혈합육어(멸치, 고등어, 황다랭이 및 날개다랭이)의 Trypsin - 1. 정제와 반응조건

변재형(Jae-Hyeung Pyeun),조득문(Deuk-Moon Cho),허민수(Min-Soo Heu) 한국식품영양과학회 1993 한국식품영양과학회지 Vol.22 No.4

혈합육어 중 연근해 온대산 멸치의 내장과 고등어의 유문수, 그리고 원양 열대산 황다랭이 및 날개다랭이의 유문수를 각각 시료로 하여 trypsin을 분리 정제하였으며, 그 분자량과 아미노산 조성 및 효소의 반응 최적조건에 관하여 비교분석하였다. 실험결과를 요약하면 다음과 같다. 1. 멸치의 내장과 고등어, 황다랭이 및 날개다랭이의 유문수에서 각각 황산암모늄염석, benzamidine-Sepharose 6B 친화성 크로마토그래피, DEAE-Sephadex A-50 크로마토그래피, Sephadex G-75 겔 여과 크로마토그래피 등의 방법을 혼용하여 고등어부터는 2종의 trypsin(고등어 trypsin A와 고등어 trypsin B로 명명함)을, 그리고 다른 어류로부터는 각각 1종의 trypsin을 분리 정제하였다. 2. 멸치 trypsin과 고등어 trypsin B는 고등어 trypsin A와 황다랭이 trypsin 및 날개다랭이 trypsin에 비하여 그 고유활성이 월등히 높았다. 3. 이들 혈합육어 trypsin의 분자량은 22kDa~26kDa 범위였다. 4. 이들 trypsin은 공통적으로 glycine, serine, aspartic acid를 많이 함유하고 있었으며, tryptophan, methionine, lysine, tyrosine의 함유량은 적었다. 5. BA-p-NA기질에 대한 반응 최적조건은 멸치 trypsin은 PH 8.0에서 45℃, 고등어 trypsin A와 B는 PH 8.0에서 50℃, 그리고 황다랭이 trypsin은 PH 9.0에서 55℃, 날개다랭이 trypsin은 PH 9.0에서 50℃였다. 6. 위에 든 실험 결과들은 혈합육 어류의 서식온도가 이들 어류의 trypsin의 반응 최적 온도와는 어느정도 관계가 있을 것으로 추측되었다. Deterioration of fish muscle is known to occur more quickly in the dark fleshed fish than in the white fleshed fish, causing by their high intestinal proteolytic activity. Muscle degradation which suffer post-mortem autoproteolysis is affected by trypsin with its unique activation function towards other enzymes. To compare physicochemical and enzymatic properties for the trypsins of the dark fleshed fish, trypsins from the viscera of anchovy (Engraulis japonica), and the pyloric caeca of mackerel(Scomber japonicus), yellowfin tuna(Thunnus albacores) and albacore (Thunnus alalunga) were purified through ammonium sulfate fractionation, benzamidine-Sepharose 6B, DEAE-Sephadex A-50, and Sephadex G-75 chromatography. Two trypsins from mackerel (designated mackerel trypsin A and mackerel trypsin B), and one each from anchovy, yellowfin tuna and albacore were isolated as electrophoretical homogeneity. The purities of anchovy trypsin, mackerel trypsin A and H, yellowfin tuna trypsin, and albacore trypsin increased to 78.1, 4.8, 9.3, 120, and 16o-fold, respectively, compared to crude enzyme solutions. Molecular weights of the trypsins from the dark fleshed fish estimated by SDS-polyacrylamide electrophoresis were ranged from 22kDa to 26kDa. The trypsins contained higher amount of glycine, serine and aspartic acid, and less amount of tryptophan, methionine, lysine and tyrosine. Optimal conditions for amidolytic reactions of the enzymes were pH 8.0 and 45℃ for anchovy trypsin, pH 8.0 and 50℃ for mackerel trypsin A and B, pH 9.0 and 55℃ for yellowfin tuna trypsin, and pH 9.0 and 50℃ for albacore trypsin. It was supposed that the habitat temperature of the dark fleshed fish is slightly connected with the optimal reaction temperature of the trypsins of the fish.

