Study On The Isolation, Purification And Antioxidant Activity Of Longan Seed Polyphenols

There are many active polyphenols in the longan seed,but they have not been fully developed and utilized.As natural active ingredients,polyphenols have good antioxidant properties.Cell metabolism,external stimuli,unhealthy eating habits and life habits can produce free radicals.Polyphenols can reduce the incidence of disease and delay aging by reducing the content of free radicals.In this paper,the main polyphenols in the longan seeds were obtained by various separation and purification methods and then analyzed by UPLC-MS for structures and contents.To expand the application of longan seed polyphenols in cosmetics,their antioxidant properties were evaluated by chemical methods and cell methods that use skin fibroblasts as a model.The main steps were as follows:Firstly,the polyphenols in the longan seeds were extracted by ultrasound,and the optimal extraction conditions were optimized by single factor experiment.The optimal extraction conditions were as follows:ethanol volume fraction of 60%,material to liquid ratio of 1/25?g/mL?,extraction temperature as 70?and sonication time as 60 min.Under these conditions,the extraction amount of longan seed polyphenols was the highest,reaching 53.68mg/g.Finally,the yield was 12.08%.Secondly,the crude polyphenols of longan seeds were extracted by chloroform,ethyl acetate and n-butanol,respectively.The results showed that the ethyl acetate phase enriched main polyphenols.Its polyphenol content was 663.50 mg/g.The antioxidant activity of the ethyl acetate phase was found to have the highest antioxidant capacity.The IC₅₀ of ABTS·and OH·was 69.11?g/mL and 73.83±6.02?g/mL,respectively,which was stronger than V_C.The IC₅₀ of the DPPH·was 21.79±0.60?g/mL;which was close to Vc.The cytotoxicity assay on human skin fibroblasts revealed that the crude polyphenols and the other three extract phases were not cytotoxic at the concentration of less than 5?g/mL.The antioxidant activity of the ethyl acetate phase was found to be the strongest by cell antioxidant experiments.At the concentration of 5?g/mL,the CAA unit was 57.71±5.19.After 600?mol/L H₂O₂ damaged,the intracellular SOD activity was increased from 0.75U/mL to 2.13 U/mL,and the GSH content from 9.93?mol/L to 16.78?mol/L.Finally,the ethyl acetate phase was subjected to further purification,structural identification and performance determination.The AB-8 macroporous resin was selected by screening six different types of macroporous adsorbent resins,and the resin elution conditions were optimized.The optimum elution conditions of AB-8 macroporous resin were as follows:the loading concentration of 1.5 mg/mL,the pH of 3.5,the elution solvent of 60%ethanol,and the elution flow rate of 0.6 BV/h.It was further purified by Sephadex LH-20 resin.The results showed that component 4 predominanted in the polyphenols,which was 757.75 mg/g.Qualitative and quantitative analysis by UPLC-MS showed that the main polyphenol of component 4 was corilagin,and the content was 631.18 mg/g.The chemical antioxidant experiments showed that the IC₅₀ of DPPH·,OH·and ABTS·of component 4 was 17.92?g/mL,65.10?g/mL and 49.73?g/mL,respectively.The reduction ability and FRAP value of100?g/mL were 0.600±0.01 and 1344.31±12.93,respectively.This is similar to the results of corilagin.The results of cell antioxidant experiments showed that the cytotoxicity of component 4 was lower,and the cell viability was 88.89±1.90%at 5?g/mL.After H₂O₂damaged cells,the protective effect of component 4 was stronger than that of corilagin.The cell viability of 1?g/mL were 72.55±2.34%and 58.62±3.13%,respectively.The clearing intracellular ROS and increasing SOD activity and GSH content effect of component 4 was similar to that of corilagin.