连续酶解花生蛋白及其花生多肽抗氧化活性研究
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摘要
花生蛋白是花生榨油后的副产物,一般作为食品或饲料的原料。本文以花生蛋白为原料,选用Alcalase碱性蛋白酶,连续酶解并得到酶解产物(即花生多肽)。同时对花生多肽的体外和体内的抗氧化性进行研究,并对其进行初步的分离纯化,进一步研究各个分离组分的氨基酸和分子量分布。此研究旨在为工业化连续酶解花生蛋白及花生多肽在功能性食品上的应用提供理论依据,并以此提高花生蛋白的附加值。主要研究内容和结果如下：
1.连续酶解花生蛋白的工艺研究。以水解度(DH)为指标,选用Alcalase碱性蛋白酶,采用酶膜反应器连续酶解花生蛋白。通过单因素试验选取试验因素与水平,并确定了操作压力为0.02MPa。同时设计了四因素(pH值、温度、底物浓度和加酶量)三水平的中心组合响应面试验。得出最佳工艺条件为：pH9.6,温度54℃,底物浓度2%,加酶量7440U／g。通过在DH最佳水解条件下进行水解,实际得到DH为26.13%。
2.花生多肽体外抗氧化活性的研究。考察了花生多肽的还原能力、DPPH·自由基清除能力、ABTS+·自由基清除能力和超氧阴离子(O2-·)自由基清除能力。结果显示,花生多肽具有良好的还原能力,且对这几种自由基均具有不同程度的清除能力。随着花生多肽浓度的增大,其还原能力和对这几种自由基清除能力都增强。同时还测定了花生多肽对DPPH·自由基和超氧阴离子(O2-·)自由基清除能力的半抑制浓度IC50值,花生多肽对DPPH·自由基清除能力的IC50值为0.65mg/ml,对O2-·自由基清除能力的IC50值为24.23mg/ml。
3.花生多肽体内抗氧化活性的研究。采用D-半乳糖诱导衰老小鼠作为模型,分别以小鼠的血清、肝脏和心脏组织中的丙二醛(MDA)含量、超氧化物歧化酶(SOD),谷胱甘肽(GSH)和谷胱甘肽过氧化物酶(GSH-Px)活性作为评价指标,探讨花生多肽在生物体内的抗氧化作用。结果显示：与模型组比较,高剂量的花生多肽能极显著降低D-半乳糖致衰模型小鼠血清、肝脏和心脏中MDA含量(p<0.01),并极显著提高SOD和GSH-Px的活力(p<0.01),同时显著增加GSH的含量(p<0.05)。同时还发现,与模型组比较,各花生多肽给药组MDA含量均有不同程度的降低,GSH含量、SOD和GSH-Px活力均有一定程度的升高,且与花生多肽的给药量存在明显的量效关系。
4.采用葡聚糖凝胶Sephadex G-15对花生多肽进行初步的分离纯化,对其各个组分进行氨基酸成分和分子量分布进行研究,探讨其与抗氧化活性的相关性。花生多肽分离纯化后得到组分Ⅰ和组分Ⅱ这两个组分,通过比较DPPH·和超氧阴离子(O2-·)自由基的清除能力,发现组分Ⅰ的抗氧化活性高于组分Ⅱ。同时对组分Ⅰ和组分Ⅱ进行氨基酸分析得出,组分Ⅰ和组分Ⅱ中不同种类氨基酸含量相差较大,且组分Ⅰ中疏水性氨基酸含量接近50%,远高于组分Ⅱ中的含量。在分子量分布研究中发现,组分Ⅰ和组分Ⅱ中94%的肽段主要集中在3000-5000Da之间,其中组分Ⅱ中分子量分布更为分散些。
Peanut proteins, which have wide source and low price, are generally used in the food processing. In this study, peanut proteins as the raw material, were continuously hydrolyzed by Alcalase protease in the enzyme membrane reactor. This paper main studied antioxidant activity in the vitro and vivo, preliminary purification of peanut peptides. And, amino acids and molecular weight distribution of the various purified fractions were further researched. This study can provide theoretical bases for the industrial continuous hydrolysis of peanut protein and functional foods application of peanut peptides. So this research also increased the added value of peanut protein. The main research content and results are following:
1. Study on the technology of continuous hydrolysis peanut protein. Using hydrolysis degree (DH) as an indicator, peanut proteins were hydrolyzed by Alcalase protease in the enzyme membrane reactor. Through single-factor tests, they select test factors and the level of test factors, and determine the operating pressure of 0.02MPa. And the four factors (pH, temperature, substrate concentration and protease dosage) and three levels response surface methodology (RSM) test were designed. It was found that the optimum condition was pH 9.6, temperature 54℃, substrate concentration 2% and protease dosage 7440u/g. Under the optimum condition, practical DH is 26.13%.
