Fermentation Production And Formulation Preparation Of Kluyveromyces Lactis ?-Galactosidase

?-Galactosidase?the lactose-hydrolyzing enzyme, which catalyzes the hydrolysisof ?-galactose glycosidic bond of ?-D-galactopyranosides. Meanwhile,?-Galactosidase with transgalactosylation activity, has been used to producegalactooligosaccharide (GOS). ?-Galactosidase has been extensively applied to foodindustries, medical science and biochemical analysis, et al. Regarding Kluyveromyceslactis ?-galactosidase as the research object, the main contents in this paper were asfollows:(1) Optimization of culture conditions by using response surface method(RSM);(2) Optimization of fermentation process and development of a new fed-batchstrategy in a7-L fermenter;(3) Development of industrial mediums by using wheypowder as the carbon source and the main energy source;(4) Study of the?-galactosidase enzymic preparation.The optium culture conditions by RSM in this paper: lactose20g/L, peptone5g/L,yeast extract12g/L, malt extract5g/L, Na2HPO412H2O0.01mol/L, MgSO40.6g/L,FeSO40.06g/L, pH8.48, tempariture27.6oC. Under the conditions, the?-galactosidase activity reached up to20.60U/mL, which was at about2-fold increasethan the initial conditions (6.85U/mL).The optium fermentation process by a ‘single factor' way in a7-L fermenter,which were both in favour of the secretion of ?-galactosidase and grow ofmicroorganisms: agitation speeds200rpm, aeration rate5L/min. Under thiscondition, ?-galactosidase activity obtained48.41U/mL and biomass reach up to7.21g/L. In order to obtain the higher ?-galactosidase production, a new fed-batch strategybased on the pH feedback control was developed successfully. The ?-galactosidaseactivity, productivity and the biomass with the new fed-batch strategy in a7-Lfermenter reached up to111.61U/mL,5.31U/(mL h) and10.73mg cell dry wt./mLafter21h, respectively, which were at about1.31-fold,3.18-fold and0.49-foldcompared with the batch culture, respectively.The desired industrial medium culture regarding the whey powder as the mainraw material: whey powder60g/L, yeast extract9g/L, peptone3g/L, MnCl20.03g/L, ZnCl20.058g/L, pH8.0. According to the analysis and synthesis in the shakenflask and the7-L fermenter, the productivity of ?-galactosidase production in the industrial medium cultures was as almost same as the above optium culture conditions.But the former was perfered to the ?-galactosidase enzymic preparation.The preparation of permeabilized cells optimized regarding the fermentationbroth cultivated in the7-L fermenter above as the raw material: the incubated cells30g/L, ethanol35%(v/v), stirred for2min, washed with buffer solution (50mM sodiumphosphate buffer, pH7.0) and resuspended in the buffer solution. Then the?-galactosidase enzymic preparation was studied by using room-temperature-drying,freeze-drying and spray drying, respectively. According to the analysis of these resultsobtained, the enzymic preparation by room-temperature-drying was good quality,which was conductive to further study.