Study On The Isolation And Identification Of Rice Residue Fermentation Products From Molds And Its Antioxidant Property

As a rice producing country, the output of rice is about200billion kg per year in China. Besides the supply of daily need for food, most of the low-value rice is used as material for starch-sugar production and industrial fermentation, which may leave a great deal of byproducts. Rice residue is a kind of byproducts produced in the processes of starch-sugar production, organic acid fermentation and monosodium glutamate production. Rice residue in which the content of protein is more than40%and rich in dextrine and polysaccharide, is an ideal protein resource and worthy of exploiting. Based on rice residue as raw material for the solid-state fermentation of fungi, product with high antioxidant activity was isolated and purified by ultrafiltration, gel chromatography and PR-HPLC, and the structure has been identified as well. The main results were as follows:1It was the first time to ferment rice residue with strains of genus of Aspergillus oryzae, Rhizopus oligosporrus, M. racemosus, Aspergillium niger and Penicillium glaucum, on solid state to produce antioxidant product. Antioxidant activity in vivo has been determined by MTT assay. Based on the results of reducibility, clearance ability of DPPH·free radical, ABTS·free radical and OH·free radical and anti-lipid peroxidation ability of each product, Aspergillium niger has been considered the optimum strain for the fermentation. Analysis for the products fermented by the five strains above have exhibited that each product showed a high degree of hydrolysis, and that the inoxidability of fermented products of rice residue mainly depended on the behavior of micromolecular peptide by the hydrolyzation of protein during the post-fermentation has been primarily concluded, combining the analysis of basic ingredients of rice residue.2Using the activity of protease and glucoamylase of koji as evaluation index, primary fermentation conditions have been concluded as follows:water content of rice residue45%,3d for the fermentation at32℃, according to single factor experiment. It has been confirmed that factors of fermentation temperature, fermentation time and water content significantly affect the inoxidability and degree of hydrolysis of RRFPsIV during the after fermentation, and higher antioxidant activity has been detected against RRFPsIV with proper degree of hydrolysis. Using the clearance of DPPH·as index, models for each factor level have been fitted according Box-Beknken response surface. The regression equation was as follow: Y=53.62-0.82X1-0.85X2+0.40X3-2.55X12-2.86X22-1.55X32+0.99X1X2-0.71X1X3According the analysis software of MINITAB16, optimum process for thefermentation of rice residue has been concluded:temperature40.4℃, time5.5d, water content83.4mL/10g rice residue, and the predicted Y of clearance of RRFPsIV against DPPH-was53.56%, degree of hydrolysis38.92%.3The antioxidative stability of RRFPsⅣ and its synergism with other natural antioxidants were studied. Results exhibited that no significant effect of inoxidability of RRFPsIV had been detected by the variation of temperature, pH value and pepsin hydrolysis, however, Cu2+and Fe3+did. Synergistic interaction has been detected when RRFPsIV was blended with Vc and tea polyphenol, but no Synergistic interaction could be detected when blended with VE.4Barrier separation for the hydrolysate of RRFPsIV has been studied with a ultrafiltration membrane of10KDa intercepting, tests showed that clearance of constituent less than10KDa was64.98±3.45%against DPPH·, and it was higher than that above lOKDa. Maximum absorption peak was scaned at210nm towards constituent less than10KDa with UV spectrum. Three constituents named F1, F2and F3was isolated by SephadexG-25from constituent less than10KDa. Clearance of F2against DPPH·was74.71±2.66%. Five main constituents from F2have been isolated by preparative RP-HPLC, of which F2d and F2e have showed a high clearance against DPPH·of77.37±1.01%and83.04±1.69%respectively. It has been confirmed that F2d and F2e were both relative single peaks, and the peak area were90.36%and91.69%respectively according to the purity analysis of analytical RP-HPLC.5It was the first time to identify the molecular weight and constitutional formula of F2d and F2e by MicroTOF-QII tandem mass spectrometry. And the molecular weight were1072.97Da and1130.5132Da respectively, their amino acid sequences have identified as follows (started from N terminal): Val-Ala-Glu-Glu-Glu-Leu-Gly-Gly-Asn-Arg Leu-Asp-Pro-Glu-Gly-Thr-Gly-Thr-Phe-Pro-Pro...