish post mortem is due to the Maillard reaction. Von Tigerstrom and Tarr (1965) also brought out the importance of D-ribose as playing a significant role in the Maillard reaction. Other researchers have also indicated that ribose is one of the reducing sugars which is a main contributor to the Maillard reaction (Tarr and Gadd. 1965; Pom- eranz et al,, 1962; Gilka, 1963; and Jones, 1962). Most browning inhibitors have been shown to be carbonyl reagents such as cyanide (Barnes and Kaufman, 1947), hydroxylamine (Burton, 1945) , hydrazine (Barnes and Kaufman, 1947) , mercaptans (Guss, 1952; Song and Chichester, 1967), bisulfite (Olcott, 1953), and glutathione (Joslyn and Ponting, 1951). Most of these chemicals, however, have undesirable side effects in meat and fish. Other methods used in inhibiting the flesh browning are (1) low storage temperatures and (2) lowering the moisture content in dehydrated foods. In spite of the commercial importance of good skin color of red snapper, there has been very little work done on preventing skin color fading. According to Tsukuda and Amano (1966) , storage temperatures as low as —30° C did not prevent skin discoloration in Lepido- trigla giintheri, Lusk et at, (1964) reported the changes in the amount of astacene pigment in freeze-dried shrimp during freeze dehydra- tion and subsequent storage both in air atmos- phere and a nitrogen atmosphere. The pigment in shrimp stored in the air atmosphere was essentially all bleached after one week whereas shrimp stored in a nitrogen atmosphere re- tained their pigment color over the same period. Yamaga and Morioka (1962) described the effectiveness of certain antioxidants used to prevent the skin color fading in certain red fishes. The purposes of this paper are to describe an experiment designed to show (1) the in- hibitory effects that certain chemicals have on the browning of the flesh of frozen snapper fillets and (2) the effect vacuum packaging has on preventing the skin discoloration of frozen snapper fillets. MATERIALS AND METHODS Preparation of Samples Yelloweye snapper (Luijanus vivanus). — Fifty-two fresh, iced, eviscerated, heads-on snapper were obtained from a local fishery. These fish were scaled and headed. Their weights averaged approximately 3 lb. Chemicals. — All chemicals were either food grade or reagent grade quality. The chemicals used in this study were 3,3'-thiodipropionic acid (TDP), glutathione (Glu), disodium ethylene- diaminetetraacetate dihydrate (Na2EDTA) in combination with propyl gallate (PG), and monotertiary butylhydroquinone (TBHQ). Method of chemical application. — -The technique involved using a hypodermic syringe to iniect a scaled eviscerated snapper prior to filleting with an aqueous solution of the desired chemical. Each side of the fish was injected with two 2-ml shots evenly spaced down the lateral line. The solutions of TDP and gluta- thione were injected in the amount necessary to produce a residual concentration of the chem- icals equivalent to 0.02 r'< of the snapper oil content. TBHQ was injected in an amount to produce a 50 ppm residual. A combination of Na2EDTA and PG was injected in the amount to produce a 50 ppm residual of the combina- tion. These chemicals can also be applied to fillets by a dipping- or injection technique as was demonstrated by this laboratory (Thomp- son and Thompson, 1969). Packaging. — All fillets were separately packaged in vacuum-sealed Cryovac plastic bags. Storage. — ■ All fillets were stored in a —10° F (—23.3° C) freezer until analyzed. Table 1 .