혈합육어 Trypsin의 효소적 성질에 대한 반응속도론적 해석

조득문 ( Deuk Moon Cho ),허민수 ( Min Soo Heu ),김형락 ( Hyeung Rak Kim ),김두상 ( Doo Sang Kim ),변재형 ( Jae Hyeung Pyeun ) 한국수산학회 1996 한국수산과학회지 Vol.29 No.1

멸치, 고등어, 황다랭이 및 날개다랭이의 혈합육어에서 정제된 trypsin을 시료로 하여 각각의 BAPNA기질에 대한 반응속도와 그 관련 성질들을 분석 검토하였다. 4종의 혈합육어에서 정제된 trypsin의 Km`와 k(cat)는 멸치 trypsin이 각각 49.3μM과 90.9min 1, 고등어 trypsin A는 53.7μM과 61.2min 1 고등어 trypsin B는 96.5μM과 76.6min 1 황다랭이 trypsin은 62.8μM과 46.4min 1 그리고 날개다랭이 trypsin은 98.3μM과 47.68min 1이었다. TLCK에 대한 K1값은 멸치 trypsin이 20.90μM, 고등어 trypsin A가 2.86μM, 고등어 trypsin B가 3.90μM, 황다랭이 trypsin이 0.96μM, 그리고 날개다랭이 trypsin이 1.82μM이었으며, 황다랭이의 trypsin이 TLCK에 대하여 가장 예민하게 반응하였다. 이들 trypsin의 효소 활성과 촉매효율은 연근해 온 대산 혈합육어인 멸치와 고등어 trypsin이 열대해역에서 온대해역에 걸쳐 널리 회유하는 혈합육어인 황다랭이와 날개다랭이의 trypsin에 비하여 높은 특징을 보였다. Kinetic properties of trypsins purified from dark-fleshed fish (anchovy, mackerel, yellowfin tuna, and albacore) were examined and analyzed on benzoyl-(D,c)-arginine-p-nitroanilide (BAPNA). The values of Km` and k(cat) of the purified trypsins from the four dark-fleshed fish were found to be 49.3μM and 90.9min-1 for anchovy, 53.7μM and 61.2min-1 for mackerel A, 96.5μM and 76.6min-1 for mackerel B, 62.8μM and 46.6min-1 for yellowfin tuna, and 98.3μM and 47.7min-1 for albacore, respectively. The values of K, on tosyl-L-lysine chloromethyl ketone (TLCK) were determined to be 20.90μM for anchovy trypsin, 2.86μM for mackerel trypsin A, 3.90μM for mackerel trypsin B, 0.96μM for yellowfin tuna trypsin, and 1.82μM for albacore trypsin. Thus yellowfin tuna trypsin was the most sensitive to TLCK among all trypsins. The activities and catalytic efficiency of the trypsins purified from the temperate zone fish, anchovy and mackerel, were higher than those of the trypsins purified from yellowfin tuna and albacore which migrate widely from the tropic zone to the temperate zone.

혈합육어(멸치, 고등어, 황다랭이 및 날개다랭이)의 Trypsin - 2. 성질과 열 안정성

조득문(Deuk-Moon Cho),허민수(Min-Soo Heu),변재형(Jae-Hyeung Pyeun) 한국식품영양과학회 1993 한국식품영양과학회지 Vol.22 No.4

