Taste Components of Japanese Spiny and Shovel-Nosed Lobsters

Fisheries Science 62(2), 283-287 (1996)

Takaaki Shirai,*1 Yoshinori Hirakawa,*1 Yoshimi Koshikawa,*1 Hiroshi Toraishi,*1 Makoto Terayama,*2 Takeshi Suzuki,*1 and Toshiyuki Hirano*1 *1Department of Food Science and Technology , Tokyo University of Fisheries, Konan, Minato, Tokyo 108, Japan *2The Miyazaki Prefectural Fisheries Experimental Station , Aoshima, Miyazaki, Miyazaki 889-21, Japan

(Received July 10, 1995)

Methanol extracts were prepared from the abdominal muscles of Japanese spiny and shovel-nosed lobsters and free amino acids, ATP related compounds, betaines, and other components in the extracts were analyzed. The two lobsters were rich in glycine, arginine, glycine betaine, trimethylamine oxide, glutamine, taurine, homarine, proline, alanine, ADP, Na+, K+, P043-, and Cl-. Glycine, arginine, pro line, alanine, glutamic acid, glycine betaine, trimethylamine oxide, AMP, IMP, NaCl, KCl, and KH2PO4. They were confirmed to be taste-active for the Japanese spiny lobster taste by the sensory taste test. As for shovel-nosed lobster, valine, isoleucine, leucine, sarcosine, and methionine were taste active in addition to the above 11 components except IMP.

Key words: Japanese spiny lobster, shovel-nosed lobster, taste, glycine, betaine, IMP, extractive components

Japanese spiny lobster Panulirus japonicus is tasty and Proximate Composition one of the most palatable lobsters. Shovel-nosed lobster The lobster abdominal muscle was dried in an air-oven Ibacus ciliatus is also popular in the western and southern at 108•Ž and the moisture was calculated. Crude protein parts of Japan and is known as having a thick tasty body. of the muscle was analyzed by the Kjeldahl method. Lipid The taste of shovel-nosed lobster is said to be similar to of the muscle was extracted with chloroform-methanol that of Japanese spiny lobster. (2:1) mixture3) and dried in an air-oven. Ash of the muscle Though many works on extractive components of fish was determined with an oven at 500•Ž. and shellfish muscles have been reported, few studies have correlated these analytical data directly with their tastes.1,2) Extract It is known that the taste of shellfish is produced by several Two grams of the muscle was homogenized with 25 ml free amino acids, 5•Œ-adenylic (AMP) or 5•Œ-inosinic acid of 100% methanol in a glass homogenizer and the

(IMP), NaCl, and other extractive components.2) To reveal homogenate was centrifuged at 8,000 rpm and 4•Ž for 10 the extractive components of Japanese spiny and shovel min. The residue was extracted twice more with 25ml of nosed lobsters and to evaluate their taste-active compo 100% methanol in the same manner. The supernatants nents, we analyzed their extractive components and then a were combined, evaporated to remove methanol, and then sensory taste test was carried out with the taste test solu defatted with 50ml of diethyl ether. The extract was evapo tions prepared with commercial chemicals and on ob rated and diluted to 50 ml with water. tained data. Extractive Components Materials and Methods Free amino acids were measured with a JEOL JLC-300 amino acid analyzer. Lobsters ATP and related compounds were determined by the The five individuals each of male and female Japanese HPLC method with an Asahipack GS-320 column