--Organoleptic scores for cooked snapper fillets stored at -10° F.JV Storage period Sample 1 month i months 6 months 9 months 12 months Control 4.6 4.0 3.2 2.0 2.4 TDP - - 3.8 3.5 2.7 Glu 4.7 4.0 3.8 2.9 2.2 Na2EDTA + PG 4.5 4.0 3.3 3.6 1.8 TBHQ - - 3.0 2.9 2.2 _1/Values presented are averages of scores given for appearance, taste, and texture based on a 5-point scale. Organoleptic Evaluation of Fillets Fillets were evaluated organoleptically by an 8-member expert taste panel at the end of 1, 3, 6, 9, and 12 months of frozen storage. The fillets were scored on appearance, taste, and texture. The scoring method consisted of giv- ing a score of from 5 for excellent to 0 for in- edible. Table 2. --Appearance scores for cooked snapper fillets stored at -10° F.i/ Storage period Sample 1 month 3 months 6 months 9 months 12 months Control 4.8 4.0 4.2 2.2 2.6 TDP - - 5.0 3.7 2.7 Glu 4.4 4.0 5.0 2.9 2.4 Na2EDTA + PG 4.6 4.0 4.2 3.6 1.8 TBHQ - - 4.2 3.4 2.2 iyvalu.es presented are averages of scores given for appearance by taste panel members based on a 5-point scale. Chemical Methods Total ribose determination. — Total ribose was extracted from minced snapper flesh ac- cording to the method of Sinnhuber (1966). Total ribose content of the extract was deter- mined by the orcinol method of Mejbum (1939). Heating time was increased to 45 min as modified by Albaum and Umbreit (1947). Free ribose determination Free ribose was extracted from minced snapper flesh ac- cording to the method of Sinnhuber (1966). Bound ribose was removed from this extract using Dowex ion exchange resins according to the method of Tarr and Leroux (1962). Free ribose was then determined by the orcinol meth- od of Mejbum (1939) using an increased heat- ing time of 45 min as modified by Albaum and Umbreit (1947). RESULTS Organoleptically the glutathione treated fil- lets received the best average score (4.7) at the end of 1 month of storage (Table 1). By the Table 3. --Free and total ribose content of snapper flesh stored at -10° ?M Stora ^e period S Jmplc 1 month 2 months 3 months 'i mnnths 9 months 12 months fg/s UB/g mb/b ub/b UB/g ub/b Free ribose Control 13.64 12.93 13.24 11.14 10.43 9.71 TDP 10.87 10.33 11.01 5.19 4.78 8.57 Glu 11.10 10.47 7.61 10.27 10.44 10.71 Na2EDTA + PC 12.15 4.18 7.34 5.07 4.78 7.33 TBHQ 19.27 8.01 16.29 3.11 11 6.88 Total ribose Control 31.15 21.51 26.21 21.16 19.88 15.77 TDP 39.15 18.27 35.82 27.39 21.51 35.19 Glu 33.54 19.78 23.84 14.32 28.05 24.91 Na2EDTA + PG 36.88 25.24 28.24 20.01 35.72 23.91 TBHQ 38.04 17.21 28.72 24.43 17.35 25.70 _1/Values presented are averages of duplicate samples. ^'Sample was lost. end of 3 months of storage, the control, gluta- thione, and the Na-FDTA + PG treated fillets all received an average score of 4.0. Not until the 6 month draw was there a considerable dif- ference in organoleptic scores observed. Then both the TDP and glutathione treated fillets received a score of 3.8 while the control fillets received a score of 3.2. By the end of 12 months of storage the TDP treated fillets received the highest score (2.7) while the NA2EDTA + PC, treated fillets received the lowest score (1.8). After averaging scores for appearance only (Table 2), the TDP treated fillets received the highest score (2.7) at the end of 12 months of frozen storage while the Na2EDTA + PG treated fillets were given the lowest score for appearance (1.8) over the same storage period. Visual observations of the progression of browning are supported by changes in free and total ribose content of snapper flesh over a 12- month frozen storage period. The greatest de- crease in free ribose content of snapper fillets between 1 and 12 months of frozen storage was exhibited by fillets treated with TBHQ, NA2- EDTA + PG, and by the control (Table 3). The fillets treated with TDP and glutathione showed the least differences in free ribose con- tent over the same period. The fillets that showed the largest decreases in total ribose content over the 12-month storage period were the control, those treated with Na2EDTA + PG, and those treated with TBHQ. The least change in total ribose content over the 12-month storage period was observed for the TDP and glutathione treated fillets. By the end of 3 months of frozen storage, the flesh of the control, the Na2EDTA + PG, and the TBHQ treated fillets