Analysis On Amino Acids Of Enzymatic Hydrolysis Of Crab And Optimization On Hydrolysis Conditions

Eriocheir sinensis（Crab）plays an important role in farming industry and food culturein China with its nutritious and pleasant aroma. At present, the processing of crab is limitedand the price of products is still lower relatively. The remaining crab legs, low-value crab,chitin, and other waste，they not only polluted the environment also caused the proteinwaste. Tuohu crab as raw materials used in this paper, the optimum conditions wereanalyzed which were from protein enzymatic hydrolysis by complex enzyme, and wewould find the correlation between taste compounds and hydrolysis degree (DH) with thehelp of High Performance Liquid Chromatography; After a series of detection andanalyzing of hydrolysis molecular weight distribution using Gel Chromatography. Then, akind of crab-extraction product was manufactured with full of flavor, nutrient-rich inproportion. In this paper, response surface method on crab hydrolysis process wasoptimized, the specific contents are as follows:First，In this paper, the hydrolysis effects were researched respectively on theappropriate hydrolysis conditions by comparing different enzymes such as Papain, neutralprotease, complex flavor enzyme, and two types enzyme flavor enzyme and Papainenzyme compound were selected using by degree of hydrolysis. Besides, time add flavorenzyme was1/4total hydrolysis time when DH value reached the maximum.Second, On the basis of probably hydrolysis conditions, pH, hydrolysis temperature,enzyme dosage and hydrolysis time4factors3levels were designed orthogonal test. Theresults of the total model design analysis showed that the factors that affect the degree ofhydrolysis were: pH> enzyme dosage> hydrolysis time> temperature. Summing up rangeanalysis and variance analysis, the preliminary results showed that better conditions forhydrolysis reaction were A1B1C1D2, that was pH5.5, temperature45℃, enzyme dosage4000U, hydrolysis time was5h.Third, Amino acids component from of the enzyme-hydrolyzate of the frozen crabwere determined by RP-HPLC using pre-column derivatisation witho-phthaldialdehyde(OPA), And glutamate injection was repeated five times on the testmethod precision which were calculated. Results indicated that amino acids was0.78%RSD retention time, peak area RSD was4.54%, indicating the method had good stableprecision, which was of determination on crab Protein Hydrolysate15kinds of amino acidsand reliable. During the results of hydrolyzate of the OPA derivatization method, glutamic acid content reached827μg/g, accounting for3.80%of Total amino acids. Flavorcharacteristics of substances mainly Glutamic and Aspartic which accounted for about7%of the total. Crab enzyme-hydrolyzate with higher levels of free amino acids, Essentialamino acids accounted for34.7%-46.8%of Total amino acids, which can be seen, crab notonly had the delicious flavor but also had a comprehensive and balanced amino acidcomposition, it，s also a good source of nutritious food.Fourth, This test discussed the total correlation analysis with flavor amino acids(Aspartic, Glutamic), Essential amino acids, Total amino acids and the degree ofhydrolysis using partial correlation analysis in SAS for data processing. The resultsshowed that fresh, Essential, Total amino acids make a positive correlation between thedegree of hydrolysis. Also there was significantly correlation between amino acids and thedegree of hydrolysis. As DH increasing, the total amount of amino acids and essentialamino acids also increased, but in comparison, the amino acid content with fresh featuresshowed much more significant growth. This also showed that the degree of hydrolysiscould be regarded as an important indicator of optimization hydrolysis process andmeasuring flavor level.Fifth, Molecular weight was analyzed by column chromatography. Elution orderbased on the degree of hydrolysis were as follows:12.71%,14.01%,15.06%,18.01%,20.85%,21.09%, the result shows with the hydrolysis degree increasing, high molecularweight peak gradually decreased and there was a gradual afterward by time trends; the lowmolecular weight peak increased after the shift.Sixth, On the basis of one-factor test, the method of response surface analysis combinedwith SAS data analysis software was adopted to built a quadratic regression equation withdegree of hydrolysis(DH) as the response value. Optimized by response surface analysis,the optimum hydrolysis conditions were obtained as follows: pH5, hydrolysis temperature55°C, enzyme dosage5500U and hydrolysis time6h. By repeating the experiment,variance and reliability analysis showed that the degree of model fit better.