전보에 이어, 정제한 혈합육어 trypsin에 대하여 효소적 성질 및 열역학적 성질에 관하여 비교 검토한 내용을 요약하면 다음과 같다. 이들 혈합육어 trypsin은 BA-p-NA와 SP-p-NA 같은 amide기질 중 BA-p-BA기질만에 대하여, 그리고 ATEE, BAEE, BTEE 및 TAME 등의 ester기질 중에서는 BAEE와 TAME에 대하여만 현저한 활성을 보였다. 이들 효소는 antipain, leupeptin, DFT, TLCK, SBTI 등 화학약제와 금속이온 Cu^(2+) 및 Hg^(2+)에 의하여서는 그 활성이 현저히 저해를 받았으나, Mg^(2+)에 의하여서는 부활하였다. 이 효소들은 모두 50℃이상의 온도에서는 불안정하였으며, 날개다랭이의 trypsin이 온도 변화에 대하여 가장 안정하였다. 그리고, 활성화에너지는 멸치 trypsin이 13.91㎉/mole, 고등어 trypsin A는 11.61㎉/mole, 고등어 trypsin B는 8.43㎉/mole, 황다랭이 trypsin은 4.35㎉/mole, 그리고 날개다랭이 trypsin은 3.76㎉/mole이었다. In the present paper, enzymatic properties of the trypsins from the four dark fleshed fish were compared with each other and thermal stabilities of the enzymes were also investigated. The trypsins from the dark fleshed fish showed their activity only in BA-p-NA substrate of the amide substrates such as BA-p-NA and SP-p-NA, and BAEE and TAME of the ester substrates such as ATEE, BAEE, BTEE, and TAME. The enzymes were strongly inhibited by the serine protease inhibitors such as antipain, leupeptin, TlCK, DFP and SBTI, and were also inhibited by such metal ions as Cu^(2+) and Hg^(2+), but fairly activated by Mg^(2+). Denaturation constants of the enzymes were 13.4×10^(-4)sec^(-1) for anchovy trypsin, 47.18×10^(-4)sec^(-1) for mackerel trypsin A, 34.06×10^(-4)sec^(-1) for mackerel trypsin B, 42.28×10^(-4) sec^(-1) for yellowfin tuna trypsin and 16.6×10^(-4) sec^(-1) for albacore trypsin at 55°C. The activation energies of the trypsins at a temperature range of 30℃ to 50℃ were estimated to be 13.91㎉/mole for anchovy trypsin, 11.61㎉/mole and 8.43㎉/mole for mackerel trypsin A and for mackerel trypsin B, 4.35㎉/mole for yellowfin tuna trypsin, and 3.76㎉/mole for albacore trypsin.

식용해조류중의 미량요소와 특수기능성 당질 - 1 산지와 채취시기별 일반성분의 조성과 무기원소의 분포

조득문,김두상,이동수,김형락,변재형 ( Deuk Moon Cho,Doo Sang Kim,Dong Soo Lee,Hyeung Rak Kim,Jae Hyeung Pyeun ) 한국수산과학회 1995 한국수산과학회지 Vol.28 No.1

Nine species of edible seaweed [green layer(Monostroma nitidium) and sea staghorn(Codium fragile) of green algae; sea mustard(Undaria pinnatifida), seaweed fusiforme (Hizikia fusiforme), gulf weed (Sargassum fulvellium), and sea tangle(Laminaria japonica) of brown algae; seaweed dilatata(Halimeniopsis dilatata), seaweed furcata(Gloiopeltis furcata), and layer(Porphyra tenera) of red algae] collected from Kijang, Chungmu, and Yosu in Korea, were examined and compared on their chemical and mineral composition depending on their harvesting season. Crude protein(N×6.25) showed about 45% on moisture free basis(the contents of every components described below are shown as moisture free basis) for layer and 30% for green layer collected from every seasons and sites. Sea tangle showed the highest content in crude lipid(10%) among brown seaweeds and green layer had 6% of crude lipid regardless their habitats. Ash and carbohydrate in sea staghorn hold about 90% of total solids but those levels were only 50% in layer. The relationship between ash and carbohydrate content showed a tendency with reverse correlation. Mineral compositions were examined on green layer, sea mustard, and seaweed furcata collected from Kijang. Sodium was eminent element(1,798-7,334㎎/100g) followed by potassium. Magnesium and calcium content were low level compared with sodium and potassium. As a micro-element, iron was appraised the highest status(165-330㎎/100g) in green layer, however, iron in sea mustard and seaweed furcata was comparatively low amount (2.7-47.4㎎/100g). The level of zinc was also comparatively high and that was varied on habitats. In conclusion, chemical compositions of these algae were distinctively varied on species and habitats, and mineral compositions were notably changed by the harvesting season.