2. Research on the vitro antioxidant activity of peanut peptides. The reducing capacity, DPPH-scavenging capacity, ABTS+·scavenging capacity and O2-·scavenging capacity of peanut peptides were studied. The results showed that peanut peptides had certain reducing capacity and different scavenging effects on different free radicals. The reducing capacity and free radical-scavenging capacity were increased with the increase of peanut peptide concentrations.The scavenging capacity of half-inhibitory concentration (IC50) value was determined on the DPPH·free radical of peanut peptides and O2-·free radical of peanut peptides. Peanut peptides on the DPPH·free radical scavenging capacity of IC50 value was 0.65mg/ml, on the O2-·free radical of IC50 value was 24.23mg/ml.
3. In order to study on the vivo antioxidant activity of peanut pep tides, using D-galactose induced aging mice as a model, the contents of malondialdehyde(MDA) and glutathione(GSH), the activities of superoxide dismutase(SOD) and glutathione peroxidase(GSH-Px) in the serum, liver and heart of each groups mice were determined. The results showed:compared with the model group, the high doses of peanut peptides could significantly(p<0.01) reduce the MDA contents in the serum, live and heart of aged mice induced by D-galactose. It also could significantly(p< 0.01) increased the activities of serum, live and heart SOD and GSH-Px, which increase(p<0.05) the contents of GSH. So peanut peptides have well vivo antioxidant activity.
4. Peanut peptides were separated and purified by sephadex G-15, which got two fractions (ⅠandⅡ). Then, the amino acids and molecular weight distribution of these fractions were further researched. Through testing the DPPH·and O2-·radical scavenging, the antioxidant activity of fractionⅠwas higher than fracitionⅡ. Then analysis of amino acids between two fractions, they were large difference among the contents of different kinds of amino acids. FractionⅠhad the hydrophobic amino acid nearly 50%, much higher than fractionⅡ. Found in the molecular weight distribution, the peptides of fractionⅠandⅡconcentrated in 3000-5000Da. The molecular weight distribution of fractionⅡwas more dispersed than fractionⅠ
1.连续酶解花生蛋白的工艺研究。以水解度(DH)为指标,选用Alcalase碱性蛋白酶,采用酶膜反应器连续酶解花生蛋白。通过单因素试验选取试验因素与水平,并确定了操作压力为0.02MPa。同时设计了四因素(pH值、温度、底物浓度和加酶量)三水平的中心组合响应面试验。得出最佳工艺条件为：pH9.6,温度54℃,底物浓度2%,加酶量7440U／g。通过在DH最佳水解条件下进行水解,实际得到DH为26.13%。
2.花生多肽体外抗氧化活性的研究。考察了花生多肽的还原能力、DPPH·自由基清除能力、ABTS+·自由基清除能力和超氧阴离子(O2-·)自由基清除能力。结果显示,花生多肽具有良好的还原能力,且对这几种自由基均具有不同程度的清除能力。随着花生多肽浓度的增大,其还原能力和对这几种自由基清除能力都增强。同时还测定了花生多肽对DPPH·自由基和超氧阴离子(O2-·)自由基清除能力的半抑制浓度IC50值,花生多肽对DPPH·自由基清除能力的IC50值为0.65mg/ml,对O2-·自由基清除能力的IC50值为24.23mg/ml。
3.花生多肽体内抗氧化活性的研究。采用D-半乳糖诱导衰老小鼠作为模型,分别以小鼠的血清、肝脏和心脏组织中的丙二醛(MDA)含量、超氧化物歧化酶(SOD),谷胱甘肽(GSH)和谷胱甘肽过氧化物酶(GSH-Px)活性作为评价指标,探讨花生多肽在生物体内的抗氧化作用。结果显示：与模型组比较,高剂量的花生多肽能极显著降低D-半乳糖致衰模型小鼠血清、肝脏和心脏中MDA含量(p<0.01),并极显著提高SOD和GSH-Px的活力(p<0.01),同时显著增加GSH的含量(p<0.05)。同时还发现,与模型组比较,各花生多肽给药组MDA含量均有不同程度的降低,GSH含量、SOD和GSH-Px活力均有一定程度的升高,且与花生多肽的给药量存在明显的量效关系。
