Study On Physicochemical And Functional Properties Of Tree Peony Seed Protein And Antioxidant Activity Of Polypeptides

Tree peony seed protein（TPSP）is a kind of plant protein with the abundant of amino acids composition.The study and development of TPSP not only relieve the current lack situation of plant protein but also enrich the resource of plant protein in China.In this paper,the extraction and separation,physicochemical and functional properties of TPSP were studied using the tree peony seed as raw material.Meanwhile,the preparation process of tree peony seed polypeptide was optimized,and its antioxidant properties were explored.The results of the experiment were as follows.（1）TPSP was prepared from defatted tree peony seed meal according to the traditional alkaline-soluble-acid precipitation method.The effect of four single factors,extraction time,extraction pH,extraction temperature and solid/liquid ratio,on the extraction rate of TPSP was studied.Based on the single factor experiment,optimum extraction conditions of TPSP were 3.5 h,pH 9.50,60oC,and 1:10 g/mL of solid/liquid ratio with the maximum extraction rate（42.74%）using response surface methodology.（2）The physicochemical properties of TPSP were studied in the experiment.The results showed that TPSP with a favorable amino acid profile was composed of a 60kDa glycoprotein with two subunits of 38 and 23 kDa.The isoelectric points of the two subunits were 3.6 and 9.0.Moreover,2-D and diagonal electrophoresis data indicated the 38 kDa subunit included three types,which two types had inter-disulphide bonds and one type had no-disulphide bonds.So did the 23 kDa subunit.Circular dichroism spectra indicated the TPSP had predominantly aβ-sheet structure with a high content of random coils.Differential scanning calorimetry analysis indicated the denaturation temperatures of TPSP at pH 5.0,7.0 and 9.0 were92.0oC,97.1oC and 95.2oC,respectively,and TPSP had a high thermal stability.Furthermore,in the alkaline environment,the surface hydrophobicity of TPSP was low while the surface free sulfhydryls content was high.（3）The functionality of TPSP was studied.The results of experiments showed that the solubility of TPSP increased with the increase of the value of pH.The lowest solubility of TPSP was 8.09%when the isoelectric point was 3.6.However,fat absorption capacity of TPSP decreased with the increase of temperature.The fat absorption capacity of TPSP at low temperature（4oC）was high（8.55 g/g）while at high temperature（100oC）was low（6.35 g/g）.The water holding capacity of TPSP decreased with the increase of the value of pH.In alkaline environment,TPSP had almost no water holding capacity.The foaming and emulsifying properties of TPSP increased with the increase of the value of pH.At pH 11.0,the foaming and emulsifying properties of TPSP had the maximum value（25%and 151.92 m²/g）.But the foam stability and emulsion stability of TPSP increased first and then decreased with the increase of the value of pH.At pH 7.0,the foam stability and emulsion stability of TPSP reached the maximum（70%and 91.95%）.By the analysis of rheological properties,TPSP has a pseudoplastic fluid with good solubility in the alkaline environment,and the minimum gel concentration was 16.00%in the neutral environment.Cytotoxicity experiments showed that TPSP had no cytotoxicity.（4）The hydrolysis of TPSP was carried out by alcalase.The effects of four single factors,such as enzyme concentration,enzymatic pH,enzymatic temperature and enzymatic time,were studied in the experiment of alcalase.On the basis of single factor experiment,optimum enzymatic hydrolysis conditions of TPSP were enzyme concentration 1.0%,enzymatic pH 8.0,enzymatic temperature 60oC,and enzymatic time 3.0 h with the maximum the value of reducing power（0.420）using response surface methodology.（5）The antioxidant properties of tree peony seed polypeptide and TPSP were studied and compared with glutathione（GSH）.The results showed that the antioxidant capacity of GSH was stronger than that of tree peony seed polypeptide and TPSP in the experiment of reducing power and O^2-·radicals scavenging.The other experiments included DPPH·radicals scavenging,ABTS⁺·radicals scavenging,·OH radicals scavenging,Fe²⁺or Cu²⁺chelating activity and inhibition rate of autoxidation of linoleic acid,the antioxidant activity of tree peony seed polypeptide was similar to GSH but always stronger than that of TPSP.