Quality Factors and Functional Components in the Edible Seaweeds - Ⅰ. Distribution of n-3 Fatty Acids in 10 Species of Seaweeds by Their Habitats

정보영(Bo-Young Jeong),조득문(Deuk-Moon Cho),문수경(Soo-Kyung Moon),변재형(Jae-Hyeung Pyeun) 한국식품영양과학회 1993 한국식품영양과학회지 Vol.22 No.5

10종 해조류(녹조류 2종, 갈조류 5종, 홍조류 3종)의 n-3 지방산 조성을 우리 나라 남해안의 주요 서식지에 따라 비교하였다. 총지질(TL)은 녹조류 중에서는 파래(약 7~10%), 갈조류 중에서는 미역과 톳 (약 3~5%), 그리고 홍조류 중에서는 김(약 5~6%)에서 각각 많았다. 미역과 톳의 TL함량은 충무에서 채취한 것이 기장 및 여수에서 채취된 것들보다 많은 경향이 있었으나, 파래의 경우에는 가장 및 여수에서 채취된 것에서 더 많았다. 녹조류는 16:4(n-3), 18:4(n-3) 및 18:3(n-3)과 같은 비교적 탄소수가 적온 n-3지방산(28.0~66.3%)이 풍부하였다. 갈조류는 녹조류에 비하여 적은 양의 n-3 지방산 (17.9~36.5%)을 함유하였으나, n-6 지방산을 상당량 함유하였다. 홍조류의 n-3 지방산은 대부분 20:5(n-3) 지방산으로 구성되었으며, 특히 낙동에서 채취된 것이 53%의 20:5(n-3)를 함유하였다. 이들 n-3 지방산의 함량은, 동일종에 속하는 해조의 경우, TL함량이 높은 종류에서 더 높은 경향을 보였다. 따라서 시험된 해조류는 TL함량과 n-3 지방산의 함량이 서식지 따라 따른 결과를 나타내었다. Total lipid contents from 10 species of marine algae(2 green, 5 brown, and 3 red algae) collected from different places in Korea were examined and their fatty acid compositions were compared among species and habitats. Total lipid (TL) was prominent in green laver (about 7.3~10.1%) of the green algae, in sea mustard and seaweed fusiforme(about 3.1~4.8%) of the brown algae and purple laver (about 4.9~6.4%) of the red algae. Sea mustard and seaweed fusiforme collected at Chungmu contained a relatively high level of TL compared with those collected at Kijang and Yosu, whereas green laver at Chungmu revealed a lower level of TL than that at Yosu and Kijang. The TL content of purple laver showed the highest portion in that collected at Nakdong. Green algae comprised the majority of n-3 fatty acids (29.0~66.3%), which mainly consisted of 16:4 (n-3)(or 16:3 (n-3)), 18:3 (n-3) and 18:4(n-3). Brown algae accounted for a low level of n-3 fatty acids (17.9~36.5%) mainly 18:4(n-3), 18:3(n-3) and 20:5(n-3), whereas the brown algae contained a significant level of n-6 fatty acids (7.23~26.5%) such as 20:4 (n-6) and 18:2 (n-6). In the case of red algae, the n-3 fatty acids consisted mostly of 20:5 (n-3) which scored 53% of polyenoic acids in purple laver collected at Nakdong. The proportion of n-3 fatty acids in algae belonging to the same species was higher in algae of high TL contents. Consequently, TL and n-3 fatty acid levels from the seaweeds studied in this paper were different from their habitats.

해삼內臟젓갈 熟成中 蛋白質分解酵素의 活性과 아미노酸 組成의 變化

이기찬(Gi Chan Lee),조득문(Deuk Moon Cho),변대석(Dae Seok Byun),주현규(Hyen Kyu Joo),변재형(Jae Hyeung Pyeun) 한국식품영양과학회 1983 한국식품영양과학회지 Vol.12 No.4