4.采用葡聚糖凝胶Sephadex G-15对花生多肽进行初步的分离纯化,对其各个组分进行氨基酸成分和分子量分布进行研究,探讨其与抗氧化活性的相关性。花生多肽分离纯化后得到组分Ⅰ和组分Ⅱ这两个组分,通过比较DPPH·和超氧阴离子(O2-·)自由基的清除能力,发现组分Ⅰ的抗氧化活性高于组分Ⅱ。同时对组分Ⅰ和组分Ⅱ进行氨基酸分析得出,组分Ⅰ和组分Ⅱ中不同种类氨基酸含量相差较大,且组分Ⅰ中疏水性氨基酸含量接近50%,远高于组分Ⅱ中的含量。在分子量分布研究中发现,组分Ⅰ和组分Ⅱ中94%的肽段主要集中在3000-5000Da之间,其中组分Ⅱ中分子量分布更为分散些。
Peanut proteins, which have wide source and low price, are generally used in the food processing. In this study, peanut proteins as the raw material, were continuously hydrolyzed by Alcalase protease in the enzyme membrane reactor. This paper main studied antioxidant activity in the vitro and vivo, preliminary purification of peanut peptides. And, amino acids and molecular weight distribution of the various purified fractions were further researched. This study can provide theoretical bases for the industrial continuous hydrolysis of peanut protein and functional foods application of peanut peptides. So this research also increased the added value of peanut protein. The main research content and results are following:
1. Study on the technology of continuous hydrolysis peanut protein. Using hydrolysis degree (DH) as an indicator, peanut proteins were hydrolyzed by Alcalase protease in the enzyme membrane reactor. Through single-factor tests, they select test factors and the level of test factors, and determine the operating pressure of 0.02MPa. And the four factors (pH, temperature, substrate concentration and protease dosage) and three levels response surface methodology (RSM) test were designed. It was found that the optimum condition was pH 9.6, temperature 54℃, substrate concentration 2% and protease dosage 7440u/g. Under the optimum condition, practical DH is 26.13%.
2. Research on the vitro antioxidant activity of peanut peptides. The reducing capacity, DPPH-scavenging capacity, ABTS+·scavenging capacity and O2-·scavenging capacity of peanut peptides were studied. The results showed that peanut peptides had certain reducing capacity and different scavenging effects on different free radicals. The reducing capacity and free radical-scavenging capacity were increased with the increase of peanut peptide concentrations.The scavenging capacity of half-inhibitory concentration (IC50) value was determined on the DPPH·free radical of peanut peptides and O2-·free radical of peanut peptides. Peanut peptides on the DPPH·free radical scavenging capacity of IC50 value was 0.65mg/ml, on the O2-·free radical of IC50 value was 24.23mg/ml.
3. In order to study on the vivo antioxidant activity of peanut pep tides, using D-galactose induced aging mice as a model, the contents of malondialdehyde(MDA) and glutathione(GSH), the activities of superoxide dismutase(SOD) and glutathione peroxidase(GSH-Px) in the serum, liver and heart of each groups mice were determined. The results showed:compared with the model group, the high doses of peanut peptides could significantly(p<0.01) reduce the MDA contents in the serum, live and heart of aged mice induced by D-galactose. It also could significantly(p< 0.01) increased the activities of serum, live and heart SOD and GSH-Px, which increase(p<0.05) the contents of GSH. So peanut peptides have well vivo antioxidant activity.
