Study On Extraction,Isolation,Purification,Structural Identification And Characterization Of Anthocyanins From Hibiscus Sabdariffa

Hibiscus Sabdariffa has a high medicinal value and edible value.For example,you can use the Hibiscus Sabdariffa calyx to make preserves,flower tea and so on;Hibiscus Sabdariffa anthocyanins have physiological effects beneficial to the body such as anti-oxidation,prevention of diabetes and anti-inflammatory Hibiscus Sabdariffa grows in subtropical regions with long flowering periods and high yields,and has attracted much attention in the food and pharmaceutical industries.The anthocyanins of Hibiscus Sabdariffa are susceptible to degradation by environmental factors,therefore,this study mainly explores the processand characteristics of anthocyanins in Hibiscus Sabdariffa,and provides experimental basis for the vigorous development of anthocyanins of Hibiscus Sabdariffa.?1?Using the Hibiscus Sabdariffa anthocyanins extraction amount as an evaluation index,the process parameters of extracting anthocyanins from Hibiscus Sabdariffa by ultrasonic microwave were optimized by response surface method.Optimal process parameter conditions:Ultrasonic power 350 W,extraction temperature 50?,microwave power 279 W,extraction time of 18 min,ethanol volume fraction 60%,liquid to material ratio 36:1?mL/g?.The extraction amount of anthocyanins from Hibiscus Sabdariffa was787.56 mg/100g by pH differential method.?2?Selecting four kinds of macroporous resins:AB-8,D101,NKA-II and HPD-100and screening the macroporous resins with the best purification effect on the anthocyanins of Hibiscus Sabdariffa by two evaluation indexes of static adsorption rate and desorption rate.According to the static purification experiment,the static adsorption rate of AB-8macroporous resin to Hibiscus Sabdariffa was 88.58%and the static desorption rate was90.37%,the experimental results showed that the purification effect of AB-8 macroporous resin was better than that of other three kinds of resins,so the subsequent dynamic purification experiment of AB-8 macroporous resin was selected.On the basis of dynamic adsorption and desorption experiment,the optimum dynamic adsorption conditions of the anthocyanins were determined by single factor experiment and response surface optimization design experiment:the upper sample liquid was pH2.5,the upper sample concentration was 3.0 mg/mL,the upper sample Velocity was 2.0 mL/min,and three groups of parallel experiments were carried out under these condition.The dynamic adsorption rate of the anthocyanin was 99.2%.The eluting solution was pH 2.0,the concentration of eluting solution was 80%,the elution velocity was 4.0 mL/min,and three groups of parallel experiments were designed underthe same conditions,and the desorption rate of the anthocyanins was 92.15%.The color price of anthocyanins increased from 2.35 to 30.2.?3?The Sephadex LH-20 dextran Gel was selected to further isolate and purify the anthocyanins of the Hibiscus Sabdariffa after the purification of AB-8 macroporous resin.To investigate the effect of upper sample concentration,elution concentration and elution velocity on the separation and purification of anthocyanins from Hibiscus Sabdariffa,the optimum process conditions for separation and purification were determined by measuring the ultraviolet absorbance values of the collected components:the upper sample concentration was 20 mg/mL,the elution concentration was 50%methanol?pH 3?,the elution velocity was 0.3 mL/min,under these conditions,the anthocyanins of the Hibiscus Sabdariffa has the best separation effect,and a component peak is collected.The anthocyanin components of the purified Hibiscus Sabdariffa were preliminarily identified by HPLC-MS and mass spectrometry information.The results showed that there were two kinds of anthocyanins in the peak of a component of the anthocyanins of the Hibiscus Sabdariffa,which were Delphinidin-3-sambubioside and Cyanidin-3-sambubioside.?4?The stability of the anthocyanins of Hibiscus Sabdariffa was evaluated by taking the absorbance and L*,a*and b*values of the anthocyanins in the Hibiscus Sabdariffa as indexes.The experimental results show that the anthocyanins of Hibiscus Sabdariffa can be well preserved at a low temperature of 4^oC,and can maintain the red color characteristics of the anthocyanins itself.pH has a great influence on the stability of the anthocyanins,and under acidic conditions,the anthocyanins of the Hibiscus Sabdariffa are very stable and the color remains red.Under alkaline conditions,anthocyanins is less stable and the original color characteristics of the anthocyanins of the Hibiscus Sabdariffa are changed.Four kinds of ions of Na⁺,K⁺,Mg²⁺and Ca²⁺have a certain auxiliary color effect on the Hibiscus Sabdariffa anthocyanins.Three kinds of ions of Fe²⁺,Cu²⁺and Al³⁺have a certain destructive effect on the stability of the anthocyanins of the Hibiscus Sabdariffa,so avoid contact with them in practical application.?5?Determination of the antioxidant activity of anthocyanins in Hibiscus Sabdariffa by DPPH method,·OH scavenging ability and total reduction ability.The experimental results show that the scavenging ability of purified anthocyanins and Vc to DPPH free radicals is comparable within a certain range but the scavenging ability of DPPH free radicals is significantly higher than that of extract anthocyanins.Under high concentration conditions,the anthocyanins can effectively remove.OH free radicals but weaker than Vc removal ability;Hibiscus Sabdariffa anthocyanins have a certain total reduction ability and the total reduction capacity is Vc>purified anthocyanins>extract anthocyanins.