Studies On Enzymatic Prepation Of Hohenbuehelia Serotina Antioxidant Protein And Stability

Hohenbuehelia serotina,the scientific name of the genus Hohenbuehelia serotina,alias yellow mushroom,frozen mushroom,is one of the famous wild edible fungi in Northeast China.Regular consumption of Hohenbuehelia serotina has the functions of enhancing immunity,anti-oxidation and anti-fatigue.In this study,the protein was extracted from Hohenbuehelia serotina,and Hohenbuehelia serotina protein was enzymatically hydrolyzed to obtain the polypeptide of Hohenbuehelia serotina.The dialysis method was used to divide the polypeptide into three different molecular sizes components.The amino acid composition and structure of the peptides were compared and analyzed,and the antioxidant capacity and stability of the three components were compared:(1)The crude protein of five kinds of edible fungi(Boletus edulis,Oyster mushroom,Fungus Inonotus,obliquus and Hohenbuehelia serotina),and mushroom was used as experimental material.The protein extraction rate of mushroom was extracted by four kinds method(Alkali extraction,Ultrasound-assisted alkali extraction,Tris-HCl extraction and Salt extraction).The highest protein extraction rate was obtained by ultrasonic assisted alkali method.Examining the isoelectric point of the protein of mushroom is pH 4.(2)Three single enzymes and two complex enzymes were used to hydrolyze the protein of mushroom.It was found that the degree of hydrolysis and DPPH·+scavenging rate were the highest after flavoring alone.Protease is the best protease for hydrolyzing the protein of mushroom.The results of Tricine SDS PAGE showed that the enzymatic hydrolysis method can effectively hydrolyze the macromolecular mushroom protein into small molecular peptide.The response surface method was used to optimize the proteolytic conditions of mushroom.Under the optimal conditions of hydrolysis,the degree of hydrolysis and DPPH·+scavenging rate are basically consistent with the theoretical values.(3)Using dialysis bag dialysis,the enzymatic hydrolysate of mushroom was divided into three components of different molecular size,P1(<3.5 kDa),P2(3.5 kDa-10 kDa),and P3(>10 kDa).The amino acid composition analysis showed that the amino acids of P1,P2 and P3 were abundant,but none contained cysteine(Cys).The infrared spectrum showed that the peak shapes and peak positions of the components P1,P2 and P3 were basically the same,amide I band and amide III band with protein infrared spectrum.the characteristic peak of the amide B band.(4)The results of antioxidant activity assay showed that DPPH·+,ABTS·+,and ·OH of component P3 had the strongest scavenging ability and relative reducing ability;component P2 inhibited lipid peroxidation strongest;The O2-·scavenging rate of three components of different molecular size is not significant;using the membership function method to rank the comprehensive antioxidant capacity of three differents molecular size of mushroom polypeptide,the comprehensive antioxidant capacity order:P3>P1>P2.(5)The effects of temperature,pH,food ingredients,metal ions,preservatives and simulated artificial gastrointestinal fluid digestion on the stability of the polypeptide were studied.The results showed that mushroom polypeptide is stable under neutral pH environment;the gastrointestinal digestion and food ingredients have no effect on mushroom polypeptide;the Cu2+,Zn2+ and Mg2+ in the metal ion have a great influence on the stability of the polypeptide;the heat of the component P3 stability and preservative stability are the best.This study shows that the protein content of the mushroom is high,the amino acid composition of the polypeptide after enzymatic hydrolysis is rich,has significant antioxidant capacity and good stability,and can be used for the development of functional foods.