spiny lobsters being 15-17 cm body length and 99-123 g (7.6 •~ 500mm), 0.2M NaH2PO4 eluate (pH 3.00), a flow body weight were collected in October 1993 locally at Udo, rate of 1ml/min, and detection at 254 nm. Miyazaki Prefecture. The five individuals each of male and Cyclic and ?-betaines were respectively analyzed by the female shovel-nosed lobsters being 13-15 cm body length HPLC methods of a Whatman Partisil 10-SCX column and 90-140 g body weight were also collected in June 1993 (4.6 •~ 250mm), 0.03 M KH2PO4 eluate (pH 2.10), a flow locally at Kawaminami, Miyazaki Prefecture. The lobsters rate of 1 ml/min, and detection at 262nm and a Cica were landed, frozen immediately, and shipped to the MERCK Hibar LiChroCART LiChrospher 100 NH2 laboratory, Tokyo through a night at -18•Ž. column (4 •~ 250mm), 75% acetonitrile eluate, a flow rate The lobsters shelled under a frozen condition. The ab of 1ml/min, and a refractive index detector. dominal muscles were removed from the tail part of the Trimethylamine and its oxide were measured with a Con lobsters and used in this study. way glass unit and picric acid toluene solution. The 1ml 284 Shirai et al. portion of the extract and 1ml of 20% formalin contain Table 1. Proximate composition of the abdominal muscles of the ing 5% MgCO3 were placed and mixed in an outer room of Japanese spiny and shovel-nosed lobsters a Conway unit and stood for 3 min. Then the saturated

(about I%) picric acid toluene solution was placed in an in ner room of the unit. Just after adding 1ml of saturated K2CO3 solution to the outer room and mixing gently, the unit was covered air-tight and stood at 37•Ž for 120min. The picric acid toluene solution was taken out and diluted to 5 ml with toluene. The optical density of the toluene so lution was measured at 410nm. Trimethylamine oxide was Values are means of 5 samples with SD. reduced with 1% TiC13 at 80•Ž for a few seconds 4) and trimethylamine formed was determined as described above. Trimethylamine oxide was calculated from this shovel-nosed lobsters and between females of both lob trimethylamine increment. sters. Superscript indicates significantly higher concentra Na+ and K+ were analyzed with a Shimadzu AA-660 tion (p<0.05). atomic absorption analyzer. For K+ analysis, NaCl was Among nitrogenous extractive components, taurine, added to the diluted extract to make a final concentration glutamine, glycine, alanine, arginine, proline, homarine, of 0.04%. glycine betaine, and trimethylamine oxide were the major Cl- was determined by the mercury thiocyanate components of the abdominal muscle of Japanese spiny method5) and P043 by the molybdenum blue method6) after lobsters. Especially, more than 1,000mg/ 100g of glycine refluxed digestion of the extract (1 ml) with 2ml of 60% was detected in male and female of Japanese spiny lob perchloric acid. sters. About 100mg/ 100g of IMP and slight amounts of Extractive nitrogen was determined by the Kjeldahl N-methylisonicotinic acid which is an isomer of homarine method. were also detected in this lobster. On the other hand, taurine, glutamine, glycine, argi Sensory Taste Test nine, proline, homarine, glycine betaine, and trimethyl Twenty-two subjects participated in the sensory taste amine oxide were the major components of the abdominal test and their mean age was 25 years. All subjects had previ muscle extract of shovel-nosed lobster. In contrast to ously participated in sensory taste tests and could dis Japanese spiny lobster, IMP seemed to be absent in criminate not only sweet, salty, sour, and bitter qualities shovel-nosed lobster. but umami taste as well, using with diluted solutions of gly The difference in the extractive component composi cine and alanine, NaC1, H3P04, KCl and KH2P04, and so tions between sexes of each lobster was not significant, ex dium glutamate and IMP, respectively. cept ATP-related compounds. The compositions of extrac Taste profiles of the natural extract and full component tive components of male and female Japanese spiny taste test solution were compared at room temperature lobsters were compared statistically (p<0.05) with those and open panel discussion. of male and female shovel-nosed lobsters, respectively. The other sensory taste test was run by the triangle differ Amounts of glycine of male and female Japanese spinylob ence test at room temperature. Subjects were presented sters were respectively about twice as large as those of male with a row of three 30 ml-brown glass cups containing 10 and female shovel-nosed lobsters. The homarine amount ml each of the extract or a test solution at each time in a of Japanese spiny lobster seemed to be larger than that of room with three isolated compartments. Subjects scored in the shovel-nosed lobster. The concentrations of glycinebe tensities of sweetness, saltiness, sourness, bitterness, uma taine of male and female Japanese spiny lobsters were mi, and taste body as -3 "very weak," -2 " weak," -I almost half of those of male and female shovel-nosed lob "slightly weak ," 0 "the same," + 1 "slightly strong," +2 sters, respectively. Concentrations of several components, "strong ," and +3 "very strong." such as threonine, valine, isoleucine, leucine, and trimethylamine oxide, were higher in both sexes of shovel Results and Discussion nosed lobsters than in these of Japanese spiny lobsters. Konosu and Yamaguchi1)reviewed the flavor components Proximate Composition in fish and shellfish. In Japanese spiny and shovel-nosed Table 1 shows moisture, protein, lipid, and ash of the ab lobsters, respectively, large amounts of glycine, 1078and dominal muscles of male and female Japanese spiny and 1159 mg/ 100 g, and arginine, 678 and 658 mg/ 100 g, were shovel-nosed lobsters. Results in the table are means of 5 detected, which were followed by alanine, 42 and 104 mg/ samples with standard deviation. There was almost no sig 100 g, proline, 116 and 36 mg/ 100 g, and taurine 68 and nificant difference between male and female individuals or 89 mg/ 100 g. Among other extractive components, large between Japanese spiny and shovel-nosed lobsters, being amounts of glycine betaine, 928 and 677 mg/ 100 g, were 77-80% moisture, 18-21 % protein, 1% lipid, and 2% ash. also detected in the Japanese spiny and shovel-nosed lob ster muscles, respectively. Except a few amino acids, such Composition of Extractive Components as taurine, serine, and alanine, almost the same amounts Table 2 shows compositions of extractive components were found in the muscles of both lobsters , respectively,in of the abdominal muscles of male and female Japanese this study. Thus the free amino acid compositions of spiny and shovel-nosed lobsters. The t-test was performed Japanese spiny and shovel-nosed lobsters were character on the analyzed data between males of Japanese spiny and ized by high concentrations of glycine, arginine , and sever Taste Components of Lobsters 285

