Fisheries Science 63(5), 772-778 (1997)

Taste Components of Boreo Pacific Gonate borealis

Takaaki Shirai, *1 Naoki Kikuchi, *1 Shingo Matsuo, *1 Shinichirou Uchida,*1 Hiroshi Inada,*2 Takeshi Suzuki,*1 and Toshiyuki Hirano*1,3

*1Department of Food Science and Technology , Tokyo University of Fisheries, Konan, Minato, Tokyo 108, Japan *2Laboratory of Fishing Technology , Tokyo University of Fisheries, Konan, Minato, Tokyo 108, Japan *3School of Marine Science and Technology , Tokai University, Orido, Shimizu, Shizuoka 424, Japan (Received September 20, 1996)

An extract was prepared with trichloroacetic acid from the and arm muscles of boreo pacific gonate squid. Free amino acids, trimethylamine oxide, 5'-ATP and related compounds, be taines, octopine, and other components in the extract were analyzed. Mantle muscle of boreo pacific gonate squid was rich in taurine, arginine, trimethylamine oxide, octopine, homarine, and glycine betaine. Amounts of free amino acids of the mantle and arm muscles of boreo pacific gonate squid were relatively lower than the reported values of common . This squid seemed to be acceptable as a food material. Amounts of acidic and neutral amino acids, Na+, and Cl of the arm muscle were higher than those of the mantle muscle, on the other hand, amounts of homarine and octopine of the mantle muscle were higher than those of the arm muscle. An ethanol extract was also prepared to reveal taste-active components of the boreo pacific gonate squid muscle. Threonine, serine, glutamic acid, gly cine, alanine, arginine, proline, glycinebetaine, trimethylamine oxide, trimethylamine, 5'-adenylic acid, NaCl, KH2PO4, and KC1 were confirmed as the taste-active components of boreo pacific gonate squid by the sensory taste test.

Key words: boreo pacific gonate squid, amino acid, extractive component, taste test

Squid is one of the major marine food resources and pos Yamaguchi5) and the squid muscle was reported to be rich sibly the most important of those which are not yet fully ex in taurine, proline, glycine, alanine, arginine, trimethyla ploited, occur in all oceans and seas. Boreo pacific gonate mine oxide. squid Gonatopsis borealis belongs to the Pacific, is jigged To reveal the extractive components of boreo pacific at 100-300 m depth, and is recently getting attention as a gonate squid and to evaluate its taste-active components, utilizable species in the fish market, because this squid is in we analyzed the extractive components and then a sensory cidentally caught frequently on the developing way of taste test was carried out with the taste test solutions. Ommastrephes bartrami jigging as a sub These were prepared with commercial chemicals and then stitutive fisheries for drift net in the central North Pacific data was obtained. Ocean." The taste of boreo pacific gonate squid is some Two kinds of extracts were prepared with trichloroace times said to be tasteless or thin and does not appear on tic acid and ethanol and were used in this study. The the market. Neither work on the taste of this squid nor the trichloroacetic acid extraction was employed from the utilization has been reported yet. Like common squids, biochemical point of view. Since the fresh squid flesh is boreo pacific gonate squid has triangular fins at the rear. often eaten as "sashimi" dishes which are raw, an extract At the front is the head and projecting from this are only that was prepared by the ethanol extraction reproduced an eight arms, and two long tentacles are lacked. Because of original lobster taste well enough.) The ethanol extraction the introduction of machinery catch into the squid fishery, was adopted to reveal the taste of raw squid taste from the this boreo pacific gonate squid is now becoming popular in food chemical view point. the seafood markets. Diacyl glycerol ether in liver lipids) and wax esters in the stomach content lipids') of boreo Materials and Methods pacific gonate squid were analyzed, however, little infor mation from the food chemical view point has been report Squid ed yet. Five individuals of boreo pacific gonate squid, Extractive components of squids were reviewed by Suya 406 •} 65.0 g body weight and 23 •} 1.3 cm mantle length, ma.4) Free amino acids, betaines, and trimethylamine which were jigged in the North Pacific Ocean at about oxide of squids were also reviewed by Konosu and 150 m depth in June-August 1995 and they were immedi- Taste Components of Boreo Pacific Gonate Squid 773 ately frozen at -35•Ž and transported to our laboratory were scored as 2 "strong," 1 "slightly strong," 0 "the in the frozen condition under -18•Ž. These squids were same," -1 "slightly weak," and -2 "weak." The differ thawed at 5•Ž for 5-6 h. A mantle muscle and 8 arms were ence in the taste test solutions was statistically judged by t-t removed from a squid, minced, and used as samples. est.

