酶解咸鸭蛋蛋清制取白蛋白肽的研究
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摘要
生物活性肽由于其高效的营养价值,功能活性,是目前营养界研究得比较多的一个热门课题,而咸鸭蛋蛋清作为食品工业生产中的废弃物虽然含有较多的优质蛋白质,却因其盐分含量较高而无法利用遭到丢弃。
本论文研究以本身利用价值不大的咸鸭蛋蛋清为原料,经过选择高效率的蛋白酶水解,制取分子量小于5000的富含白蛋白肽的生物活性肽产品,并利用膜过滤技术去除原有的以及水解过程中增加的盐分,使得产品中的盐含量大大降低,符合人体食用的要求。
本文首先研究了咸鸭蛋蛋清制取白蛋白肽的工艺及工艺优化。工艺优化的重点是酶解反应条件的优化。通过对酶种类、底物浓度、酶浓度、酶解体系的pH、酶解温度和酶解时间等进行单因素试验,以及随后的正交试验综合分析,得到了酶解反应的最佳条件:选用酸性蛋白酶、稀释倍数为1、酶用量为3000U/g、pH为2.5、反应温度为45℃、酶解时间为6h。
接着,本文又研究了脱盐方法的选择。根据对原料脱盐和水解以后对水解液脱盐的优缺点的分析,选择采用膜过滤技术对原料或水解液进行脱盐试验。利用超滤技术对咸鸭蛋蛋清直接脱盐,选用截留分子量为10000的超滤膜可使脱盐率达到83.81%,蛋白质截留率达到86.1%。利用纳滤技术和电渗析技术对水解液进行脱盐,当pH3.2时用NF-90的纳滤膜使脱盐率达到94.3%,蛋白质截留率达到84.27%。在pH4.2时利用电渗析使脱盐率达到96.69%,蛋白质截留率达到71.4%。
Bioactive peptide has a high nutrition of value and more functional activities. So it is a popular subject in nutrition now. The egg white of salted duck eggs contains abundance of high quality protein, but it can't be used directly, and is commonly discarded because of its high salt content. That induced serious environment problem and great waste of the high quality protein.This paper researched on the bioactive peptides made from salted duck eggs white which was rich of ovalbumin and was of little commercial value through hydrolyzing by proteases and subsequent membrane filtration to get rid of the extra salt so that the final product could be more useful for human and animals.Firstly, the technology of making ovalbumin peptides was optimized. The enzymatic reaction conditions such as enzyme species, substrate concentration, enzyme concentration, pH, temperature and time of reaction was studied. The optimal conditions of enzymatic treatment was obtained as follows: chose acid proteinase, dilution rate was 1, dosage of enzyme was 3000U/g, pH was 2.5, temperature was 45 ℃, and raction for 6hours.Then the desalting method of the final hydrolyzed product was compared and chosen. The advantage and disadvantage of desalting to the egg white and desalting to the hydrolyzed egg white were evaluated. The membrane filtration technology was chosen to be used for desalting. The results showed that ultra filtration by the molecular mass cutoff 10000 was suitable for egg white, the desalinization rate was 83.81%, and the protein retention was 86.1%. And the nanofiltration and electro dialysis were suitable for hydrolyzed product, the desalinization rate and protein retention of nanofiltration could reach 94.3% and 84.27% using NF-90 at pH=3.2. The desalinization rate and protein retention of electro dialysis could reach 96.69% and 71.4% at pH=4.2.