4. Peanut peptides were separated and purified by sephadex G-15, which got two fractions (ⅠandⅡ). Then, the amino acids and molecular weight distribution of these fractions were further researched. Through testing the DPPH·and O2-·radical scavenging, the antioxidant activity of fractionⅠwas higher than fracitionⅡ. Then analysis of amino acids between two fractions, they were large difference among the contents of different kinds of amino acids. FractionⅠhad the hydrophobic amino acid nearly 50%, much higher than fractionⅡ. Found in the molecular weight distribution, the peptides of fractionⅠandⅡconcentrated in 3000-5000Da. The molecular weight distribution of fractionⅡwas more dispersed than fractionⅠ
引文
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56. Cao G H, Sofic E, Prior R L. Antioxidant and prooxidant behavior of flavonoids: structure-activity relationship. Free Radical Bio & Medicine,1997,22 (5): 749-760
57. Chen H M, Muramoto K, Yamauchi F. Antioxidative properties of histidine-containing peptides designed from peptide fragments found in the digests of a soy-bean protein. J Agric Food Chem,1998,46:49-53
58. Chen H M, Muramoto K, Yamauchi F. Structural analysis of antioxidative peptides from soybean β-conglycinin. J Agric Food Chem,1995,43:574-578
59. Chi-Yue C, Kuei-Ching W, Shu-Hua C. Antioxidant properties and protein compositions of porcine haemoglobin hydrolysates. Food Chemistry,2007,100: 1537-1543
60. Duh PD, Yen GC. Antioxidant efficacy of Methanolic extracts from peanut hulls. Journal of the American oil chemists'society,1993,70(4):383-385
61. D.Belhocine, H.Mokrane, H.Grib, H.Lounici, A.Pauss, N.Mameri. Optimization of enzymatic hydrolysis of haemoglobin in a continuous membrane bioreactor. Chemical Engineering Joural,2000,76:189-196
62. D. Misun, L. Curda, P. Jelen. Batch and continuous hydrolysis of ovine whey protein. Small Ruminant Research,2008,79:51-56
63. Eresha Mendis, Niranjan Rajapakse, Se-Kwon Kim. Antioxidant properties of a radical-scavenging peptide purified from enzymatically prepared fish skin gelatin hydrolysate. Agric and Food Chemis,2005(53):581-587
64. G.M.Rios, M.P.Belleville, D.Paolucci, J.Sanchez. Progress in enzymatic membrane reactors-a review. Journal of Membrane Science,2004,242:189-196
65. Ho W S, Sirkar K K. Membrane Handbook. New York:Van NostranReinhold, 1992,353-413
66. Hee-Guk Byun, Se-Kwon Kim. Purification and characterization of angiotensin I converting enzyme (ACE) inhibitory peptides from Alaska pollack(Theragra chalcogramma) skin. Process Biochemistry,2001,36:1155-1162
67. Jeon Y J, Byun H G, Kim S K. Improvement of functional properties of cod frame protein hydrolysates using ultrafiltration membrane. Process Biochem, 1999,35(5):471-478
68. Jae-Young Je, Pyo-Jam Park, Se-Kwon Kim.Antioxidant activity of a peptide isolated from Alaska Pollack (Theragra chalcogramma) frame protein hydrolysate.Food Research International,2005(38):45-50
69. Kong B, Xiong Y L. Antioxidant activity of zein hydrolyates in a liposome system and the possible mode of action. J. Agric. Food Chem,2006, (54): 6059-6068
70. M.Prata-Vidal, S.Bouhallab, G.Henry, P.Aimar. An experimental study of caseinomacropeptide hydrolysis by trypsin in a continuous membrane reactor. Biochemical Engineering Journal,2001,8:195-202
71. Mimouni B, Azanza J L, Raymond J. Influence of double enzymic hydrolyses on gluten functionality. Journal of the Science of Food and Agriculture,1999,79: 1048-105
72. Marklund S, Marklund G. Involvement of the superoxide anion radical in the autoxidation of pyrogallal and a convenient assay for superoxide dismutase. European Journal of Biochemistry.1974,47(3):467-474
73. N.Prevot-D'Alvise, C. Lesueur-Lambert, A. Fertin-Bazus. Continuous enzymatic solubilization of alfalfa protein in an ultrafiltration reactor. Enzyme and Microbial Technology,2004,34:380-391
74. N.D'Alvise, C.Lesueur-Lambert, B.Fertin, P.Dhulster, D.Guillochon. Hydrolysis and large scale ultrafiltration study of alfalfa protein concentrate enzymatic hydrolysate. Enzyme and Microbial Technology,2000,27:286-294
