• Food Science and Biotechnology
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• 제18권5호
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• pp.1173-1179
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• 2009

Trypsin inhibitors and lectins are defense proteins produced by many organisms. From Chinese 'Large Black Soybeans', a 60 kDa lectin and a 20 Da trypsin inhibitor (TI) were isolated using chromatography on Q-Sepharose, Mono Q, and Superdex 75. The TI inhibited trypsin and chymotrypsin with an $IC_{50}$ of 5.7 and $5{\mu}M$, respectively. Trypsin inhibitory activity of the TI was stable from pH 3 to 13 and from 0 to $65^{\circ}C$. Hemagglutinating activity of the lectin was stable from pH 2 to 13 and from 0 to $65^{\circ}C$. The TI was inhibited by dithiothreitol, signifying the importance of disulfide bond. The TI and the lectin inhibited HIV-1 reverse transcriptase ($IC_{50}$=44 and $26{\mu}M$), and proliferation of breast cancer cells ($IC_{50}$=42 and $13.5{\mu}M$) and hepatoma cells ($IC_{50}$=96 and $175{\mu}M$). The hemagglutinating activity of the lectin was inhibited most potently by L-arabinose. Neither the lectin nor the TI displayed antifungal activity.

• 한국식품영양과학회지
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• 제7권2호
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• pp.7-12
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• 1978

본(本) 실험(實驗)은 한국산(韓國産) 대두(大豆) 5 품종(品種)(수원62호, 수원81호, 수원 82호, 수원 83호, 경남 3 호)의 수용(水溶), 단백질(蛋白質)을 gel filtration하여 trypsin inhibitor를 fractionation하여 대두품종별(大豆品種別) trypsin inhibitor의 pattern을 조사(調査)하고 또 이의 내열성(耐熱性)을 살펴보기 위한 목적(目的)으로 시도하여 다음과 같은 결과(結果)를 얻었다. 대두(大豆)의 수용성(水溶性) 단백질(蛋白質)을 추출(抽出)하여 sephadex G-75로 column chromatography하여 분획한 결과(結果) 3개(個)의 peak를 얻었으며, TIA는 두번째 peak 부근에 걸쳐 있었고 trypsin inhibitor는 모두 4개의 fraction(FI, FII, FIII, FIV)으로 나뉘어졌으며, trypsin inhibitor pattern은 수원81호, 수원82호, 수원83호는 FIII의 TIA가 높았으며, 경남 3호 및 수원62호는 FIV에서 높게 나타났다. gel filtration에서 분획(分劃)한 4개의 fraction에 있어서 단백질(蛋白質) mg당(當) TIA는 모두 FIV가 가장 높게 나타났으며 이 fraction의 내열성(耐熱性)은 $100^{\circ}C$에서 20분간(分間) 가열처리(加熱處理)를 기준으로 했을 때 대체로 수원81호가 가장 내열성(耐熱性)이 약했으며 경남 3 호, 수원38호, 수원82호, 수원62호의 순서로 내열성(耐熱性)이 강하게 나타났고 fraction별(別)로는 수원82호를 제외하고는 모두 FIV가 가장 내열성(耐熱性)이 강하게 나타났으며 대체로 FI의 내열성(耐熱性)이 약하였으며 경남 3 호의 FI은 $100^{\circ}C$ 20분간(分間) 처리(處理)에서 완전(完全)히 소실되었다.

• International Journal of Industrial Entomology and Biomaterials
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• 제8권2호
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• pp.195-200
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• 2004

Trypsin can be immobilized on silk fibroin fiber (SFF) by introducing several spacer arms, such as ethylene diamine (ED), bovine serum albumin (BSA) and silk sericin (SS). Direct immobilization on silk fiber (SFFGA) has low activity because of the steric hindrance between the trypsin and substrate. The introduction of spacer arms onto SFF-GA can enhance the activity of trypsin by reducing the steric hindrance. When ED is used as a spacer arm, the activity of trypsin has increased but its stability decreased due to the increased hydrophobicity of SFF. BSA and SS, as a spacer arm, have better results in both activity and stability. SFF-BSA shows some decrease in the specific activity due to improper immobilizatin. SFF-SS maintained 90% of its initial activity even after 12 hrs incubation at $50^{\circ}C$. In the case of repeated hydrolysis of silk sericin with immobilized trypsin, SFF-GA and SFF-ED lost 50% of their initial activity right after first run, whereas SFF-BSA and SFF-SS maintained 80% of their initial activities even after 5 runs. Higher operational stability is due to increased hydrophilicity of SFF by introducing hydrophilic spacer arms such as BSA and SS. The high content of serine in SS increases the hydrophilicity of SFF resulting the best results among other spacer arms.

