Preparation Of Scallop Oligopeptides With High Fisher Ratio And Study On Its Effect In Vivo And In Vitro Of Anti-alcoholism And Liver Protection Combined With Traditional Chinese Medicine

The liver damage caused by long-term excessive drinking is called alcoholic liver disease,which endangers life and health seriously.It is of great significance to look for natural and safe biological active substances with the function of relieving alcohol and protecting liver.High Fischer ratio oligopeptides and Chinese herbal medicines such as Pueraria,Flos Puerariae,Hovenia dulcis thunb,turmeric and acanthopanax senticosus all have certain anti-alcoholism effects.Therefore,in this paper,the key technologies of enzymatic hydrolysis and aromatic amino acid removal for preparing high Fischer ratio oligopeptides from shellfish by-products were studied,the protective effects of Chinese herbal medicines and high Fischer ratio oligopeptides on acute alcoholic liver injury were studied and analyzed:(1)The protein,water content and amino acid composition of 9 raw materials were determined and analyzed,and the Fischer ratio was calculated.The by-products of processing of chlamys farreri were selected as the raw materials for the preparation of high Fischer ratio oligopeptides The optimal conditions for enzymatic hydrolysis of three combinations of enzymes(pepsin and flavoring protease,complex protease and flavoring protease,alkaline protease and flavoring protease)were optimized,the highest degree of hydrolysis of the three combinations was 60.14%,45.75% and 46.84%respectively,and the combination of pepsin and flavor protease had the highest degree of hydrolysis,the optimal conditions were as follows: pepsin activity 1000 U/g,temperature 42 °C,p H 2.5,time 6 h,flavor protease activity 1000 U/g,temperature55 °C,p H 7.0,time 7 h.In order to improve the hydrolysis efficiency,the raw materials were treated by microwave,ultrasonic and boiling water bath,and the degree of hydrolysis was increased most at 30 min of ultrasonic treatment.(2)The enzymatic hydrolysates from the by-products of scallop chlamys farreri were used as raw materials to remove aromatic amino acid,and the types of activated carbon were screened,the optimum adsorption conditions were as follows: solid-liquid ratio 1:10,p H 6.0,temperature 35 °C,adsorption time 2 h.After adsorption,the high Fischer ratio oligopeptide solution reached 33.22,which met the preparation requirements(Fischer ratio>20),high Fischer ratio oligopeptide solution,a by-product of the processing of chlamys farreri,was successfully prepared.(3)The molecular weight of the oligopeptides with high Fischer ratio was determined and their structures were characterized.The molecular weight of the oligopeptides with high Fischer ratio was less than 1000 Da,and more than 85% of the oligopeptides accorded with the requirement of the molecular weight range of the oligopeptides.The UV spectra of high Fischer ratio oligopeptides showed no obvious absorption peak near the characteristic absorption peak of 280 nm,which indicated that the high Fischer ratio oligopeptides had lower aromatic amino acid content in the aromatic amino acid.The IR spectra of high Fischer ratio oligopeptides show the characteristic absorption peaks in the characteristic bands(amide a band,amide I band,amide III band).(4)The activity of ADH and SOD of 5 kinds of Chinese herbal medicines which have potential for relieving alcoholism and protecting liver were tested in vitro.It was found that the activities of ADH and SOD of Flos Puerariae were 23.17% and 34.38%,respectively,the combination ratio of Flos Puerariae and high Fischer ratio oligopeptide was 1:2.By establishing an animal model of acute alcoholic liver injury in mice,the protective effects of Flos Puerariae,high Fischer ratio oligopeptide,high Fischer ratio oligopeptide and Flos Puerariae compound(1:2)on alcoholic liver injury were compared and analyzed,the results showed that the high Fischer ratio oligopeptide group,Flos Puerariae group and compound group could prolong the drunk time,shorten the sober time,increase the activity of ADH and ALDH in mice liver,accelerate the metabolism of ethanol;The activities of ALT and AST in serum,the activities of GSH-Px,SOD and CAT in liver and the content of MDA in serum were significantly decreased.Compared with high Fischer ratio oligopeptide and Flos Puerariae group,the effect of alcohol detoxification and liver protection in the compound group was more significant.In summary,on the basis of screening out the by-products from the processing of chlamys farreri as raw materials for the preparation of high Fischer ratio oligopeptides,high Fischer ratio oligopeptides with Fischer ratio greater than 20 were successfully prepared by pepsin and flavor protease two-step enzymatic hydrolysis and activated carbon dearomatization.The combination of high Fischer ratio oligopeptide and Flos Puerariae has a good protective effect on acute alcoholic liver injury in mice,which provides a theoretical support for the prevention and treatment of alcoholic liver disease.