Analysis of Proximate Composition Results and Discussion Sample muscle was dried at 110•Ž and its moisture was measured. Crude protein was determined by the Kjeldahl Proximate Composition method. Lipid was extracted with chloroform/methanol Table 1 shows moisture, crude protein, lipid, and ash of mixture of Bligh and Dyer7) and dried in an air-oven. Ash the mantle and arm muscles of boreo pacific gonate squid. was measured with an oven at 550•Ž. Values are means of 5 individuals with SD. Moisture, crude protein, and ash of mantle muscles of flying squids, Preparation of Extracts cuttlefish, and common squids were respectively ranging For the trichloroacetic acid extraction, 2 g of the mantle from 76-81, 18-23, and 1-2%10) and these values were or arm muscle was homogenized with 50 ml of 5% almost the same as values of the mantle muscle of boreo trichloroacetic acid in a glass pestle homogenizer accord pacific gonate squid. There is little information about prox ing to the usual method.8) imate composition of the squid arm muscle. An ethanol extract was prepared with 2 g of sample ac cording to the reported method.6) Extractive Components Table 2 shows the composition of free amino acids, Analysis of Extractive Components trimethylamine oxide, ATP and related compounds, be Free amino acids, 5'-adenosine triphosphate (ATP) and taines, and the other extractive components of the related compounds, trimethylamine and its oxide, and trichloroacetic acid extracts of the mantle and arm muscles cyclic and w-betaines were measured with the previously of boreo pacific gonate squid. reported methods.) Octopine was estimated by the Among free amino acids, both of the mantle and arm Sakaguchi reaction9) after passing through a weak acidic muscles were rich in taurine and arginine amounting about ion-exchange Amberlite IRC-50 column.10) Na+, K+, Cl-, to 204 and 455 mg/ 100 g for taurine and 140 mg/ 100 g for and PO43- were also determined by the reported methods. 6) arginine. Alanine, glutamine, proline, lysine, and histidine Extractive nitrogen of the above extracts was deter were also detected in higher amounts being in levels of 55 mined by the Kjeldahl method. 70, 45, 30-55, 10-35, and 10-35 mg/ 100 g, respectively. A portion of the extract was dried up at 110•Ž and the The amount of taurine in the arm muscle was twice as residue was designated as a dry matter. much higher than that of the mantle muscle. Amounts of taurine, glycine, arginine, and proline of Measurement of pH the boreo pacific gonate squid mantle muscle were lower Water (10 ml) was added to 5 g of the mantle muscle than those of flying squid, cuttlefish, and common squids. and homogenized with an Ultra-Turrax (Janke-Kunkel) Especially the amounts of glycine and proline were about and the pH of the homogenate was measured with a pH 30 and 55 mg/ 100 g for the boreo pacific gonate squid meter. The pH of the ethanol extract was also measured. mantle muscle and were lower than those of the flying squid, cuttlefish, and common squid mantle muscles being Sensory Taste Test about 100-300 and 400-900 mg/ 100 g,11)respectively. The Eight subjects whose ages were from 20s to 40s were sums of free amino acids of the mantle and arm muscles of participated in the sensory taste test and most of the tests boreo pacific gonate squid amounting 770 and 1,100 mg/ were repeated 20 times with these subjects. An aqueous 100 g, respectively, were lower than those being 1,900 taste test solution was prepared with commercial chemi 2,400 mg/ 100 g of the mantle muscles of those flying cals. Before practicing this sensory taste test, the subjects squid, cuttlefish, and common squids. Comparing with the were trained in sweetness, umami, saltiness, sourness, and reported data obtained from several squids," the mantle astringency using glycine, alanine, and glycinebetaine, and arm muscles of boreo pacific gonate squid seemed not monosodium glutamate and 5'-adenylic acid (AMP), to be rich in free amino acids. NaCl, and KH2PO4 and KCI, respectively. The stimuli Ammonia was detected in the chromatograms of free were dissolved in distilled and deionized water and present ed to subjects at room temperature, 22-25•Ž. Subjects were given a row of two 20 ml glass cups and were asked to identify tastes. They were also trained to taste body and to Table 1. Proximate composition of the mantle and arm muscles of taste the profile of squid with a mixture of chemicals, the boreo pacific gonate squid Gonatopsis borealis above ethanol extract, and the minced squid mantle mus cle for description of each taste and its profile. Each senso ry taste test was carried out and repeated 20 times. Remov ing chemicals from the test solution until the initial taste profile of this solution was weakened or changed to carry out the omission test. The intensities of sweetness, umami, saltiness, sourness, and astringency and taste profile of the taste test solution Means of 5•}S.D. 774 Shirai et al.

