| In this thesis,preparation of polypeptide from tilapia meat by enzymatic hydrolysis was studied.The optimum hydrolysis conditions of papain and neutral protease were optimized.Then the refining processes of hydrolysis product with inorganic ceramic membrane microfiltration and macroporous resin adsorption were discussed.And the physicochemical properties of hydrolysis product before and after the refining process were studied.The aim was to explore new ways for the use of tilapia meat,to improve the fish flavor of protease hydrolysis product and to provide theoretical guidance for the preparation of bioactive peptide.The main findings are as follows:1.Taking the degree of hydrolysis,soluble protein content,the nitrogen utilization rate and molecular weight distribution as indexes,the optimum conditions of tilapia meat by papain enzyme were optimized through L25(56)orthogonal experiments: temperature 50?,time 5h,enzyme dose 850 u/g,the ratio of material to solvent 1:2.5,natural p H.On this condition the degree of hydrolysis,soluble protein content and nitrogen recovery of hydrolysis product were 14.10%,9.84 g/100 g,88.93%,respectively.The relative molecular mass of less than 1 k Da and less than 5 k Da were 36.53% and 80.98%.And the optimum process conditions of tilapia meat by neutral protein enzyme : temperature 50?,time 5h,enzyme dose 1000 u/g,the ratio of material to solvent 1:2.5,natural p H.Under this condition the degree of hydrolysis,soluble protein content and nitrogen recovery of hydrolysis product were 16.75%,10.20 g/100 g,83.48%,respectively.The relative molecular mass of less than 1 k Da and less than 5k Da were 62.34% and 97.17%.While,on the basis of orthogonal experiment,combining with the nitrogen utilization rate and molecular weight distribution,the optimum process conditions of tilapia meat by neutral protease and papain compound enzyme: temperature 50?,time 5h,enzyme dose of neutral protease and papain 500 u/g and 425 u/g,the ratio of material to solvent 1:2.5,natural p H.Results showed that the degree of hydrolysis was 17.24%,soluble protein content was 10.74 g/100 g,nitrogen recovery rate was 94.15%,the relative molecular mass of less than 1 k Da and less than 5k Da were 64.18% and 97.66%,respectively.2.To discuss the hydrolysis product refining process,firstly,the ceramic membrane flux of 200 nm and 50 nm,physical and chemical properties of permeate liquid with the change of micro filtration time and membrane cleaning condition were investigated.In condition of TBM,circular flow of 20 L/min and transmembrane pressure difference of 0.06 MPa,after 11 min,the membrane flux of 200 nm and 50 nm were stable and were 91.0 L/(m2·h)and 71.0 L/(m2·h),respectively.Soluble protein content of permeate liquid was stable after 10 min,nitrogen recovery rate was 87.58% and 85.84%.Ceramic membrane can significantly reduce the color difference value of enzymolysis liquid and effect of 50 nm was better than 200 nm.Using 0.5% HNO3 and 0.5% Na OH to clean step by step,the recovery of ceramic membrane flux of 200 nm and 50 nm were 92.56% and 88.75%,respectively.Secondly,to discuss the effect of ethanol elution,the macroporous resin AB-8 and DA201-C?dynamic adsorption of enzymolysis liquid ceramic membrane micro filtrate through adsorption curve and curve were observed.The best sample volume of micro filtrate were 7.5BV and 9.5BV,respectively,the maximum adsorption capacity were 54.23 mg/m L and 33.64 mg/m L.Best elution solution: AB-8 eluent was 60% ethanol and elution velocity was 3 BV/h,DA201-CI eluent was 40% ethanol and elution velocity was 3 BV/h.3.Physical and chemical properties of before(enzymolysis polypeptide powder)and after(microfiltration polypeptide powder,adsorption polypeptide powder and elution polypeptide powder)the refined products were analyzed.The basic component content of microfiltration polypeptide powder and enzymolysis polypeptide powder were nearly same.The protein loss rate of AB-8 adsorption polypeptide powder and elution polypeptide powder were 1.99% and 3.07%,respectively.And ash content and fat content were all increased.Ceramic membrane microfiltration and macroporous resin adsorption had good effect on decolorization.Results showed that ceramic membrane microfiltration of enzymolysis polypeptide powder had a little influence on molecular weight distribution.The 2-5 k Da peptides proportion of AB-8 adsorption polypeptide powder was reduced by 23.17% than that of enzymolysis polypeptide powder,while in the elution polypeptide powder increased by 41.83%.Ceramic membrane microfiltration can enrich the lysine.AB-8 adsorption induced the decline of the content of the total amino acids,essential amino acids,bitter amino acids and hydrophobic amino acid,but increase in flavor amino acids and sweet amino acid.For peptide adsorption ability was stronger,free amino acid adsorption was less,amino acid adsorption was selective.The results of reverse phase liquid phase showed elution polypeptide powder contained more hydrophobic peptides than enzymolysis polypeptide powder,microfiltration polypeptide powder and adsorption polypeptide powder.The HS-SPME/GC-MS was used to analyze volatile components of each peptide powder and found volatile component species of enzymolysis polypeptide powder,microfiltration polypeptide powder,adsorption polypeptide powder and elution polypeptide powder were 26,24,34 and 40,respectively.And each of them had high lipid components.Ceramic membrane microfiltration could remove nitrogen and partial ketone,and ketone content reduced by 40.90%.Macroporous resin mainly adsorbed esters and acids,and reduced by 13.62% and 13.62%,respectively. |
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