水産名産品에 屬하는 해삼內臟젓갈의 食品營養學的인 評價 및 그 加工에 關한 基礎資料를 얻고자 본 연구를 착수하였으며, 해삼內臟 組織中에 分布하는 蛋白質分解 粗酵素를 抽出하여 그 活性條件과 젓갈 熟成中의 遊離아미노酸 및 蛋白質 構成 아미노酸의 組成變化에 關하여 分析 檢討한 結果를 要約하면 다음과 같다.<br/> 1. 해삼內臟 組織中에는 pH 3.1 50℃, pH 5.7 50℃ 그리고 pH 7.7 45℃에 各各 最適活性條件을 갖는 세 種類 以上의 蛋白質分解酵素의 存在가 確認되었다.<br/> 2. 이들 酵素中 pH 3.1 50℃, pH 7.7 45℃에 最適活性을 갖는 酵素들은 食鹽濃度 1%以上의 濃度에서 漸次 높은 阻害를 받았으며, pH 5.7 50℃에 最適活性條件을 갖는 酵素는 食鹽濃度 1%에서 賦活되었으며, 5%以上에서 阻害를 받았다.<br/> 3. 몇가지 鹽類이온에 의한 影響을 檢討한 結果 pH 3.1 50℃에서 最適條件을 보인 酵素는 實驗에 쓴 全 鹽類이온에 의하여 조금씩 阻害를 받았으며, pH 5.7 50℃에서 最適條件을 보인 酵素는 Cu^(2+)을 除外한 모든 鹽類이온에 의하여 賦活을 받았고, pH 7.7 45℃에서 最適條件을 보인 酵素는 Ca^(2+)과 Mn^(2+)에 의하여서는 賦活을 받았으며, Ba^(2+)에 의하여서는 影響이 없었고 Co^(2+), Mg^(2+)에 의하여서는 阻害를 받았다. 그리고 特히 이 酵素들은 Cu^(2+)에 의하여서는 甚한 阻害를 받았다.<br/> 4. 해삼內臟 組織에 分布하는 酵素들의 젓갈熟成日數의 經過에 따른 活性에 미치는 影響을 檢討한 結果, pH 3.1 50℃와 pH 7.7 45℃에서 活性最適條件을 보인 酵素들은 活性이 조금씩 弱化하여 갔으나, pH 5.7에서 活性最適條件을 보인 酵素는 熟成日數의 經過에도 影響이 없었다.<br/> 5. 해삼內臟젓갈 熟成 8日째일 때는 生 內臟中의 蛋白質 構成아미노酸의 大部分이 減少하였으며, 特히 두드러지게 減少한 아미노酸은 arginine, alanine, glutamic acid, glycine, serine, valine, threonine 및 lysine 等이었고, methionine와 histidine 및 isoleucine 等은 그 減少幅이 낮았다.<br/> 6. 遊離아미노酸 組成을 分析한 結果, 8日間 熟成한 試料中에는 glutamic acid, aspartic acid, leuine 및 lysine은 많은 量이 含有되어 있었고, histidine, methionine, proline 및 tyrosine 等은 그 含量이 낮았다. 그리고 젓갈 熟成過程中에는 大部分의 遊離아미노酸은 增加하였으며, 特히 lysine, histidine, threonine, glutamic acid, methionine, valine, leucine 等은 많은 增加를 보였다. This study was undertaken to ascertain food and nutritional evaluating data on the processing of fermented sea cucumber (Stichopus japonicus) entrails.<br/> In the experiment, the crude proteolytic enzyme from the entrails tissue of raw and fermented sea cucumber during the days of ripening was extracted. The optimal activity condition for the crude enzyme and the compositional changes of amino acid of the protein and free amino acid in the raw and fermented sample were also investigated.<br/> 1. Less than three kinds of proteolytic enzymes that each enzyme has optimal activity condition at pH 3.1 50℃(A-enzyme), pH 5.7 50℃(B-enzyme) and pH 7.7 45℃(C-enzyme), respectively were believed to be exist in the entrails tissue of sea cucumber.<br/> 2. A-enzyme and C-enzyme were strongly inhibited with the increase of the salt concentration, and B-enzyme was activated at the 1% salt concentration and was inhibited above the 5% salt concentration.<br/> 3. The result of the effect of several salt ions on the proteolytic activity showed that A-enzyme was slightly inhibited in the presence of all salt ions added, B-enzyme was activated in the presence of the all salt ions except Cu^(2+) and C-enzyme was activated in the presence of Ca^(2+) and Mn^(2+), and inhibited by Cu^(2+), Co^(2+) and Mg^(2+).<br/> 4. When the effects of the ripening days on the proteolytic activity of the crude enzymes were analysed, the activity of the A-enzyme was slightly weakened with the lapse of the fermentation days, whereas the B-enzyme was not influenced by the fermentation days.<br/> 5. In the analysis of amino acid composition of the protein of the samples, the 8 days fermented sea cucumber entrails showed the diminution of all kinds of amino acid. Apparently diminished amino acids were arginine, alanine, glutamic acid, glycine, serine, valine, threonine and lysine etc., and methionine, histidine and isoleucine were slightly decreased.<br/> 6. In the analysis of free amino acid composition of the 8 days fermented sample, glutamic acid, aspartic acid, leucine and lysine were rich, while histidine, methionine, proline and tyrosine were poor. The most of free amino acids were increased during the fermentation procedure and especially in lysine, histidine, threonine, glutamic acid, methionine, valine and leucine.