Table 2. Extractive components of the abdominal muscles of the Japanese spiny and shovel-nosed lobsters (mg/ 100g)

Values are means of 5 samples with SD. The sum of extractive components was divided by the dry matter and is expressed as the dry-matter recovery. * Significantl y different at p<0.05 between the male spiny and shovel-nosed lobsters and between these female lobsters.

al betaines and are similar to those of prawns and shrimps, 0.37-1.09 ƒÊmol/g, was found in both lobsters. Adenosine such as the coonstripe shrimp Pandalus hypsinotus, pink was reported to be rare in the crab, lobster, and prawn shrimp P. borealis,1) kuruma prawn Penaeus japonicus,7) muscles under both of the fresh condition and during the and mantis shrimp Oratosquilla oratoria.8) 10-day ice-storage.10) The feature of free amino acid composition of the shov Amounts of dry matter of the Japanese spiny and el-nosed lobster, being very rich in glycine, arginine, and shovel-nosed lobster extracts ranged from 5300 to 5600 betaines and relatively rich in other several amino acids mg/100g. The total extractive components shown in was similar to that of crab.9) A small amount of adenosine, Table 2 was divided by the dry matter and expressed as the 286 Shirai et al. dry-matter recovery. These recoveries were 98-107%, Table 3. Composition of the taste test solutions for the sensory showing that almost all components in the extracts were taste test of Jananese sninv and shovel-nosed lobsters analyzed in this study. The extractive-nitrogen recovery (mg/100ml) was calculated by dividing the sum of the nitrogen amounts of the analyzed nitrogenous components by the extractive nitrogen. These recoveries for the lobster ex tracts were also satisfactory, being 93-103%.