Table 2. Trichloroacetic acid-extractable components of the man be fluctuated according to death and storage conditions. tle and arm muscles of boreo pacific gonate squid Gonatopsis ATP is degraded enzymatically in muscle, however, large borealis amounts of ATP amounting to 104 and 65 mg/ 100 g were detected in the mantle and arm muscles, respectively, of boreo pacific gonate squid in this study and suggested that sample squids were fresh. AMP amounting to 30 and 37 mg/ 100 g and hypoxanthine, 79 and 46 mg/ 100 g, were de tected in the mantle and arm muscles, respectively, however, 5'-inosinic acid (IMP) was absent in these mus cles in this study. Suwetja et al. 13)analyzed ATP related compounds in marine invertebrates during ice storage and more than 20 mg/ 100 g each of hypoxanthine was detect ed in the fresh muscles of cuttlefish Sepia esculenta and squid Loligo japonica at the 0 day storage. Relatively large amounts of glycinebetaine and homa rine amounting to about 460 and 310 mg/ 100 g, was detect ed in the mantle muscle, respectively. These values are almost the same level as those of the mantle muscles of flying squid, cuttlefish, common squid, and other squids.4) On the other hand, boreo pacific gonate squid was rich in octopine that are a guanidino compound and these amounts, being 1,800 and 930 mg/ 100 g for the mantle and arm muscles, respectively, were slightly higher than levels as amounts being 450-1,130 mg/ 100 g of the mantle muscles of several kinds of squids.4) The mantle muscle of boreo pacific gonate squid was rich in Na+ and Cl amounting to higher than 200 and 500 mg/ 100 g, respectively. These two components of the arm muscle were twice as high as those of the mantle mus cle. On the other hand, PO43- of the arm muscle detected 300 mg/ 100 g was about a half of that of the mantle mus cle. The inorganic ions of the mantle muscles of common and American jumbo squids") were reported to be 215 and 280-360 for Na+, 520 and 290-620 for K+, 240 and 320 800 for Cl-, and 715 and 380-740 mg/ 100 g for PO43-, re spectively. Na+ and Cl of the mantle muscles of Europe an cuttlefish S. officinalis and European arrow squid L. for besi were reported to be 50 and 129 mg/ 100 g for Europe an cuttlefish and 60 and 116 mg/ 100 g for European arrow squid, respectively.14) Amounts of acidic and neutral amino acids, Na+, and Cl of the arm muscle of boreo pacific gonate squid were higher than those of the mantle muscle, on the other hand, amounts of homarine and octopine of the mantle muscle were higher than those of the arm muscle. Table 3 shows ethanol-extractable components of the mantle and arm muscles of boreo pacific gonate squid. Since the extract was prepared with 80% ethanol had the Values are means of 5 samples with S.D. original taste of the raw lobster muscle as well,') the * Significantly higher at p<0 .05 between the mantle and arm muscles. ethanol extract was prepared in this study from the food chemical view point to reveal the taste and taste-active com ponents of the raw mantle and arm muscles of boreo amino acid analysis and their amounts of mantle and arm pacific gonate squid. Each amount of the ethanol-extracta muscles were less than 30 mg/ 100 g for both of the mantle ble components in Table 3 was slightly lower than the and arm muscles. About 35, 85, and 240 mg/ 100 g of am those of the trichloroacetic acid-extractable components in monia were detected in the raw mantle muscle of Ameri Table 2, however, almost no difference between both the can jumbo squids which were caught at three different overall compositions of ethanol-extractable and trichloro areas. 12) acetic acid-extractable components was seen and they were Like other squids,5) a large amount of trimethylamine similar to each other. oxide was detected. The mantle muscle was richer than the Amounts of the dry matter of the ethanol extracts pre arm muscle in trimethylamine oxide. pared with the mantle and arm muscles were about 5,400 Amounts of ATP and related compounds are known to and 5,500 mg/ 100 g, respectively. The total amounts of ex- Taste Components of Boreo Pacific Gonate Squid 775