식용 갈조류의 알긴산 조성

이동수(Dong Soo Lee),김형락(Hyeung Rak Kim),조득문(Deuk Moon Cho),남택정(Taek Jeong Nam),변재형(Jae Hyeung Pyeun) 한국수산과학회 1998 한국수산과학회지 Vol.31 No.1

Uronate compositions and molecular weights of alginates from the four kinds of brown algae, sea mustard(Undaria pinnatifida), sea tangle(Laminaria japonica), gulf weed(Sargassum fulvellum), and seaweed fusiforme(Hizikia fusiforme), in regard with the harvesting season were investigated. Sea mustard contained the highest amount of alginates in the four kinds of brown algae. _D-Mannuronic acid to _L-guluronic acid(M/G) ratio of the alginates was high in order of seaweed fusiforme, gulf weed, sea mustard, and sea tangle, and especially in water-soluble alginate. Molecular weights of the alginates were greater with the growing period ranging in 4,500-4,800kDa for sea tangle, 4,000-4,200kDa for sea mustard, 3,300-3,400kDa for seaweed fusiforme, and 3,000-3,200kDa for gulfweed. In water-soluble alginate of sea mustard, M/G ratio was much higher in sporophyll than in midrib and blade.

식용해조류중의 미량요소와 특수기능성 당질: 2. 산지와 채취시기별 식이성섬유질 함량의 변화와 해조다당류의 분포

김두상 ( Doo Sang Kim ),이동수 ( Dong Soo Lee ),조득문 ( Deuk Moon Cho ),김형락 ( Hyeung Rak Kim ),변재형 ( Jae Hyeung Pyeun ) 한국수산학회 1995 한국수산과학회지 Vol.28 No.3

우리 나라 대표적인 식용해조 (녹조류-참홑파래, 청각; 갈조류-미역, 톳, 모자반, 다시마; 홍조류-얼룩도박, 참김, 불등풀가사리) 9종을 대상으로 가용성 식이섬유질, 불용성 식이섬유질 및 총 식이섬유질의 함량을 주요 산지 (기장, 충무, 여수)와 채취 시기별로 측정·검토하고, 특히 갈조류와 홍조류에 대하여는 해조 다당류의 분포를 분석 비교하였다. 총 식이섬유질은 녹조류 중의 참홑파래가 25.4~38.1% (이하 건물중의 비율), 청각은 35.4~43.8%를, 갈조류 중의 미역은 34.2~48.8%, 톳은 37.5~47.8%, 모자반은 42.9~71.3%, 다시마는 37.1~45.1%를, 그리고, 홍조류 중의 참김은 31.3~40.5%, 얼룩도박은 51.5~60.4%, 불등풀가사리는 57.1~65.8%를 각각 함유하였다. 가용성 식이섬유질과 불용성 식이섬유질을 공통적으로 많이 함유한 해조류는 각각 불등풀가사리와 모자반이었다. 총 식이섬유질에 대한 가용성 식이섬유질의 함유 비율은 녹조류 중에서는 참홑파래 (43.7~64.8%), 갈조류 중에서는 미역 (17.5~31.3%), 그리고 홍조류 중에서는 불등풀가사리 (44.7~63.2%)가 높은 값을 보였다. 해조 다당류의 분포상의 특정을 보면, 갈조류는 알칼리 가용성 알긴산을 가장 많이 함유하였으며 (9.0~15.1%), 홍조류 중의 참김은 폴피란 (5.8%)을, 그리고 불등풀가사리와 얼룩도박은 한천 (20.0%)과 카라기난 (23.8%)을 각각 많이 함유하는 차이를 보였다. This report described on the soluble, insoluble, and total dietary fiber level depending on the harvesting seasons and habitats in 9 species of marine algae. Total dietary fiber contents were comprised 25.4~38.1% (dry basis) in green laver and 35.4~43.8% in sea staghorn of green algae, 34.2~48.8% in sea mustard, 37.5~47.8% in seaweed fusiforme, 42.9­~71.3% in gulf weed, and 37.1~45.1% in sea tangle of brown algae, and 31.3~40.5% in laver, 51.5~60.4% in seaweed dilatata, and 57.1~65.8% in seaweed furcata of red algae. Relatively high levels of both soluble and insoluble dietary fibers were found in seaweed furcata and gulf weed. The ratio of soluble dietary fiber to total dietary fiber was the highest in green laver (43.7~­64.8%), sea mustard (17.5~31.3%), and seaweed furcata (44.7~63.2%) in their respective groups. The highest level of algal polysaccharides was confirmed to be an alkali-soluble alginic acid (9.0~15.1%) in whole brown algae, porphyran (5.8%) in laver, agar (20.0%) in seaweed furcata, and carrageenan (23.8%) in seaweed dilatata of red algae.