本论文研究以本身利用价值不大的咸鸭蛋蛋清为原料,经过选择高效率的蛋白酶水解,制取分子量小于5000的富含白蛋白肽的生物活性肽产品,并利用膜过滤技术去除原有的以及水解过程中增加的盐分,使得产品中的盐含量大大降低,符合人体食用的要求。
本文首先研究了咸鸭蛋蛋清制取白蛋白肽的工艺及工艺优化。工艺优化的重点是酶解反应条件的优化。通过对酶种类、底物浓度、酶浓度、酶解体系的pH、酶解温度和酶解时间等进行单因素试验,以及随后的正交试验综合分析,得到了酶解反应的最佳条件:选用酸性蛋白酶、稀释倍数为1、酶用量为3000U/g、pH为2.5、反应温度为45℃、酶解时间为6h。
接着,本文又研究了脱盐方法的选择。根据对原料脱盐和水解以后对水解液脱盐的优缺点的分析,选择采用膜过滤技术对原料或水解液进行脱盐试验。利用超滤技术对咸鸭蛋蛋清直接脱盐,选用截留分子量为10000的超滤膜可使脱盐率达到83.81%,蛋白质截留率达到86.1%。利用纳滤技术和电渗析技术对水解液进行脱盐,当pH3.2时用NF-90的纳滤膜使脱盐率达到94.3%,蛋白质截留率达到84.27%。在pH4.2时利用电渗析使脱盐率达到96.69%,蛋白质截留率达到71.4%。
Bioactive peptide has a high nutrition of value and more functional activities. So it is a popular subject in nutrition now. The egg white of salted duck eggs contains abundance of high quality protein, but it can't be used directly, and is commonly discarded because of its high salt content. That induced serious environment problem and great waste of the high quality protein.This paper researched on the bioactive peptides made from salted duck eggs white which was rich of ovalbumin and was of little commercial value through hydrolyzing by proteases and subsequent membrane filtration to get rid of the extra salt so that the final product could be more useful for human and animals.Firstly, the technology of making ovalbumin peptides was optimized. The enzymatic reaction conditions such as enzyme species, substrate concentration, enzyme concentration, pH, temperature and time of reaction was studied. The optimal conditions of enzymatic treatment was obtained as follows: chose acid proteinase, dilution rate was 1, dosage of enzyme was 3000U/g, pH was 2.5, temperature was 45 ℃, and raction for 6hours.Then the desalting method of the final hydrolyzed product was compared and chosen. The advantage and disadvantage of desalting to the egg white and desalting to the hydrolyzed egg white were evaluated. The membrane filtration technology was chosen to be used for desalting. The results showed that ultra filtration by the molecular mass cutoff 10000 was suitable for egg white, the desalinization rate was 83.81%, and the protein retention was 86.1%. And the nanofiltration and electro dialysis were suitable for hydrolyzed product, the desalinization rate and protein retention of nanofiltration could reach 94.3% and 84.27% using NF-90 at pH=3.2. The desalinization rate and protein retention of electro dialysis could reach 96.69% and 71.4% at pH=4.2.
引文
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6. Emanuele S M. Gobbetti.Bioactive peptides in dairy products: synthesis and interaction weth proteolytic enzymes.Food Microbiology, 2000, 17: 129～141.
7. Cornelia L.etc.Purification of novel pepetide antibiotics from human milk[J].Journal of Chromatagraphy B, 2001, 752: 369～377.
8.张琦.生物活性肽制备的研究.畜牧兽医杂志,2003,22(3):20～21.
9. Nishimura T, Kato H. Taste of free amino acids and peptides. Food Rev Int, 1988 (4): 175～194
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12.张延坤,张东祥.生物活性肽的特殊保健功能及其食品应用.食品开发.2005(7)19
13.程远芳,宋代军.生物活性肽在饲料工业中的应用.中国饲料.2003(10):19～20
14.齐继成.我国生物活性肽开发应用前景.中国保健食品,2004(4):22～25.
