动物营养学报2018，Vol.30Issue(1)：262-273,12.DOI:10.3969/j.issn.1006-267x.2018.01.032
单宁酶水解石榴皮多酚的工艺优化及酶解产物鉴定
Process Optimization of Hydrolysis of Pomegranate Peel Polyphenols by Tannase and Hydrolysates Identification
摘要
Abstract
Polyphenols from pomegranate peel has multiple biological activities such as anti-inflammatory, an-tioxidant and anti-cancer activities. However,the high molecular ellagitannins such as punicalagin in pomegran-ate peel polyphenols can not be absorbed by animal gastrointestine directly, but have to be hydrolyzed into small molecules such as ellagic acid first and then be absorbed into bloodstream and exerts physiological effects. This study was conducted to hydrolyze the high molecule tannins(relative molecular mass>500)in pomegran-ate peel polyphenols into small molecule phenolic acids by using tannase, optimize the hydrolysis process and hydrolysates purification process,and clarify the composition changes before and after the hydrolysis. By doing this, we hope it could provide basic data for the exploration of pomegranate peel polyphenols as functional feed additives promoting animal health. By comparing the high performance liquid chromatography fingerprints be-fore and after the hydrolysis, we determined ellagic acid increasing rate and punicalagin degradation rate as in-dicators of enzymolysis degree. Using the single factor design and orthogonal combination design, we opti-mized the enzymolysis conditions,and established the macroporous resin purification conditions of hydrolysates by using the methods of static adsorption and dynamic adsorption. Moreover, the composition changes before and after hydrolysis of pomegranate peel polyphenols were determined by a liquid chromatography-mass spec-trometry. The results showed that the optimum reaction conditions for the enzymolysis were pH 5.2,45℃,en-zyme volume 30 mL/g substrate,substrate concentration 4 g/L,16 h. For enzymatic products purification pa-rameters, D101 macroporous resin was tested to be the best absorbing material,the best elution was 95% etha-nol,the best loading volume was 300 mL,and the best collecting volume of elution was 120 mL. The contents of punicalagin(relative molecular mass was 1 084)and ellagic acid(relative molecular mass was 302)in the pomegranate peel polyphenols were 43.64% and 4.85%, respectively. Under the optimal enzymatic hydrolysis conditions and the optimal purification conditions,the enzymatic products of pomegranate peel polyphenols was obtained, the punicalagin content of which was 0.00%, and the ellagic acid content of which was 45.73%. There were seven kinds of main polyphenols in the pomegranate peel polyphenols had been determined, the mass-to-charge ratio(m/z)of them were 1 083,1 083,783,757,633,401 and 300,respectively,and the relative molecular mass was from 302 to 1 084. There were four kinds of main polyphenols in the enzymatic products pomegranate peel polyphenols had been determined, the m/z of them were 600,300,291 and 247, respectively, and the relative molecular mass was from 248 to 602. This experiment optimized the reaction con-ditions of tannase hydrolyzing pomegranate peel polyphenols and the hydrolysates purification conditions of the enzymatic products of pomegranate peel polyphenols, obtained hydrolysates rich in small molecular polyphe-nols, and this work could provide basic data for the development and utilization of pomegranate peel polyphe-nols as new functional animal feed additives.关键词
石榴皮多酚/单宁酶/酶解/工艺优化Key words
pomegranate peel polyphenols/tannase/enzymolysis/process optimization分类
农业科技引用本文复制引用
孙雨晴,陶新,徐子伟..单宁酶水解石榴皮多酚的工艺优化及酶解产物鉴定[J].动物营养学报,2018,30(1):262-273,12.基金项目
浙江省公益技术研究项目(LGN18C170004) （LGN18C170004）
浙江省重点研发计划项目(2018C02035,2018C02044) （2018C02035,2018C02044）
现代农业产业技术体系建设专项(CARS-36) （CARS-36）
浙江省农业(畜禽)新品种选育重大科技专项(2016C02054) （畜禽）
