| As far as the biodiesel manufacture by transesterification biocatalized on fats, itcould increase the mole ratio of methonal to oil in the transesterification reaction systemand the reaction efficiency to enhance the methanol tolerance of biocatalysts.In this paper, it was realized to enhance the methanol tolerance of biocatalysts forbiodiesel manufacture by two ways, one of which was to screen and acclimate thelipase-producing strain, the other was to immobilize lipase.The lipase-producing strains were isolated from soils. Six strains of the34isolateswere selected by the larger transparent zone on the selective medium with victoria blue.Strain FM-1was the best strain producing intracellular lipase with high activity132u/gcell. The growth of strain FM-1was improved by adding0.1%Tween80in media. Tocrush the cells of strain FM-1culivated for60h by ultrasonic, the best condition was tooperate for15min (5s with15s intermittent everytime) at560w.The strain FM-1was acclimated repeatedly by the traditional cultivation in themedium containing mehanol, which became the strain tolerant to methanol. Afteracclimation in the medium containing1.1%(v/v) methanol, the relative lipase activityproduced by strain was30%, the original strains could not grow under the samecondition.The lipase was immobilized by adsorpted on the gel pellet of PVA-alginate，the addition of active carbon could increase the adsorption capacity of carrier. Resultsshow that the activity of immobilized lipase is higher when the lipase is immobilized byadsorption for30mg/mL enzyme liquid on PVA-alginate gel pellet containing1%active carbon at25℃and pH5.5on the shaker at60rpm.Compared with free lipase, the optimal reaction conditions of immobilized lipasewere changed, in which the opimal temperature was10℃higher, the optimal pH wasshifted from7.0to7.2and the pH range was widened.The stability of immobilized lipase had been improved in temperature, pH, storage and operation, especially in methanoltolerence. |
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