Sensory Taste Test Since almost all extractive components of Japanese spiny and shovel-nosed lobsters were analyzed in this study, the taste test solutions were prepared by reagent grade commercial chemicals according to the analytical data. These test solutions designated as the full component test solutions were used for the sensory taste test. The sen sory taste test was done initially using the full component test solutions formulated from about 40 commercial chemi cals, regarding mg/ 100 g muscle listed in Table 2 as mg/ 100 mt water excepting phosphoserine, ƒ¿-aminobutyric acid, ammonia, N-methylisonicotinic acid, trigonelline, ATP, and hypoxanthine. NaCl, KCl, and KH2PO4 represented Na+, K+, Cl-, and P043-. The sensory taste test began with the open panel discus sion and the subjects stated that the full component test so lutions for the Japanese spiny and shovel-nosed lobsters Table 4. Sensory taste test and t-values for the taste intensity of the respectively possessed fairly well the lobster tastes. test solutions of shovel-nosed lobster against that of the Japanese spiny lobster The full component test solution for Japanese spiny lob ster was judged to own its taste definitely with strong sweet ness and bitterness. Almost all subjects also stated that dis tinguishable umami, saltiness, and sourness were present in this test solution. The full component test solution for the shovel-nosed lobster also judged to have strong sweet ness and bitterness. Umami, saltiness, and sourness were stated to be present in this solution perceptibly. All sub jects stated that the taste body of the full component test solution for the shovel-nosed lobster was stronger and n=22. thicker than that of the full component test solution for * The shovel-nosed lobster taste was significantly (p<0.01) different from the Japanese spiny lobster. Japanese spiny lobster taste. Successively one or several components were omitted ev ery time from the sensory tests and separate from the full component test solutions for Japanese spiny and shovel components of the boiled snow crab meat,2) glycine, ala nosed lobsters. Then attention was paid to the characteris nine, glutamic acid, arginine, glycine betaine, AMP, gua tics of those lobster tastes until each of the taste character nylic acid, cytidylic acid, Na+, K+, Cl and P043- were istics of the test solutions was weakened statistically at reported to construct the core of the taste of the boiled p<0.05. The most simplified taste test solutions were ob snow crab meat. This composition was similar to those of tained finally and Table 3 indicates the compositions of the the lobsters. most simplified taste test solutions. The test solution for Table 4 indicates t-values of the sensory taste test scor the Japanese spiny lobster taste consisted of 12 compo ing the difference in taste intensity between the 12-compo nents, especially a large amount of glycine was present. nent test solution representing Japanese spiny lobster and Sarcosine, valine, methionine, isoleucine, and leucine were the 16-component test solution representing shovel-nosed absent in this 12-component test solution. On the other lobster. The 16-component test solution was judged hand, the test solution for the shovel-nosed lobster taste statistically at p < 0.01 to have stronger sweetness, bitter consisted of 16 components. As compared to the 12-com ness, and taste body. ponent test solution, this 16-component test solution is Glycine is well known to have sweetness. The 16-compo characterized by the presence of valine, methionine, isoleu nent test solution possessed stronger sweetness than the cine, and leucine and also large amounts of glycine be 12-component test solution though less glycine quantity. taine, trimethylamine oxide, and arginine. The 12-compo Glycine betaine is also known to have sweetness. A large nent test solution showed clearly the lobster taste having amount of glycine betaine probably contributed to stron strong sweetness. This test solution also had umami, salti ger sweetness of the 