Research On The Extraction,Purification And Antioxidant Activity Of Anthocyanin From Black Chokeberry

Black chokeberry is rich in anthocyanins and can be used as a new source of natural pigments.In order to improve the utilization of black chokeberry,the enzymatic hydrolysis-solvent extraction technology was used to extract the anthocyanins and the extraction conditions were optimized.AB-8 macroporous resin was applied to isolate and purify anthocyanins,and optimiontal purification process was determined through static and dynamic adsorption and desorption experiments.Evaluating antioxidant activity of anthocyanins by measuring the capacity of anthocyanins on DPPH free radicals and hydroxyl radical scavenging,.The H2O2-induced HepG2 cell oxidative injury model was established to investigate the protective effect and mechanism of anthocyanin on oxidative damage of cells.The main results were as follows:1.The optimal conditions for cellulose enzymatic-ethanol extraction were determined by single factor and response surface design:the enzyme dosage was 0.03%,the enzyme treatment temperature was 51.76℃,and the enzyme treatment time was 1.29 h.Under this condition,the yield of anthocyanin extraction was 3.16 mg/g,which was increased by 45.24%compared with the simple alcohol extraction method,indicating that cellulase could significantly increase the extraction amount of anthocyanins.2.The crude anthocyanin was isolated and purified by using AB-8 macroporous resin.The static-dynamic adsorption and desorption conditions were studied to determine the best purification conditions:sample concentration was 10 mg/mL,the pH of sample was 2.0.The flow rate was 1.5 mg/mL,the sample volume was 12 BV,the amount of water was 7 BV,the ethanol concentration was 90%,and the amount of ethanol required for elution was 2 BV.Under this condition,the purified anthocyanin was purple-black powder,and its content was 80.8%,which was about 6 times higher than that before purification.The anthocyanin color value after purification was 252.7,which was increased by about 15 times.3.The anthocyanins of black chokeberry had good antioxidative activity,and the DPPH free radical and hydroxyl free scavenging capacity increased with the increase of concentration.IC50 was 0.009 mg/mL and 0.444 mg/mL,respectively.4.The anthocyanins of the black chokeberry have protective effects on oxidative damage of HepG2 cells.The conditions for establishing the oxidative damage model of HepG2 cells were:H2O2 treatment at 210 μmol/L for 4 h,the survival rate of the model group cells was 67.92%.The low-,medium-and high-density(20 μg/mL,100 μg/mL,and 200 μg/mL)anthocyanins were used to pre-protect cells and the survival rates were increased to 79.34%,94.55%and 87.72%,respectively.In addition,anthocyanin could significantly decrease the activity of LDH and the content of MDA in the culture medium,and increase the level of SOD in the cells(p<0.05).The result of flow cytometry showed that the chokeberry anthocyanin could reduce the early apoptosis rate of oxidative damage cells from 9.74%to 0.06%-4.41%.