해산복족류의 소화관조직중에 분포하는 단백질분해효소의 활성

변재형,김장양,변대석,조득문 한국수산학회 1983 한국수산과학회지 Vol.16 No.3

This work was undertaken to obtain some characteristics of proteolytic enzyme of marine gastropods such as sea hare, Aplysia kurodai, top shell, Turbo cornutks, and abalone, Haliotis discus hannai. An influence of pH, temperature and some chemicals on proteolytic activity of the crude enzyme extracted from digestive tract of the samples was taken into account and the stability of the enzyme during the storage at low temperature was also discussed. In comparison of the activities of the crude enzyme from the samples to the optimum conditions, it was characterized that abalone has twice or the more times higher activities than the other two species of the gastropoda in the acid and weak acid region, while in the alkaline region, sea hare has six or the more times higher activities than the other two species. The proteolytic activity was facilitated by Mn2+, some reducing agents, EDTA and DTT, and inhibited by Hg2+ and SDS, but the other chemicals were not significantly affected to the activity. The low temperature storage of the enzymes of sea hare and top shell at 0�� or -20�� was not affected to the enzymic activity under the optimum pH condition except in the alkaline region. On the other hand, the low temperature storage was brought about no significant effect on the activity of the enzymes extracted from abalone under the optimum condition of the weak acid region, but apparently influenced to the activity under the optimum condition of the acid and alkaline region.

복상어와 고등어의 Trypsin 에 관한 비교 효소학적 연구 : 1 . Trypsin 의 정제와 반응조건 1 . Purifications and Reaction Conditions of the Trypsins