16-component test solution. Fuke et ness, bitterness, and weak sourness. Moreover, the 16-com al.11) investigated the enhancing effect of AMP on prawn ponent test solution possessed much thicker taste body taste of a test solution prepared with commercial chemi than the 12-component test solution. Among taste-active cals and reported that the umami was detected in the taste Taste Components of Lobsters 287 test solution at the AMP concentration of 100 mg/100ml. tion and purification. Can. J. Biochem. Physiol., 37, 911-917 The synergistic elevation of AMP caused a further increase (1959). in umami, sweetness, and saltiness. Since the 16-compo 4) J. J. Bystedt, L. Swenne, and H. W. Aas: Determination of nent test solution lacked IMP, its umami was considered trimethylamine oxide in fish muscle. J. Sci. Food Agric., 10, 301 304 (1959). to be caused mainly by AMP. More than 500mg/ 100g of 5) Japan Soc. Anal. Chem. (Hokkaido): Chloride Ion (2), in "Analy arginine that is known to possess bitterness and whose sis of Water", Kagakudojin, Kyoto, 1979, pp. 206-208. (in threshold concentration was reported to be 50mg/100 Japanese) ml12)was added to 100ml of both of the 12 and 16-compo 6) A. Kawamura: Determination of phosphorus, in "Shin-pan Shoku nent test solutions. Bitterness of the 16-component test hingaku-Jikkenhou", Asakura, Tokyo, 1975, pp. 56-58 (in solution was stronger than that of the 12-component test Japanese). solution, however, the former test solution was still accept 7) M. Matsumoto, H. Yamanaka, and K. Hatae:. Effect of "arai" treatments on the biochemical changes in the kuruma prawn mus able and held well the lobster taste. Michikawa and cle. Nippon Suisan Gakkaishi, 57, 1383-1387 (1991) (in Japanese). Konosu13)1 reported that glutamic acid, AMP, and NaC1, es 8) T. Hirano, M. Yamaguchi, T. Shirai, T. Suzuki, and M. Suyama: pecially NaCl, reduced the bitterness of arginine at a con Free amino acids, trimethylamine oxide, and betaines of the raw centration of 300mg of arginine/100ml. and boiled meats of mantis shrimp Oratosquilla oratorio. Nippon Several subjects stated that the 16-component test solu Suisan Gakkaishi, 58, 973 (1992). tion had definitely the shovel-nosed lobster taste with 9) S. Konosu, K. Yamaguchi, and T. Hayashi: Studies on flavor com strong taste body and that its taste was similar to the crab ponents in boiled crabs. 1. Amino acids and related compounds in the extracts. Nippon Suisan Gakkaishi, 44, 505-510 (1978). taste. 10) 1. K. Suwetja, K. Hori, K. Miyazawa, and K. Ito: Changes in con tent of ATP-related compounds, homarine, and trigonelline in ma Acknowledgments We wish to express our sincere thanks to Dr. K. rine invertebrates during ice storage. Nippon Suisan Gakkaishi, 55, Sakuramoto of Tokyo University of Fisheries for his help in performing 559-566 (1989). t-test. 11) S. Fuke, K. Watanabe, and S. Konosu: Enhancing effect of nucleo tides on sweetness of heated prawn muscle, in "Olfaction and Taste References XI." (ed. by K. Kurihara, N. Suzuki, and H. Ogawa), Springer-Ver lag, Tokyo, 1994, pp. 357-360. 12) M. Yoshida, T. Ninomiya, S. Ikeda, S. Yamaguchi, T. Yoshikawa, 1) S. Konosu and K. Yamaguchi: The flavor components in fish and and M. Ohara: Studies on the taste of amino acids. Part I. Determi shellfish, in "Chemistry & Biochemistry of Marine Food Products" nation of threshold values of various amino acids. Nippon Nogei (ed. by R. E. Martin, G. J. Flick, C. E. Hebard, and D. R. Ward), Kagaku Kaishi, 40, 295-299 (1966) (in Japanese). AVI Publ., Westport, CT, 1982, pp. 367-404. 13) K. Michikawa and S. Konosu: Sensory identification of effective 2) T. Hayashi, K. Yamaguchi, and S. Konosu: Sensory analysis of components for masking bitterness of arginine in synthetic extract taste-active components in the extract of boiled snow crab meat. J. of scallop, in "Olfaction and Taste XI" (ed. by K. Kurihara, N. Food Sci., 46, 479-483 & 493 (1981). Suzuki, and H. Ogawa), Springer-Verlag, Tokyo, 1994, p. 278. 3) E. G. Bligh and W. J. Dyer: A rapid method of total lipid extrac