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秋刀鱼制备黄嘌呤氧化酶抑制肽的工艺优化

Technology optimization on preparation of XOD inhibition peptide from Saury

  • 摘要: 为探讨秋刀鱼制备黄嘌呤氧化酶(XOD,xanthine oxidase)抑制肽的最佳工艺,该文以氮回收率和体外XOD抑制活性为指标,首先通过单因素试验确定中性蛋白酶和胰酶为水解秋刀鱼制备XOD抑制肽的最佳蛋白酶。在此基础上,采用响应面分析法研究加酶量、酶解时间和中性蛋白酶所占比例对酶解氮回收率及酶解产物XOD抑制活性的影响,进一步优化并最终确定了水解秋刀鱼制备XOD抑制肽的最优工艺:料液比1∶2 (g/g),总加酶量0.3%(中性蛋白酶∶胰酶质量比=6∶4),在pH值7.0和55℃条件下酶解6 h,其理论氮回收率和酶解产物XOD抑制率分别为72.69%和30.32%,对应实际值分别为72.03%和30.96%,预测模型可靠性高,可用于秋刀鱼XOD抑制肽的酶法制备,为工业化利用秋刀鱼生产降尿酸肽提供一定的理论和技术指导。

     

    Abstract: Abstract: During the metabolism in human body, the purines from food, nucleic acid and adenosine triphosphate (ATP) would all turn into hypoxanthine, and then the hypoxanthine would change into uric acid treated by xanthine oxidase (XOD). Thus the XOD plays a key role during the whole process. In addition, it has been reported that the XOD can not only combine with hypoxanthine, but also do with pyrazolopyrimidines, purines, phenylprazole derivatives, flavones and so on. Therefore, uric acid can be decreased by combining heterocyclic compounds with XOD. Recently, the western medicines have been widely used in the treatment of the high uric acid and gout, but some side effects from these western medicines may appear on human body. Allopurinol is the only drug in the medicine market which reduces the production of uric acid through the inhibition of XOD, but it is also proved to have side effects on liver and kidney. With the development of the enzymatic hydrolysis technology, more and more bioactive peptides have been prepared and studied for drug use or health care. The small peptides have many advantages, for example, they are safe without any side effects and can be absorbed more quickly. The saury is mostly used to only produce can and dry foods due to the backward science and technology, which doesn't fully tap its potential economic and nutritional value. In fact, the saury is very rich in protein and it might have many special functions due to the hard environment in the sea. Therefore, the saury might be a perfect raw protein material for the preparation of the urate-lowering peptide. This can bring not only better nutrition but also higher economic value. In the present paper, the XOD inhibition peptide was prepared by hydrolyzing saury. By detecting the protein recovery and inhibition activity of XOD of different hydrolysates, the neutral protease and pancreatin were proved the best protease through the single factor experiment. Then the effects of protease concentration, hydrolysis time and ratio of neutral protease on protein recovery and inhibition activity of XOD were studied by using response surface method to improve the preparing technology. Results showed that protease concentration and ratio of neutral protease had a significant influence on the protein recovery (P<0.001), while protease concentration and hydrolysis time had a significant effect on inhibition activity of XOD (P<0.001). The best technology of preparing the XOD inhibition peptides from saury was optimized based on the research results above. The best process parameters were as follows: the ratio of liquor to material was set to 1:2, the total protease concentration was 0.3% (the pancreatin accounted for 40% and the neutral protease accounted for 60%), the pH value of the enzymatic hydrolysis system was 7.0, the temperature was 55℃, and the hydrolysis time was 6 h. The theoretical protein recovery rate could reach 72.69%, and the theoretical inhibition activity of the XOD was 30.32%. The established model was of high reliability, and it could be used to prepare the XOD inhibition peptide from saury hydrolysate. The test of molecular weight distribution indicated that saury hydrolysate mainly consisted of the peptide fragments smaller than 3 kDa (76.35%). The paper indicated that the saury hydrolysate with XOD inhibition peptide was a potential material for decreasing uric acid in food industry.

     

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