were noticeably turning brown. By the end of 6 months of storage, these fillets had turned even browner, approaching the color of a brown pasteboard box. The flesh of the TDP and glutathione treated fillets, however, was nearly as white as that of a freshly filleted snapper. The TDP treated fillets received the highest organoleptic score for appearance throughout the storage period. Between 9 and 12 months of frozen storage, the texture of all the fillets became woody or slightly tough. The skin color deterioration problem was successfully met by packaging the fillets in vac- uum-sealed Cryovac bags. None of the samples deteriorated appreciably with regard to skin color over the 12-month frozen storage period. There was no difference noted in skin color between the control fillets or the fillets treated with the chemicals used in this experiment. DISCUSSION The organoleptic scores for both TDP and TBHQ treated fillets at the 1- and 3-month draws were not obtained since both of these chemicals were still considered experimental. Subsequently, we learned that both chemicals were approved for use in foods by the Food and Drug Administration (FDA) and organ- oleptic studies were initiated on the fillets treated with these chemicals. Bound ribose is present in fish flesh in the form of nucleic acids such as RNA, nucleotides such as IMP, and nucleosides such as inosine, to name a few. The majority of the free ribose present in fish flesh is produced by the post mortem degradation of one or more of the gen- eral classes of compounds mentioned above. Several researchers (Von Tigerstrom and Tarr, 1965; Tarr and Gadd, 1965; and Pomeranz et al., 1962) have indicated that it is the free ribose which is involved in the Maillard reaction in post mortem browning of fish muscle. The test pack of red snapper was stored in a —10° F (—23.3° C) freezer. At the slightly warmer storage temperature of — 18° C, the nucleotide IMP (inosine monophosphate) in frozen swordfish steaks is degraded at the rate of only 0.(129 /^moles per gram per week. IMP degrades at the rate of 0.24 /umoles per gram per week at —8° C (Dyer and Hiltz, 1969). This type of degradation results in a change in the free and bound ribose content of fish flesh. According to Dyer et al. (1966) , no sig- nificant change in content of IMP, inosine, and hypoxanthine in the ordinary muscle of fast- frozen swordfish steaks occurs during freezing and frozen storage at — 26° C for a period of from 4 to 5 months. Therefore, at the tem- perature at which this pack was stored (ap- proximately — 26° C), no changes in free and bound ribose due to enzymatic degradation of nucleotides and nucleosides should have oc- curred. Consequently, the progress of brown- ing in the frozen snapper flesh can be followed by analyzing the flesh for free and total ribose. As browning progresses in frozen snapper flesh, the amount of free ribose decreases due to a reaction between ribose and a free amino acid or the amino group of a protein; this re- action is the first of a series of complicated reactions which occur in the browning process. The greatest decrease in free ribose content occurred in the control fillets and those which had been treated with TBHQ and Na2EDTA + PG. The least amount of decrease in free ribose content occurred in those fillets which had been treated with TDP and glutathione. These free ribose data parallel the visual ob- servations of the progression of the browning; of the snapper flesh. The flesh of the TDP treated fillets, when visually observed, was con- sistently lighter in color at each draw than the flesh of any of the other fillets. Taking into account the ribose data, organ- oleptic scores, and visual observations of the progression of browning, the TDP treated fil- lets showed a much better inhibition of brown- ing, as well as preservation of overall quality, than fillets treated with other chemicals. The foregoing combination of observations