75. Oyaizu M. Antioxidative activities of browning products of glucosamine fractionated by organic solvent and thin-layer chromatography. Nippon Shokuhin Kogyo Gakkaishi,1988,35:771-775
76. R.Amarowicz, F.Shahidi. Antioxidant activity of peptide fractions of capelin protein hydrolysates. Food Chemistry,1997,58(4):355-359
77. Seronei Chelulei Cheison, Zhang Wang, Shi-Ying Xu. Use of response surface methodology to optimize the hydrolysis of whey protein isolate in a tangential flow filter membrane reactor. Journal of Food Engineering,2007,80:1134-1145
78. Seronei Chelulei Cheison, Zhang Wang, Shi-Ying Xu. Use of response surface methodology to optimize the hydrolysis of whey protein isolate in a tangential flow filter membrane reactor. Journal of Food Engineering,2007,80:1134-1145
79. Koichiro Saito, Dong-Hao Jin, Tomohisa Ogawa, Koji Muramoto, Eiko Hatakeyama, Tadashi Yasuhara, Kiyoshi Nokihara. Antioxdative properties of tripeptide libraries prepared by the combinatorial chemistry. J Agric Food Chem, 2003,51:3668-3674
80. Saiga A, Tanabe S, Nishimura T. Antioxdiant activity of peptides obtained from porcine myofibrillar proteins by protease treatment. J Agric Food Chem,2003, 51(12):3661-3667
81. Wen-Dee Chiang, Chieh-Jen Shih, Yan-Hwa Chu. Functional properties of soy protein hydrolysate produced from a continuous membrane reactor system.Food Chemistry,1999,65:189-194
82. Wen-Dee Chiang, May-June Tsou, Zong-Yao Tsai, Tsun-Chung Tsai. Angiotensin I-converting enzyme inhibitor derived from soy protein hydrolysate and produced by using membrane reactor. Food Chemistry,2006,98:725-732
83. Hui-Chun Wu, Hua-Ming Chen, Chyuan-yuan Shiau. Free amino acids and peptides as related to antioxidant properties in protein hydrolysates of mackerel (Scomber austriasicus). Food Research International,2003,36:949-957
84. Xinyan Peng, Youling L.Xiong, Baohua Kong. Antioxdant activity of peptide fractions from whey protein hydrolysates as measured by electro spin resonance. Food Chemistry,2009,113:196-201
85. Zhengjun Xie, Junrong Huang, Xueming Xu, Zhengyu Jin. Antioxidant activity of peptides isolated from alfalfa leaf protein hydrolysate. Food Chemistyr,2008, 111:370-376
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54. Antonio Guadix, Fernando Camacho, Emilia M.Guadix. Production of whey protein hydrolysates with reduced allergenicity in a stable membrane reactor. Journal of Food Engineering,2006,72:398-405
55. Aide Perea, Unai Ugalde. Continuous hydrolysis of whey proteins in a membrane recycle reactor. Enzyme and microbial Technology,1996,18:29-34
56. Cao G H, Sofic E, Prior R L. Antioxidant and prooxidant behavior of flavonoids: structure-activity relationship. Free Radical Bio & Medicine,1997,22 (5): 749-760
57. Chen H M, Muramoto K, Yamauchi F. Antioxidative properties of histidine-containing peptides designed from peptide fragments found in the digests of a soy-bean protein. J Agric Food Chem,1998,46:49-53
58. Chen H M, Muramoto K, Yamauchi F. Structural analysis of antioxidative peptides from soybean β-conglycinin. J Agric Food Chem,1995,43:574-578
59. Chi-Yue C, Kuei-Ching W, Shu-Hua C. Antioxidant properties and protein compositions of porcine haemoglobin hydrolysates. Food Chemistry,2007,100: 1537-1543
60. Duh PD, Yen GC. Antioxidant efficacy of Methanolic extracts from peanut hulls. Journal of the American oil chemists'society,1993,70(4):383-385
61. D.Belhocine, H.Mokrane, H.Grib, H.Lounici, A.Pauss, N.Mameri. Optimization of enzymatic hydrolysis of haemoglobin in a continuous membrane bioreactor. Chemical Engineering Joural,2000,76:189-196
62. D. Misun, L. Curda, P. Jelen. Batch and continuous hydrolysis of ovine whey protein. Small Ruminant Research,2008,79:51-56
63. Eresha Mendis, Niranjan Rajapakse, Se-Kwon Kim. Antioxidant properties of a radical-scavenging peptide purified from enzymatically prepared fish skin gelatin hydrolysate. Agric and Food Chemis,2005(53):581-587
64. G.M.Rios, M.P.Belleville, D.Paolucci, J.Sanchez. Progress in enzymatic membrane reactors-a review. Journal of Membrane Science,2004,242:189-196