15.王强,李大魁等.人血白蛋白临床应用的药物利用评价.中国药学杂志2005,40(13):1036～1038
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17. Newey H, Smyth D. H. lntracellular hydrolysis of dipeptides duringintestinal absorption. J. Physiol, 1960, 152: 367
18.夏前明,陆国庆,张彦等.人血白蛋白注射液致严重过敏反应.药物不良反应杂志.2005(3):219～220
19. Yoshinori M. The understanding of egg white protein functionality. Trends in food science & technology. 1995 (6): 225～232
20.王政腾,万添春.褐色菜鸭青白壳蛋之理化性质及其碱化过程比较.中国农业化学会志.1997,25(3):263～272
21. Tian, Hui-Guang. Dietary sodium and potassium, socioeconomic status and blood pressure in a Chinese population. Appetite. 1996, 26: 235～246
22. Goodle, F. The food industry fights for salt. British medical journal. 1996.(312): 1239～1240
23. Thelle, D.S. Salt and blood pressure revisited. British Mdeical Journal.1996, 312(704): 1240～1241
24. Antonios, T. F. T.;Macgregor, G.A. Salt-more adverse effects.Lancet.1996(348): 250～251
25.刘茂宏,张胜善.蛋白溶菌酵素对贡丸之保存效果.中国畜牧学会会志.1994(23):244～245
26.林庆文,姜延年.咸鸭蛋蛋白液之乳化性及其于法兰克福香肠之应用.食品科学.1996,23(2):244～254
27.王学松.膜分离现状及发展趋向.化学进展.1994,6(4):321～335
28.王继焕,刘启觉.膜技术及其在食品工业中的应用.武汉食品工业学院学报.1994(3):21～25
29.陈仪男,超滤技术在食品工业中的应用.漳州职业大学学报.2003(2):104～106
30.张文会,陈国海等.食品高新技术—超滤.西藏农业科技.2005,27(3):38～41
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32.陈少洲,陈芳.膜分离技术与食品加工.化学工业出版社.2005
33.张永忠,孙艳梅.纳滤膜及其在食品工业中应用.食品工业.2002(2):48～49
34.时钧,袁权等.膜技术手册.化学工业出版社.2001.
35.黄万抚,罗凯.电渗析技术应用研究进展.中国资源综合利用.2003(11):15～19
36.刘湘菊,刘国栋.用离子交换树脂脱除氨基酸与盐混合物中的盐.离子交换与吸附.2000,16(6):521～527
37.冯杰龙,林炜铁.生物活性肽及其蛋白酶水解法制备探索.广州食品工业科技.2002,18(3):36～38
38.宁正祥.食品成分分析手册.中国轻工业出版社.1998
39.Adler-N. Enzymatic hydrolysis of food proteins. Essex Elsevier applied science publishers LTD, 1986.
40.赵新淮,冯志彪.蛋白质水解物水解度的测定.食品科学.1994(11):65～67
41.吕艳.酶解小麦蛋白制取谷氨酰胺活性肽的研究.[学位论文].杭州:浙江大学食品系.2005
42.王永江,刘通讯.SPI热性性后SH基含量变化与其酶解关系研究.2005,26(1):86～88
43.孙崇荣,李玉民.蛋白质化学导论.上海:复旦大学出版社.1991
44.方林求,褚楠,薜静.鸡卵清蛋白对蛋白酶抑制作用的研究.中国生化药物杂志.1995,16(6):262～265
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2.史延平,孙永梅,王万东.生物活性肽——一个改变了动物氮代谢观念的新领域.丹东纺专学报,2002,9(2):1～3
3.张嫂,周光宏,徐幸莲.生物活性肽的生理活性和应用前景.食品研究与开发,2002,23(6):3～6
4.葛轶群,俞建瑛,宋聿文等.生物活性肽的研究进展.中国生化药物杂志.1998,19(6):404～406
5.曾晓波,王海英,林永成.食物中的生物活性肽:生物活性及研究进展.食品工业科技,2004.25(4):151～155.
6. Emanuele S M. Gobbetti.Bioactive peptides in dairy products: synthesis and interaction weth proteolytic enzymes.Food Microbiology, 2000, 17: 129～141.
7. Cornelia L.etc.Purification of novel pepetide antibiotics from human milk[J].Journal of Chromatagraphy B, 2001, 752: 369～377.
8.张琦.生物活性肽制备的研究.畜牧兽医杂志,2003,22(3):20～21.
9. Nishimura T, Kato H. Taste of free amino acids and peptides. Food Rev Int, 1988 (4): 175～194
10.唐传核,彭志英.功能性食品基料蛋白质及肽类开发现状.粮食与油脂,2001(1):39～41.