변재형,조득문,허민수 한국수산학회 1991 한국수산과학회지 Vol.24 No.5

연골어류와 경골어류의 진화에 관한 생리생화학적 차이를 특정한 소화효소의 성질과 구조면에서 밝혀 보고자 본 연구를 시도하였다. 양 어종을 대표하는 종류로서 복상어와 고등어를 시료로 선정하고, 소화효소 중 trypsin을 대상으로 하여, 우선 그 정제방법의 확립, 분자량의 측정, 반응 및 안정성에 미치는 pH 및 온도의 영향, 그리고 금속이온과 화학약제가 반응에 미치는 영향 등을 분석 검토하였으며, 그 내용을 요약하면 다음과 같다. 1. 정제방법에 관하여, 먼저 복상어 trypsin은 4 염화탄소로서 탈지 추출한 조효소에 대하여 50∼70% 포화 황산암모늄 염석, DEAE-Sephadex A-50칼럼 크로마토그래피, benzamidine-Sepharose 6B, 친화성 크로마토그래피, Sephadex G-75∼120 겔여과법을 거쳐 disc- 전기영동법, SDS-PAG 전기영동법 및 겔 여과법상 균질상태로 얻었다. 또, 고등어의 trypsin은 역시 4염화탄소로서 탈지 추출한 조효소에 대하여 30∼70% 포화 황산암모늄 염석, benzamidine-Sepharose 6B 친화성 크로마토그래피, DEAE-Sephadex A-50 크로마토그래피 등의 재크로마토그래피, benzamidine-Sepharose 6B에 의한 재크로마토그래피 등의 정제과정을 거쳐 disc- 전기영동법, SDS-PAG 전기영동법 및 겔 여과법으로 균질의 가칭 tryspin-A와 B를 달리 하였다. 2. 이들 유래를 달리하는 각 trypsin의 분자량은 SDS-PAG 전기영동법으로 측정했을 때, 복상어 trypsin 31,700, 고등어 trypsin-A 30,000, 고등어 trypsin-B 29,000, 그리고 겔 여과법으로 측정했을 때, 복상어 21,500, 고등어 trypsin-A 23,700, 고등어 trypsin-B 21,500이었다. 3. 이들 효소들의 알맞은 반응조건은 복상어 trypsin pH 9.0, 45∼50℃, 고등어 trypsin-A와 B pH 8.0, 50℃였다. pH와 온도조건에 따른 안정성을 각각 pH와 온도조건별로 30분간 전처리한 후에 그 활성에 미치는 영향으로 비교해 본 결과, pH의 변화에 대하여는 복상어 trypain은 pH 10.0에서, 그리고 고등어 trypsin-A는 pH 7.0∼9.0에서, 고등어 trypsin-B는 pH 8.0에서 안정하였고, 온도조건에 대하여는 복상어 trypsin은 25℃까지, 그리고 고등어는 trypsin-A가 10℃까지, 고등어 trypsin-B는 35℃까지는 안정하였으나, 그 이후부터는 불안정하였다. 4. 이들 효소들은 공통적으로 금속이온 Ag^(2+), Cu^(2+) 및 Hg^(2+)에 의하여 저해를 받았으며, 그 저해의 정도는 고등어 trypsin-A와 B가 보다 심하게 받았다. 또 이들 효소들은 antipain, leupeptin, TLCK(tosyllysine chloromethyl ketone) 및 SBTI(soybean trypsin inhibitor)에 의하여 현저한 저해를 받았고, PMSF(phenylmethane sulfonylfluoride), DFP(diisopropyl fluorophosphate)및 benzamidine에 의하여 어느정도 저해를 보여 모두 serine- 계의 trypsin임을 뒷받침하였다. To elucidate the physiological and biochemical differences between chondrichthyes and osteichthyes, the properties of the specific digestive enzymes in cat-shark, Cephaloscyllium umbratile, and mackerel, Scomber japonicas, were studied. Homogenous trypsin proved through the disc-electrophoresis, SDS-PAG electrophoresis and gel filtration was obtained from the pancreas of cat-shark by 50∼70% saturated ammonium sulphate fractionation, DEAF-Sephadex A-50 column chromatography, benzamidine-Sepharose 6B affinity chromatography and Sephadex G-75∼120 gel filtration. Two types of trypsins were also obtained from the pyloric caeca of mackerel by 30∼70% saturated ammonium sulphate fractionation and the slightly modified procedure from the method adopted in the purification of cat-shark trypsin. The two trypsins, designated trypsin A and B, were proved their homogeneity by disc- and SDS-PAG electrophoresis and gel filtration. The molecular weights of the trypsins were estimated to be 31,700 for cat-shark trypsin, 30,000 for mackerel trypsin A and 29,000 for mackerel trypsin B by SDS-PAG electrophoresis, but those were estimated to be 21,500 for cat-shark trypsin, 23,700 for mackerel trypsin A and 21,500 for mackerel trypsin B by gel filtration. The trypsins exhibited their optimum conditions at pH 9.0 and on temperature ranged from 45℃ to 50℃ for cat-shark, and at pH 8.0 and a temperature of 50℃ for mackerel trypsin A and B, respectively. The cat-shark trypsin was stable at pH 10.0 and the temperature below 10℃, whereas the mackerel trypsin A and B were stable in the range over pH 7.0 to pH 9.0 below 10℃.