Study On The Preparation Of Chitooligosaccharides By Enzymatic Hydrolysis,Biological Activity And Processing Of Functional Beverage

Chitosan is a product of chitin hydrolysate and has satisfactory biological activity.However,the large molecular weight of chitosan leads to poor water solubility,which makes the biological activity of chitosan unable to function better.Chitooligosaccharides is the product of chitosan degradation.Chitooligosaccharides have small molecular weight and good water solubility,while maintaining satisfactory biological activity.Chitooligosaccharide is an alkaline polysaccharide and has a variety of biological activities due to the chemical structure containing amino,hydroxyl and other groups,such as antibacterial,antioxidant,and hypoglycemic.In this study,chitooligosaccharides were prepared by enzymatic hydrolysis.The hypoglycemic activity and antibacterial activity of each monomer were separated and analyzed,and then a functional beverage was prepared.The main content is as follows:The activity of chitosanase was determined by 3,5-dinitrosalicylic acid method,and it was 102.53 U/g.The degree of deacetylation of chitosan was determined by acid-base titration,and it was 92.6%.The chitooligosaccharide mixture was prepared by three singlefactor experiments of substrate concentration,reaction temperature and reaction time,and the monomer composition of the chitooligosaccharide mixture was determined by thin-layer chromatography and MALDI-TOF.The optimal reaction conditions were: The substrate concentration was 1 g;the reaction temperature was 50 ℃;the reaction time was 8 h.Thinlayer chromatography analysis and MALDI-TOF showed that the chitooligosaccharide mixture under the reaction conditions contained three monomers: chitobiose,chitotriose and chitotetraose.Cation exchange resins are used to separate and purify chitobiose,chitotriose and chitotetraose.Thin-layer chromatography analysis and MALDI-TOF were used to determine the purity of chitobiose,chitotriose,and chitotetraose.The results showed that chitobiose,chitotriose and chitotetraose had high purity and no impurities,and the separation effect of cation exchange resin was excellent.NMR was used to detect the acetyl groups in the three chitooligosaccharide monomers,and the results showed that none of the three chitooligosaccharide monomers contained acetyl groups,indicating that they were highly deacetylated.The α-glucosidase inhibition test was used to measure and compare the hypoglycemic activities of the three chitooligosaccharide monomers.The results showed that all the three chitooligosaccharide monomers had hypoglycemic activity,and the hypoglycemic activity of chitotetraose was the best..The inhibition types of the three chitooligosaccharide monomers are all reversible competitive and non-competitive mixed inhibition.Salmonella was used to compare the antibacterial activities of three chitooligosaccharide monomers,and the results showed that chitotriose and chitotetraose had the antibacterial activity of Salmonella,and the antibacterial activity of chitotetraose was better than that of chitotriose.Orthogonal experiments were conducted to study the effects of amount of chitooligosaccharides,lemon juice,and xylitol and optimized them.The precipitation rate and stability coefficient of four types of stabilizers of agar,carrageenan,sodium carboxymethyl cellulose,and xanthan gum were studied,and stabilizers suitable for beverages were selected.Experimental results showed that the optimal process conditions were 7% chitooligosaccharide addition,16% lemon juice addition,and 12% xylitol addition.Carrageenan was most suitable for the beverage as a stabilizer,with a precipitation rate of0.3% and the highest stability coefficient.Beverages prepared under this condition had a harmonious taste and a refreshing lemon flavor.