• Asian-Australasian Journal of Animal Sciences
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• 제16권12호
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• pp.1822-1829
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• 2003

Porcine colostrum and milk were separated into the acid-soluble and casein fractions by acidification followed by centrifuge. The acid-soluble fraction of porcine colostrum was further separated by liquid chromatography and anisotropic membrane filtration. Trypsin and chymotrypsin inhibitory capacity in porcine colostrum, milk and their components was determined by incubating bovine trypsin or chymotrypsin in a medium containing their corresponding substrates with or without addition of various amounts of porcine colostrum, porcine milk or their components. The inhibition of insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF) degradation in pig small intestinal contents by porcine colostrum was measured by incubating iodinated IGF-I or EGF with the intestinal contents with or without addition of porcine colostrum. Degradation of labeled IGF-I or EGF was determined by monitoring the generation of radioactivity soluble in 30% trichloroacetic acid (TCA). The results showed that porcine colostrum had high levels of trypsin and chymotrypsin inhibitory activity and increased the stability of IGF-I and EGF in pig intestinal contents. The inhibitory activity declined rapidly during lactation. It was also found that trypsin and chymotrypsin inhibitory activity and the inhibition on IGF-I and EGF degradation in the acid-soluble fraction were higher than that in the casein fraction. Heat-resistance study indicated that trypsin inhibitors in porcine colostrum survived heat treatments of $100^{\circ}C$ water bath for up to 10 min, but exposure to boiling water bath for 30 min significantly decreased the inhibitory activity. Compared with the trypsin inhibitors, the chymotrypsin inhibitors were more heatsensitive. Separation of the acid-soluble fraction of porcine colostrum by liquid chromatography and anisotropic membrane filtration revealed that the trypsin and chymotrypsin inhibitory capacity was mainly due to a group of small proteins with molecular weight of 10,000-50,000. In conclusion, the present study confirmed the existence of high levels of protease inhibitors in porcine colostrum, and the inhibition of porcine colostrum on degradation of milk-borne growth factors in the pig small intestinal tract was demonstrated for the first time.

• Asian-Australasian Journal of Animal Sciences
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• 제17권5호
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• pp.712-718
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• 2004

This study was carried out to separate the calcium-binding protein derived from enzymatic hydrolysates of cheese whey protein. CWPs (cheese whey protein) heated for 10 min at $100^{\circ}C$ were hydrolyzed by trypsin, papain W-40, protease S, neutrase 1.5 and pepsin, and then properties of hydrolysates, separation of calcium-binding protein and analysis of calcium-binding ability were investigated. The DH (degree of hydrolysis) and NPN (non protein nitrogen) of heated-CWP hydrolysates by commercial enzymes were higher in trypsin than those of other commercial enzymes. In the result of SDS-PAGE (sodium dodecyl sulphate polyacrylamide gel electrophoresis), $\beta$-LG and $\alpha$-LA in trypsin hydrolysates were almost eliminated and the molecular weight of peptides derived from trypsin hydrolysates were smaller than 7 kDa. In the RP-HPLC (reverse phase HPLC) analysis, $\alpha$-LA was mostly eliminated, but $\beta$-LG was not affected by heat treatment and the RP-HPLC patterns of trypsin hydrolysates were similar to those of SDS-PAGE. In ion exchange chromatography, trypsin hydrolysates were shown to peak from 0.25 M NaCl and 0.5 M NaCl, and calcium-binding ability is associated with the large peak, which was eluted at a 0.25 M NaCl gradient concentration. Based on the results of this experiment, heated-CWP hydrolysates by trypsin were shown to have calcium-binding ability.