indi- cates that browning in snapper flesh is that of the Maillard type. The skin color deterioration of frozen snap- per fillets is an oxidative process whereby the red skin pigment of snapper, astaxanthin, is converted to astacene or further degradation products. The red pigment in marine fishes is predominantly astaxanthin (Tsukuda and Amano, 1966). In an earlier experiment con- ducted at this laboratory, several of the more common antioxidants used in the food industry were applied to snapper fillets (Love and Thompson, 1969). Application of these anti- oxidants deters the fading of the skin color for only a few months. In the present exper- iment, however, packaging the fillets in Cryo- vac vacuum-sealed plastic bags inhibited the oxidative process sufficiently to increase the frozen storage shelf life to 12 months. After this period, a slight discoloration or fading of the red pigment was noted. This indicated that the oxidative process which converts the red pigment, astaxanthin, to astacene or prod- ucts further down the degradation chain is not entirely due to the presence of air. Enzymatic oxidation also probably plays an important role in the skin color deterioration of red snapper. The application of TDP to red snapper fillets followed by Cryovac vacuum packaging and subsequent freezing of the product results in a superior quality product for a 12-month storage period. The TDP can be applied by either dipping or injecting the fillets as was shown by previous experiments at this labora- tory (Thompson and Thompson, 1969) . Either method of application produces the same re- sults. This treatment is by no means a "cure- all" for the snapper industry, but it is a be- ginning since the previous storage life of a frozen snapper fillet was somewhere between 1 and 3 months. LITERATURE CITED ALBAUM, H. G., and W. W. UMBREIT. 1947. 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Fish. 32: 334-345. WARREN, A. F. 1898. The red snapper fisheries, their past, present, and future. Bull. U.S. Fish Comm. 17 (1897): 331-335. YAMAGA, M., and K. MORIOKA. 1962. Chemical preservation of the body color of red fishes. Bull. Jap. Soc. Sci. Fish. 28: 1104-1108. GPO 999-437 MBL WHOI Library > nil, I, III 5 WHSE 01824 621. Predation by sculpins on fall chinook salmon, Oncorhynchus tshawytscha, fry of hatchery or- igin. By Benjamin G. Patten. February L971, iii + 14 pp., 6 figs., 9 tables. 622. Number and lengths, by season, of fishes caught with an otter trawl near Woods Hole, Massa- chusetts, September 1961 to December 1962. By F. E. Lux and F. E. Nichy. February 1971, iii + 15 pp., 3 figs., 19 tables. Apparent abundance, distribution, and migra- tions of albacore, Th minus alalunga, on the North Pacific longline grounds. By Brian J. Rothschild and Marian Y. Y. Yong. September 1970, v + 37 pp., 19 figs., 5 tables. 624. Influence of mechanical processing on the quality and vield of bay scallop meats. Bv N. B. Webb and F. B. Thomas. April 1971, iii + 11 pp., 9 figs., 3 tables. 625. Distribution of salmon and related oceanographic features in the North Pacific Ocean, spring 1968. By Robert R. French, Richard G. Bakkala, Ma- sanao Osako, and Jun Ito. March 1971, iii + 22 pp., 19 figs., 3 tables. 626. Commercial fishery and biology of the fresh- water shrimp, Macrobrachium, in the Lower St. Paul River, Liberia, 1952-53. By George C. Mil- ler. February 1971, iii + 13 pp., 8 figs., 7 tables. 627. Calico scallops of the Southeastern United States, 1959-69. By Robert Cummins, Jr. June 1971, iii + 22 pp., 23 figs., 3 tables. 628. Fur Seal Investigations, 1969. By NMFS, Ma- rine Mammal Biological Laboratory. August 1971, 82 pp., 20 figs., 44 tables, 23 appendix A tables, 10 appendix B tables. 629. Analysis of the operations of seven Hawaiian skipjack tuna fishing vessels, June-August 1967. By Richard N. Uchida and Rav F. Sumida. March 1971, v + 25 pp., 14 figs., 21 tables. For sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - 35 cents. 