Table 3. Ethanol extractable-components of the mantle and arm tively. muscles of boreo pacific gonate squid Gonatopsis borealis Sensory Taste Test First, 8 subjects tasted the ethanol extract and their im pressions were collected. All subjects stated that the ethanol extract had a slight sweetness, sourness, and astrin gency, stronger umami and saltiness, and a thick taste body. This extract had the taste profile of the boreo pacific gonate squid muscle very well. Then in order to carry out the sensory taste test between the test solution and the squid muscle, the taste test solu tion shown in Table 4 was prepared with 23 commercial chemicals on the basis of the composition of ethanol-ex tractable components of the mantle muscle of boreo pacific gonate squid. Aspartic acid, citllurine, cystathio nine, cystine, ornithine, and lysine amounting to less than 10 mg/ 100 g were not added to this test solution. Amounts of ATP and related compounds analyzed and ap peared in the Table 3 were regarded as AMP in the taste test solution. Ammonia, homarine, picolic acid, and octo pine were excluded from the test solution. To compare the taste of this test solution with another squid, the test solu tion simulating the neon flying squid taste was also pre pared on the basis from the reported values.) In this neon flying squid taste test solution, amounts of ATP and relat ed compounds and inorganic components of the mantle muscle of boreo pacific gonate squid were adopted, since these amounts were absent there. The pH of the mantle muscle was 6.6. The pH of the

Table 4. Taste test solutions for the mantle muscles of boreo pacific gonate and neon flying squids

Values are means of 5 samples with S.D. * Significantly higher at p<0.05 between the mantle and arm muscles.

tractive components shown in Table 3 divided by the dry matter and was designated as the dry matter recovery and wasalso shown in Table 3. These recoveries were 96% for the mantle muscle and 102% for the arm muscle, showing that almost all components in the extract were analyzed in thisstudy . The extractive nitrogen recovery was estimated by dividing the sum of the nitrogen amounts calculated fromthe analyzed nitrogenous components by the amount of extractive nitrogen. These extractive nitrogen recoveries for the mantle and arm muscles of boreo pacific gonate Giutamic acid was used as sodium glutamate, trimethylamine oxide as squid were also satisfactory, being 80 and 98%, respec- TMAO.2H2O, trimethylamine as TMA-HCl, and AMP as AMP -Na2 6H2O. 776 Shirai et al. ethanol extract was also measured and it was 6.8. Then the pacific gonate squid taste test solution was judged to have pH of taste test solution was adjusted to 6.8 for the boreo the taste of boreo pacific gonate squid. From the good pacific gonate squid taste test solution and 7.0 for neon recoveries of the dry matter and extractive nitrogen (Table flying squid taste test solution with a few drops of diluted 3) and the taste similarity between the minced mantle mus NaOH or HCI solution. cle and ethanol extract (Table 5), almost all extractive com This boreo pacific gonate squid taste test solution was in ponents, especially taste-active components, of the boreo itially sensory evaluated with the minced mantle muscle of pacific gonate squid were judged to be analyzed in this boreo pacific gonate squid. Table 5 shows the results of t study. test in which the boreo pacific gonate squid taste test solu An attempt was made to simplify the test solution con tion were scored against the mantle muscle of boreo pacific sisting of 23 chemicals, another omission test was carried gonate squid. The taste of the test solution appeared imme out. Five basic tastes and taste body between the test solu diately in a mouth since the solution was aqueous liquid so tion consisting of 23 chemicals and the test solution lack that saltiness of this taste test solution was judged to be ing one or several components from the above solution. stronger than the minced muscle. Significant (p<0.05) The results were shown in Table 6. Removing valine, difference in saltiness was obtained, however, umami of methionine, isoleucine, leucine, tyrosine, phenylalanine, the taste test solution and the muscle and their taste glutamine, histidine, and taurine from the test solution profiles were definitely similar to each other. The boreo simulating the Gonatopsis boreolis taste was scored not to be weakened or changed the taste of the original test solu tion at p<0.05. The omission of NaCl led to a decrease in saltiness, sweetness, and taste body. Michikawa and

Table 5. Sensory taste test and t-values for the taste intensity of the Konosu15) reported that 0.071% of NaCl decreased the test solution consisted of 23 chemicals in Table 4 for the boreo strength of bitterness due to 323 mg of arginine in 100 m/ pacific gonate squid mantle muscle against the minced mantle in the synthetic extract of scallop. The omission of KCI muscle of boreo pacific gonate squid from the test solution decreased saltiness and astringency. Removing AMP and trimethylamine oxide decreased uma mi and sweetness, respectively. Fuke et a!. 16)reported that the sweetness was increased by adding 50 mg of AMP to 100 ml of the synthetic extract simulating the taste of the kuruma prawn meat and AMP imparted umami when its level was elevated to more than 100 mg. The omission of serine and trimethylamine oxide decreased sweetness and of glutamic acid, glycine, alanine, and proline weakened sweetness and umami. Ugawa et al. 17)reported that the n=20. * The taste of the test solution was significantly (p<0.05) different from the sweetness of 100 mM sweet amino acids such as glycine, minced muscle. alanine, and serine was enhanced by 30 mM NaCl. Concen