• 한국식품과학회지
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• 제27권5호
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• pp.708-715
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• 1995

Pepsin, papain 및 trypsin 처리 참깨박 단백질의 기능성의 변화를 조사한 결과 용해도에 있어 pH 4에서 2%의 대조군에 비해 $53{\sim}94%$까지 뚜렷한 증가를 보였으며, trypsin에 의한 10%, 20% 가수분해도 처리군은 등전점에서 약 6배, papain에 의한 10% 가수분해도 처리군은 약 4.5배 가량의 유화능의 향상을 보였다. 기포 형성력은 각 효소 처리군의 30% 가수분해도 처리군의 알칼리 영역을 제외한 나머지 영역에서 전반적인 증가를 보였다. trypsin, papain 처리군의 겉보기 밀도와 수분 흡착력은 약 0.1 g/ml와 $0.3{\sim}0.7\;ml/g$ 정도 감소하였으나, 유지흡착력은 약 1 ml/g 정도의 증가를 보였다.

• 농업과학연구
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• 제25권1호
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• pp.52-67
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• 1998

본 연구는 포유동물의 젖에 함유되어 있는 lactoferrin(LF)의 생리적 특성을 밝혀 기능성 식품 특히 항균작용을 이용한 식품제조업에의 이용을 위한 기초자료를 얻고자 실시하였다. 따라서 젖소의 초유로부터 lactoferrin을 분리 정제하여, 소화부위에 따른 lactoferrin의 항균활성을 분석하고자 pepsin, trypsin 및 chymotrypsin으로 분해하여 lactoferrin의 항균력을 측정하고, gel-filtration으로 분리 정제된 각각의 fraction별로 단백질을 정량하여 Escherichia coli와 Staphylococcus aureus에 접종하여 항균효과를 지니는 각 효소별 fraction의 분자량과 peptide fragment를 검토하였다. 1. 젖소의 초유로부터 분리 정제된 lactoferrin(LF)을 단백질 분해 효소인 pepsin, trypsin 및 chymotrypsin으로 분해한 결과 bovine lactoferrin은 SDS-PAGE를 수행하여 band를 확인할 수 있었다. pepsin으로 분해된 lactoferrin은 분자량 14KDa까지에서도 band를 확인할 수가 없었으며, trypsin과 chymotrypsin으로 분해한 lactoferrin은 분해되지 않은 lactoferrin이 존재함을 나타내고, 33KDa에서도 band를 확인할 수 있었다. 2. Sephadex G-50 column을 사용하여 bovine lactoferrin를 효과적으로 정제하였다. Sephadex G-50 column chromatography를 수행 한 결과 bovine lactoferrin은 Tris-HCl사이에서 용출되었으며, pepsin, trypsin 및 chymotrypsin으로 분해한 lactoferrin은 각각 2, 3, 2 개의 peak를 보였고, HPLC 분석결과 첫번째 peak는 주로 분해되지 않은 lactoferrin 수용체가 존재하는 것으로 확인되었으며, trypsin과 chymotrypsin처리에서 항균효과도 유사점을 관찰할 수 있었다. 3. 효소처리한 lactoferrin의 항균효과를 알아보기 위하여 Escherichia coli와 Staphylococcus aureus를 접종하여 항균활성을 비교하였다. 그 결과 각 효소에 대하여 pepsin으로 처리한 lactoferrin을 접종한 시험구가 대조구에 비하여 낮은 생장율을 보여 현저한 항균활성을 지님을 알 수 있었다. 그리고 trypsin과 chymotrysin에 의한 분해물도 미생물 배양 8시간까지는 효소 처리전 bovine lactoferrin보다는 항균활성을 나타냄을 알 수 있었다. 4. Sephadex G-50 column을 사용하여 분리 정제된 bovine lactoferrin fraction별로 SDS-PAGE를 실시한 결과 lactoferrin fraction의 경우는 chromatography 수행 결과와 비교하여, pepsin과 chymotrypsin 분해물은 저분자량임을 알 수 있었고, trypsin에 의한 lactoferrin만이 단일 band를 보임으로서 단백질 분해의 특징을 볼 수 있었다.

