Separation And Identification Of Polyphenols In Pomegranate Fruit And Its Antioxidant Activities

Polyphenols is the secondary metabolites in plant.Modern scientific research has confirmed that polyphenols have varieties of biological activities and have positive impacts on human health. Pomegranate(Punica grantum L.) blongs to Punica L.of Punicaceae,it has been cultured for more than 2000 years in China.China is rich in pomegranate resources and now it ranks first in the world with the planting area of 64668 hm².Pomegranate is not only rich in nutrients,but also rich in biological active materials,so it has a variety of effects on preventing and curing diseases. Polyphenols is the most abundant class of bioactive substances in pomegranate.In the background of that the disease prevention and cure functions of natural active components are being increasing concerned,study on the pomegranate polyphenols will not only deepen people's understanding of the value of pomegranate,but also lay basis for the rational development and use of pomegranate.In this study,pomegranate fruit as the research object,pomegranate juice(PJ) and pomegranate peel(PP),kernal residue(PR)-by-products in pomegranate product processing as raw materials,the separation and purifying methods,the compositions and contents and the antioxidant activities in vitro and in vivo of polyphenols in various parts of pomegranate had been researched.This is a relatively comprehensive and systematic study.The results are as follows:(1) The optimal extraction conditions of polyphenols from PP are as follows:extractant is 60% ethanol,temperature is 60℃,Solid-Liquid Ratio is 1:20,and extraction time is 2h,extracting twice. Using this process,the pomegranate peel polyphenols(PPPs) yield is 23.39%.Ultrasonic-assisted extraction conditions are:the best processing time is 30 min,extracting twice,with a polyphenols yield of 23.68%.(2) The best extraction conditions of polyphenols from PR are as follows:extractant is 50% ethanol,temperature is 60℃,Solid-Liquid Ratio is 1:20,and extraction time is 4h,extracting twice. Using this process,the pomegranate residue polyphenols(PRPs) yield is 0.69%.Ultrasonic-assisted extraction conditions are:the best processing time is 30 min,extracting twice,with a polyphenols yield of 0.79%.The drying method of raw materials use oven drying.(3) Selected an easy operated purifying and enriching method of pomegranate polyphenols(An organic solvent extraction).Using this method,the purifying multiples of PP,PJ and PR extracts respectively were 2.75,14.08 and 7.84,and the polyphenol contents of purification products respectively were 79.83%,10.56%and 23.76%.(4) Composition and content analysis showed that Pomegranate fruit is rich in polyphenols.PP has the most abundant content of polyphenols with total phenol content of 23.83%.Nine components had been identified in PPPs extracts,and the major components is pulicalagin with the content of 116.23 mg/g,followed by less contents of gallic acid,epicatechin,chlorogenic acid and so on.PJ has the largest components of polyphenols.The contents of total polyphenols in PJ is 0.11%.Eleven phenols components had been identified in PJ and pomegranate juice polyphenols(PJPs) extracts, and the components with higher content are pulicalagin,epicatechin,chlorogenic acid,catechin and caffeic acid,and so on.The contents of total polyphenols in PR is 0.83%.Ten components had been identified in PRPs extracts,and the components with higher content are pulicalagin,epicatechin, rutin,gallic acid,catechin,and so on.(5) In vitro tests,it showed that PPPs,PJPs and PRPs had high antioxidant and scavenging free radicals activities.The deoxidization capability order is:PRPs(EC₅₀ 53.75mg/L)＞PPPs(EC₅₀ 54.70mg/L)＞PJPs(EC₅₀ 59.45mg/L).The scavenging effects on superoxide anion radical order is: PRPs(IC₅₀ 14.66mg/L)＞PJPs(IC₅₀ 18.88mg/L)＞PPPs(IC₅₀ 22.41mg/L).The scavenging effects on hydroxyl radical order is:PJPs(IC₅₀ 308.70mg/L)＞PRPs(IC₅₀ 331.50mg/L)＞PPPs(IC₅₀ 350.31mg/L).The scavenging effects on DPPH radical order is:PJPs(IC₅₀ 3.89mg/L)＞PRPs(IC₅₀ 4.74mg/L)＞PPPs(IC₅₀ 4.74mg/L).The inhibitory effects on lipid peroxidation order is:PRPs(IC₅₀ 12.29 mg/L)＞PJPs(IC₅₀ 17.66 mg/L)＞PPPs(IC₅₀ 24.37 mg/L).Generally speaking,the antioxidant activities of PRPs and PJPs are stronger than PPPs.(6) In vivo tests,it showed that the PPPs can increase the activities of superoxide dismutase (SOD),catalase(CAT) and glutathione peroxidease(GSH-Px) and decrease the amount of malondial -dehyde(MDA)in blood,liver and brain tissue of mice,but will not have significant negative impacts on the body weight and organ indices of mice.These results indicated that pomegranate polyphenols had antioxidant activity in vivo,and the role of size was positively correlated with the dose.