Extraction,Separation,Purification And Safety Evaluation Of Amyloid Fiber-EGCG Hydrogels From Tea In Summer And Autumn

In this experiment,tea polyphenols were extracted by hot water extraction method,and the extraction time,extraction temperature and liquid-material ratio were used as single factor experiments.After the crude polyphenol was prepared under optimal conditions,the pure polyphenol was purified by polyamide resin.the prepared pure product polyphenols were used for the deter mination and molecular interaction of amyloid fibers.The results showed that there was a strong affinity between polyphenols and protein molecules,and the affinity became stronger with the increase of polyphenol concentration.On the basis of molecular interaction,the experiment of making hydrogel by polyphenol and protein amyloid fibrils was carried out.Compared with crude polyphenols,pure polyphenol formed gel with amyloid fibrils and formed stable and inverted hydrogel.the hydrogel was then micro-characterized,and the image of polyphenol attached to amyloid fiber was observed under transmission electron microscope,and the diameter of amyloid fiber was measured for different concentrations of polyphenol loading.The results showed that the high concentration of polyphenols increased the diameter of amyloid fiber,that is,the loading amount of amyloid fiber was increased.polyphenol-fiber complex also showed a better chemical antioxidant effect.Hydrogel has a broad application prospect,so finally,the safety of amyloid-fibre-EGCG hydrogel mice in gavage was systematically evaluated,mainly in the aspects of daily diet,drinking water,serum index,blood routine parameters,main organ coefficient,histopathological exa mination,liver enzyme activity and so on.The main findings are as follows:1.Extraction and purification of polyphenols from summer and autumn teaThe extraction time,the extraction temperature and the ratio of liquid to material will affect the extraction effect of tea polyphenols in summer and autumn.The extraction time range was deter mined to be 30-50 min,the extraction temperature range was 85-95℃ and the ratio of liquid to material was 10-20 mL/g.According to the single factor experiment,the optimum extraction conditions of tea polyphenols in hot water extraction were determined by using three factors and three levels experiment:extraction time was 35 minutes,extraction temperature was 95℃,liquid-to-material ratio was 16 mL/g,extracted twice,the content of tea polyphenols extracted under this condition was 123.73 mg gallic acid/g.2.HEWL of interaction between amyloid fibrils and polyphenolsThe binding kinetics of polyphenols to HEWL amyloid fibers was deter mined by ForteBio Octet system.when the polyphenol concentration was 3.125 μg/mL、6.25μg/mL、12.5 μg/mL、25 μg/mL、50μg/mL,the affinity of different concentrations of polyphenols to amyloid fibers was deter mined,and the changes of binding kinetics before and after polyphenol purification were compared.Amyloid fibers and polyphenols can form inverted hydrogels.the concentration range of for ming hydrogels was deter mined by pre-experiment.Crude polyphenols can form hydrogels between more than 0.125 mg/mL,less than 2 mg/mL.while purified polyphenols can form hydrogels between more than 0.0625 mg/mL,less than 8 mg/mL.The highest concentration is increased by about 4 times.The as-prepared pure product polyphenol-fiber complex was microcharacterized.after measurement,the pure fiber diameter was 5.835±0.905 nm,when the polyphenol concentration was 1 mg/mL,the fiber diameter reached 6.955 and 0.743.when the polyphenol concentration increased to 4 mg/mL,the fiber diameter reached 7.454±0.875 nm,and compared with the pure fiber,there were significant differences,which indicated that polyphenols interacted with the fiber and attached to the fiber.The polyphenol-fiber complex has a good antioxidant effect on free radical scavenging rate of 60-70%.3.Safety evaluation of HEWL amyloid fiber-EGCG hydrogelAnimal experiments designed amyloid fiber-EGCG hydrogel low,medium and high three experimental doses,with concentration gradient distribution,and set the corresponding dose of individual EGCG for comparison,at the same time set different fiber concentrations.The results showed that there was no oral toxicity to amyloid fibrils,Neither EGCG nor EGCG-amyloid fibrils showed toxicity.