• 생명과학회지
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• 제30권1호
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• pp.82-87
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• 2020

쌍별 귀뚜라미(G. bimaculatus)가 식량공급이 제한적이거나 starvation상태를 어떻게 극복하는지를 알기 위하여 소화기관(전장, 중장, 후장)에서 소화작용의 가장 중요한 3대 효소인 amylase, trypsin, lipase의 유전자발현을 조사하였다. G. bimaculatus의 전장에서는 amylase, trypsin, lipase 유전자, 중장은 amylase 유전자, 후장은 amylase, trypsin 유전자의 발현이 먹이 의존적이지만, 중장의 trypsin, lipase 유전자와 후장의 lipase 유전자는 먹이 비의존적인 발현으로 항상 일정한 상태를 유지한다. 이 결과는 곤충이 starvation과 같은 외부환경에 적응 및 생존하는지를 관련 유전자들의 발현수준에서 접근할 수 있는 중요한 실마리를 제공할 것이다.

• 한국수산과학회지
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• 제44권1호
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• pp.8-17
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• 2011

To identify and examine the distribution of proteolytic inhibitory activity in crude extracts from fish eggs, and to determine the applicability of these protease inhibitors as anti-degradation agents in surimi-based products and fish meat, we compared the inhibitory activities of various extracts from fish eggs to those of commercial proteases, such as trypsin and papain. We used the optimal conditions for the screening of trypsin activity: 30 ug/uL of 0.1% trypsin and 0.6 mM Na-benzoyl-L-arginine-p-nitroanilide (BAPNA) with a pH of 8.0 at $40^{\circ}C$ for 60 min. The activities of papain and four commercial proteases were investigated after mixing with 100 ug/uL enzymes and 0.3% casein with a pH of 8.0 at $40^{\circ}C$ for 60 min. We performed a screening assay to detect the inhibitory activity (%) of crude extracts from eight species of fish eggs against the target proteases trypsin and papain. The assay revealed a wide distribution of trypsin and papain inhibitors in fish eggs. The specific inhibitory activities (11.6.28.6 U/mg) of crude extracts from fish eggs against trypsin and BAPNA substrate were higher than that (0.64 U/mg) of egg whites, used as a commercial inhibitor. The inhibitory activities of crude extracts from fish eggs against trypsin, and of egg whites against casein substrate (1.94.4.51 U/mg), were higher than those of papain (0.24.1.57 U/mg) and commercial protease (0.04.0.32 U/mg). The extracts from fish eggs were rich in protease inhibitors that exhibited strong inhibitory activity against trypsin, a serine protease, and papain, a cysteine protease.

• 한국식품영양과학회지
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• 제41권6호
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• pp.840-845
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• 2012

본 연구는 감태 물 추출물의 일반특성과 trypsin 저해활성을 알아보고 산업적으로 이용가능성을 확인하기 위하여 열 및 pH에 대한 안정성을 실시하였다. 감태 물 추출물의 색도 및 pH를 측정한 결과, 색도는 명도가 86.21로 높고 적색도 및 황색도가 각각 0.38 및 15.49로 낮게 나타났으며 pH는 6.17로 약산성으로 나타났다. 감태 물 추출물의 trypsin 저해활성은 5, 2.5 및 1 mg/mL 농도에서 각각 76.21%, 62.41% 및 60.41%를 나타냈으며 $IC_{50}$값은 0.83 mg/mL이었다. 열 및 pH에 대한 안정성을 측정한 결과, $80^{\circ}C$까지 열처리에 안정하였고 pH 2~8 범위에서 안정하였으나 $100^{\circ}C$와 $121^{\circ}C$ 열처리와 pH 10에서 활성이 약간 감소하였으나 전체적으로 높은 활성을 유지하여 열 및 pH에 안정한 것을 확인하였다. 이상의 결과를 통해 감태 물 추출물이 지니는 trypsin 저해활성이 열 및 pH에 대해 안정성을 지녀 식품산업에 응용 가능할 것으로 사료된다.