Optimization Of The ?-Glycosidase Production Of Oenococcus Oeni And Its Application In Winemaking

Wine quality is predominantly affected by aroma,so aroma is one of the most important factors to evaluate wine quality.In winemaking,?-glucosidase derived from wine-related lactic acid bacteria?LAB?,especially Oenococcus oeni?O.oeni?,is the most important glycosidase,which can contribute to wine aroma by malolactic fermentation?MLF?,because it could hydrolyze the grape-derived aroma precursors which couldn't be perceived.In this study,a strain,CS-7b was choosed from 12 O.oeni strains possessing important oenological characteristics,the commercial strains 31-DH was used as control to optimize their enzyme production conditions by single factor experiment and orthogonal experiment,and explore their enzymatic properties.Then,the O.oeni cells were crushed by ultrasonic processor to extract ?-glucosidase,and evaluate its wine-making adaptability.Finally the crude enzyme extraction was added to wine,then the change of aroma substances in wine was analysed by GC-MS,exploring the effect of O.oeni ?-glycosidase on wine aroma and sensory quality.The main results are as following:?1?The enzymatic activity of O.oeni CS-7b is significantly higher than other strains,it is0.359?mol/?g·min?,twice as much as the others,and quite stable.?2?Through series of single factor experiment,it was confirmed that the optimal growth period of producing ?-glycosidase was middle of logarithmic phase,the optimum carbon source was maltose,and the optimum nitrogen source was yeast powder,initial pH of inoculated medium was near 6.8;Then the best content and proportion of various substances of the medium was determined by the orthogonal experiment?1L medium?: maltose 7g,yeast powder 20 g,MgSO₄·7H₂O 0.1g,cysteine hydrochloride 0.5g,MnSO4·4H₂O 0.03 g,medium initial pH value was 6.8.Under this condition,?-glycosidase activity can be increased to1.3mmol/?g·min?as compared to the original 0.3mmol/?g·min??3?The enzymatic properties of O.oeni were characterized as follows: O.oeni produced relatively high ?-glycosidase activity at pH 5.0 and 37 ?;the?-glycosidase activity wasmildly effected by a low content of sugar,whereas,the activity of enzyme from most strains was tremendously suppressed by high sugar concentration;Moreover,alcohol resulted in significant decline of ?-glycosidase activity among all the tested strains,in addition,the tested strains lost its overall enzymatic activity in the presence of 14%?V/V?ethanol.?4?O.oeni strain CS-7b showed the prime enzymatic characteristics compared with the other tested strains.After crushing these cells,?-glycosidase was found still clinged to the pieces of broken cells,and broken fragments were picked up as crude enzyme extraction to explore the adaptability of extracellular enzyme in winemaking.Finally,addition of the crude enzyme extract was performed to the wine and analysis aroma component by GC-MS,the differences of aroma in wine showed that ?-glycosidase produced by O.oeni strains CS-7b has potential application for winemaking as fumette.