630. Blue crab meat. I. Preservation bv freezing. July 1971, iii + 13 pp., 5 figs., 2 tables! II. Effect of chemical treatments on acceptability. By Jurgen H. Strasser, Jean S. Lennon, and Fred- erick J. King. July 1971, iii + 12 pp., 1 fig., 9 tables. 631. Occurrence of thiaminase in some common aquat- ic animals of the United States and Canada. By R. A. Greig and R. H. Gnaedinger. July 1971, iii + 7 pp., 2 tallies. 632. An annotated bibliography of attempts to rear the larvae of marine fishes in the laboratory. By Robert C. May. August 1971, iii + 24 pp., 1 ap- pendix I table, 1 appendix II table. For sale by the Superintendent of Documents, U.S. Govern- ment Printing Office, Washington, D.C. 20402 - 35 cents. 633. Blueing of processed crab meat. II. Identification of some factors involved in the blue discoloration of canned crab meat Callinectes sapidns. Bv Melvin E. Waters. May 1971, iii + 7 pp., 1 fig!, 3 tables. 634. Age composition, weight, length, and sex of her- ring, Clupea pallasii, used for reduction in Alas- ka, 1929-66. By Gerald M. Reid. July 1971, iii + 25 pp., 4 figs., 18 tables. 635. A bibliography of the blackfin tuna, Thunnus atlanticus ( Lesson ) . By Grant L. Beardsley and David C. Simmons. August 1971, 10 pp." For sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - 25 cents. 636. Oil pollution on Wake Island from the tanker R. ('. Stoner. Bv Reginald M. Gooding. May 1971, iii + 12 pp., 8 figs., 2 tables. For sale by the Superintendent of Documents, U.S. Govern- ment Printing Office, Washington, D.C. 20402 - Price 25 cents. 637. Occurrence of larval, juvenile, and mature crabs in the vicinity of Beaufort Inlet, North Carolina. By Donnie L. Dudley and Mayo H. Judy. August 19*71, iii + 10 pp., 1 fig., 5 tables. For sale bv the Superintendent of Documents, U.S. Govern- ment Printing Office, Washington, D.C. 20402 - Price 25 cents. 638. Length-weight relations of haddock from com- mercial landings in New England, 1931-55. By Bradford E. Brown and Richard C. Hennemuth. August 1971, v -f 13 pp., 16 fig., 6 tables, 10 appendix A tables. For sale by the Superintend- ent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - Price 25 cents. 639. A hydrographic survey of the Galveston Bay system, Texas 1963-66. By E. J. Pullen, W. L. Trent, and G. B. Adams. October 1971, v + 13 pp., 15 figs., 12 tables. For sale by the Super- intendent of Documents, U.S. Government Print- ing Office. Washington. D.C. 20402 - Price 30 cents. 640. Annotated bibliography on the fishing industry and biology of the blue crab, Callinectes sapidus. By Marlin E. Tagatz and Ann Bowman Hall. August 1971, 94 pp. For sale by the Superinten- dent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - Price $1.00. 641. Use of threadfin shad, Dorosoma petenense, as live bait during experimental pole-and-line fish- ing for skipjack tuna, Katsuwonus pelamis, in Hawaii. By Robert T. B. Iversen. August 1971, iii + 10 pp., 3 figs., 7 tables. For sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - Price 25 cents. 642. Atlantic menhaden Brevoortia tyrannus resource and fishery — analysis of decline. By Kenneth A. Henry. August 1971, v + 32 pp., 40 figs., 5 appendix figs., 3 tables, 2 appendix tables. For sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402 - Price 45 cents. 646. Dissolved nitrogen concentrations in the Colum- bia and Snake Rivers in 1970 and their effect on chinook salmon and steelhead trout. By Weslev J. Ebel. August 1971, iii -f 7 pp., 2 figs., 6 tables. For sale by the Superintendent of Doc- uments, U.S. Government Printing Office, Wash- ington, D.C. 20402 - Price 20 cents. UNITED STATES DEPARTMENT OF COMMERCE NATIONAL OCEANIC & ATMOSPHERIC ADMINISTRATION NATIONAL MARINE FISHERIES SERVICE SCIENTIFIC PUBLICATIONS STAFF BLDG. 67. NAVAL SUPPORT ACTIVITY SEATTLE, WASHINGTON 98115 OFFICIAL BUSINESS POSTAGE AND FEES PAID U.S. DEPARTMENT OF COMMERCE