Inducing Characteristics And Subcellular Locations Of Hexanol-degrading Enzymes In Geotrichum Sp. And Application

1-Hexanol is a kind of higher alcohols,widely existing in wines and food flavor.High content of hexanol can distort the food quality and cause hazard to human nervous system.Currently,with people paying more attention on food quality and safty,it is necessary to explore an efficient way to degrade hexanol in foods.Lots of researchs have focused on microbial and enzymatic degradation for their advantages on efficient,safe and convenient specificity.The paper used a hexanol-degrading Geotrichum sp.strain and carried out a serious studies in aspects of investigating the influences of induction treatment with different higher alchols,localizingthe key degrading enzymes using subcellulare fraction technology,and testing the application of the strain in different conditions.The obtained main results were listed as the follows:(1)In enzyme isolation,liquid nitrogen grinding method was superior to ultrasonic disruption method with yielding more enzymes and higher hexanol-degrading activity.Induction with different higher alcohols showed that the capability of the strain in decreasing hexanol changed with the higher alcohols used in the induction treatments.The highest hexanol-degrading activity occured in the induction with 10 g/L hexanol and the obtained hexanol decreasing activity was two times than that without induction treatment.The maximum dehydrogenase activity,being 80%of that without induction treatment,occurred after induction with 1 g/L hexanol.The dehydrogenase activity was the minimum after induction with 10 g/L isoamyl alcohol,being only 24%of that without induction treatment.Comparatively,induction with 1-hexanol resulted in high dedydrogenase activity,whereas,induction with other higher alcohols did not.(2)Cell composition was devided into three parts named after P27,P100,and SN100.NADH oxidase,a marker enzyme located in cell membrane,was detected in the P100 fraction corresponding to the plasma membrane,but not found in the other two fractions.Hexanol-degrading activities were detected in fractions P27 and P100,up to 3.0 ?mol/min and 3.6 ?mol/min,respectively.Maximum dehydrogenase activity was found in fraction SN100 with activity of 0.12 U.Some evidences showed hexanol-degrading enzyme might be located in cell membrane and cell wall.However,the hexanol-degrading enzyme activity was not positively correlated with hexanol dehydrogenase activity in each fraction.Therefore,maybe the enzymes other than dehydrogenase also contributed to the decrease of higher alcohols.(3)Whole cells of Geotrichum sp.CCTCC AF 2012006 were immobilized by alginate beads.Free and immobilized cells were used to treat hexanol under conditions with different pH,NaCl concentration and ethanol concentration to test the capability of this strain to decrease hexanol under different conditions.Results showed free cells performed better at condiitons of pH 5.0-9.0,ethanol concentration 0-20%,and 0-0.3 mol/L KCl and they could be used for two times.However,immobilized cells had better activity in decreasing hexanol at conditions ofpH 4.0,0.6-1.2 mol/L KCl and ethanol content20-40%and they could be used for above four time.Therefore,immobilized cells might be preferred in harsh conditions for practical application.