65. Ho W S, Sirkar K K. Membrane Handbook. New York:Van NostranReinhold, 1992,353-413
66. Hee-Guk Byun, Se-Kwon Kim. Purification and characterization of angiotensin I converting enzyme (ACE) inhibitory peptides from Alaska pollack(Theragra chalcogramma) skin. Process Biochemistry,2001,36:1155-1162
67. Jeon Y J, Byun H G, Kim S K. Improvement of functional properties of cod frame protein hydrolysates using ultrafiltration membrane. Process Biochem, 1999,35(5):471-478
68. Jae-Young Je, Pyo-Jam Park, Se-Kwon Kim.Antioxidant activity of a peptide isolated from Alaska Pollack (Theragra chalcogramma) frame protein hydrolysate.Food Research International,2005(38):45-50
69. Kong B, Xiong Y L. Antioxidant activity of zein hydrolyates in a liposome system and the possible mode of action. J. Agric. Food Chem,2006, (54): 6059-6068
70. M.Prata-Vidal, S.Bouhallab, G.Henry, P.Aimar. An experimental study of caseinomacropeptide hydrolysis by trypsin in a continuous membrane reactor. Biochemical Engineering Journal,2001,8:195-202
71. Mimouni B, Azanza J L, Raymond J. Influence of double enzymic hydrolyses on gluten functionality. Journal of the Science of Food and Agriculture,1999,79: 1048-105
72. Marklund S, Marklund G. Involvement of the superoxide anion radical in the autoxidation of pyrogallal and a convenient assay for superoxide dismutase. European Journal of Biochemistry.1974,47(3):467-474
73. N.Prevot-D'Alvise, C. Lesueur-Lambert, A. Fertin-Bazus. Continuous enzymatic solubilization of alfalfa protein in an ultrafiltration reactor. Enzyme and Microbial Technology,2004,34:380-391
74. N.D'Alvise, C.Lesueur-Lambert, B.Fertin, P.Dhulster, D.Guillochon. Hydrolysis and large scale ultrafiltration study of alfalfa protein concentrate enzymatic hydrolysate. Enzyme and Microbial Technology,2000,27:286-294
75. Oyaizu M. Antioxidative activities of browning products of glucosamine fractionated by organic solvent and thin-layer chromatography. Nippon Shokuhin Kogyo Gakkaishi,1988,35:771-775
76. R.Amarowicz, F.Shahidi. Antioxidant activity of peptide fractions of capelin protein hydrolysates. Food Chemistry,1997,58(4):355-359
77. Seronei Chelulei Cheison, Zhang Wang, Shi-Ying Xu. Use of response surface methodology to optimize the hydrolysis of whey protein isolate in a tangential flow filter membrane reactor. Journal of Food Engineering,2007,80:1134-1145
78. Seronei Chelulei Cheison, Zhang Wang, Shi-Ying Xu. Use of response surface methodology to optimize the hydrolysis of whey protein isolate in a tangential flow filter membrane reactor. Journal of Food Engineering,2007,80:1134-1145
79. Koichiro Saito, Dong-Hao Jin, Tomohisa Ogawa, Koji Muramoto, Eiko Hatakeyama, Tadashi Yasuhara, Kiyoshi Nokihara. Antioxdative properties of tripeptide libraries prepared by the combinatorial chemistry. J Agric Food Chem, 2003,51:3668-3674
80. Saiga A, Tanabe S, Nishimura T. Antioxdiant activity of peptides obtained from porcine myofibrillar proteins by protease treatment. J Agric Food Chem,2003, 51(12):3661-3667
81. Wen-Dee Chiang, Chieh-Jen Shih, Yan-Hwa Chu. Functional properties of soy protein hydrolysate produced from a continuous membrane reactor system.Food Chemistry,1999,65:189-194
82. Wen-Dee Chiang, May-June Tsou, Zong-Yao Tsai, Tsun-Chung Tsai. Angiotensin I-converting enzyme inhibitor derived from soy protein hydrolysate and produced by using membrane reactor. Food Chemistry,2006,98:725-732
83. Hui-Chun Wu, Hua-Ming Chen, Chyuan-yuan Shiau. Free amino acids and peptides as related to antioxidant properties in protein hydrolysates of mackerel (Scomber austriasicus). Food Research International,2003,36:949-957
84. Xinyan Peng, Youling L.Xiong, Baohua Kong. Antioxdant activity of peptide fractions from whey protein hydrolysates as measured by electro spin resonance. Food Chemistry,2009,113:196-201
85. Zhengjun Xie, Junrong Huang, Xueming Xu, Zhengyu Jin. Antioxidant activity of peptides isolated from alfalfa leaf protein hydrolysate. Food Chemistyr,2008, 111:370-376