Table 6. Sensory taste test and t-values for the simplified test solution omitting stimuli or stimulus against the test solution consisting of commercial chemicals simulating the boreo pacific gonate squid taste

n=20. The taste test solution consisted of valine, isoleucine, leucine, tyrosine, phenylalanine, glutamine, histidine, taurine, threonine, serine, glutamic acid-Na, glycine,ala nine, arginine, proline, NaCl, KCI, KH2,PO4,AMP •Na2-6HZO,glycinebetaine, trimethylamine oxide.2H2O, trimethylamine•HCl and its composition was appeared in Ta ble 4. * Tested solution was significantly strong or weak (-) at p<5% against the test solution consisting of 23 chemicals simulating the Gonatopsis borealis taste. Taste Components of Boreo Pacific Gonate Squid 777 trations of these three amino acids were lower than 100 Table 9. Sensory taste test and t-values for the squid taste with the mM in our test solution and the same effect was not taste test solution of boreo pacific gonate squid and that of neon confirmed. When NaCl, KCI, KH2PO4, AMP, glycine flying squid betaine, trimethylamine, threonine, serine, glutamic acid, glycine, alanine, arginine, and proline were omitted, the taste body of the test solution simulating the Gonatopsis borealis taste was weakened significantly. Then these 13 components and trimethylamine oxide were taste-active components for the Gonatopsis borealis taste. This omission test identified 14 components for taste-ac tive components for the boreo pacific gonate squid muscle. Table 7 shows 14 chemicals as taste-active components for the boreo pacific gonate squid muscle and Table 8 shows the results of t-test with this simplified taste test solution and the taste test solution consisted of 23 chemicals. No significant (p<0.05) difference in the taste and the squid n=20. * Taste test solution of boreo pacific gonate squid was significantly strong or taste profile was found between the taste test solution that weak (-) at p<0.05 against that of neon flying squid. consisted of 23 chemicals and that consisted of 14 chemi cals. Subjects judged that 2 mg of trimethylamine con tributed a good characteristic flavor of seafood in the squid taste and a large amount (about 1 g/ 100 ml) of trimethylamine oxide contributed the taste body. On the other hand, the test solution consisted of 23 Table 7. Simplified taste test solutions consisted of 14 chemicals chemicals for the boreo pacific gonate squid taste was com for the boreo pacific gonate squid and neon flying squid pared with the taste test solution for the neon flying squid taste which are listed in Table 4, since the glycine amount of the mantle muscle of boreo pacific gonate squid was rel atively lower than other squids.4) Moreover, the simplified two taste test solutions consisted of 14 taste-active compo nents those were confirmed in this study for the boreo pacificgonate squid were also compared to each other and Table 9 shows the results of t-test. Compared with the neon flying squid taste test solution, the overall taste of boreo pacific gonate squid mantle muscle has a weaker sweetness and stronger saltiness. These taste profiles seemed to be week because lower amounts of glycine, ala nine, and proline were existed in the mantle muscle of boreo pacific gonate squid. Weak sweetness of the boreo pacific gonate squid taste seemed to enhance stronger salti ness in the taste. Subjects pointed that boreo pacific gonate squid lacked sweetness, however, it had a stronger taste body. The components of off-flavor, such as very salty, sour, bitter, and fishy tastes, in the muscle of Ame Glutamic acid was used as sodium glutamate, trimethylamine oxide as TMA0 2H20, trimethylamine as TMA-HCI, and AMP as AMP•Na2.6H20. The rican jumbo squid was reported to be caused by a large composition of test solution for neon flying squid was referred to data of Suyama.4) amount (300-800 mg/ 100 g) of chloride ion existing as NH4C1 in the squid muscle.") The amounts of Cl and ammonia in the mantle and arm muscles were 584 and 960 mg/ 100 g and 27 and 24 mg/ 100 g, respectively. No Table 8. Sensory taste test of the boreo pacific gonate squid taste, subject stated the existence of off-flavor in the mantle taste intensity, and t-values with simplified taste test solution and arm muscles of boreo pacific gonate squid, extracts, consisted of 14 chemicals against the taste test solution consist and taste test solution in the sensory taste test. Squids ed of 23 chemicals such as oval squid, arrow squid, cattlefish, and common and neon flying squids were said to have a good taste in this order. 18)This squid had a characteristic taste and seemed to be acceptable as a food material.

Acknowledgments We are grateful to the Japan Marine Fishery Resources Research Center for collecting squid samples.

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