11.张琦.生物活性肽的制备的研究.畜牧兽医杂志.2003,22(2):20～21
12.张延坤,张东祥.生物活性肽的特殊保健功能及其食品应用.食品开发.2005(7)19
13.程远芳,宋代军.生物活性肽在饲料工业中的应用.中国饲料.2003(10):19～20
14.齐继成.我国生物活性肽开发应用前景.中国保健食品,2004(4):22～25.
15.王强,李大魁等.人血白蛋白临床应用的药物利用评价.中国药学杂志2005,40(13):1036～1038
16.陈栋梁,刘莉,于兰等.白蛋白多肽对核苷的促吸收及增强小鼠细胞免疫功能的作用.食品科学.2004,25(1):163～166
17. Newey H, Smyth D. H. lntracellular hydrolysis of dipeptides duringintestinal absorption. J. Physiol, 1960, 152: 367
18.夏前明,陆国庆,张彦等.人血白蛋白注射液致严重过敏反应.药物不良反应杂志.2005(3):219～220
19. Yoshinori M. The understanding of egg white protein functionality. Trends in food science & technology. 1995 (6): 225～232
20.王政腾,万添春.褐色菜鸭青白壳蛋之理化性质及其碱化过程比较.中国农业化学会志.1997,25(3):263～272
21. Tian, Hui-Guang. Dietary sodium and potassium, socioeconomic status and blood pressure in a Chinese population. Appetite. 1996, 26: 235～246
22. Goodle, F. The food industry fights for salt. British medical journal. 1996.(312): 1239～1240
23. Thelle, D.S. Salt and blood pressure revisited. British Mdeical Journal.1996, 312(704): 1240～1241
24. Antonios, T. F. T.;Macgregor, G.A. Salt-more adverse effects.Lancet.1996(348): 250～251
25.刘茂宏,张胜善.蛋白溶菌酵素对贡丸之保存效果.中国畜牧学会会志.1994(23):244～245
26.林庆文,姜延年.咸鸭蛋蛋白液之乳化性及其于法兰克福香肠之应用.食品科学.1996,23(2):244～254
27.王学松.膜分离现状及发展趋向.化学进展.1994,6(4):321～335
28.王继焕,刘启觉.膜技术及其在食品工业中的应用.武汉食品工业学院学报.1994(3):21～25
29.陈仪男,超滤技术在食品工业中的应用.漳州职业大学学报.2003(2):104～106
30.张文会,陈国海等.食品高新技术—超滤.西藏农业科技.2005,27(3):38～41
31.陈颖.超滤技术及其在乳品工业中的应用.黄牛杂志.2003,29(3):35～37
32.陈少洲,陈芳.膜分离技术与食品加工.化学工业出版社.2005
33.张永忠,孙艳梅.纳滤膜及其在食品工业中应用.食品工业.2002(2):48～49
34.时钧,袁权等.膜技术手册.化学工业出版社.2001.
35.黄万抚,罗凯.电渗析技术应用研究进展.中国资源综合利用.2003(11):15～19
36.刘湘菊,刘国栋.用离子交换树脂脱除氨基酸与盐混合物中的盐.离子交换与吸附.2000,16(6):521～527
37.冯杰龙,林炜铁.生物活性肽及其蛋白酶水解法制备探索.广州食品工业科技.2002,18(3):36～38
38.宁正祥.食品成分分析手册.中国轻工业出版社.1998
39.Adler-N. Enzymatic hydrolysis of food proteins. Essex Elsevier applied science publishers LTD, 1986.
40.赵新淮,冯志彪.蛋白质水解物水解度的测定.食品科学.1994(11):65～67
41.吕艳.酶解小麦蛋白制取谷氨酰胺活性肽的研究.[学位论文].杭州:浙江大学食品系.2005
42.王永江,刘通讯.SPI热性性后SH基含量变化与其酶解关系研究.2005,26(1):86～88
43.孙崇荣,李玉民.蛋白质化学导论.上海:复旦大学出版社.1991
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