Degrading Enzyme Of Organophosphorus Pesticides Optimization Of Fermentation Conditions And Enzyme Properties Research

In this paper, genetic engineering bacteria E.coliBL2l （DE3）/pET30a-mpd fermentationconditions were optimized. Investigated by single factor experiments medium carbon,nitrogen,phosphate type and concentration, the initial fermentation pH, inoculum size, medium volume,fermentation temperature, and lactose content and induction time on the degradation oforganophosphorus pesticides Enzyme, obtained from degrading enzyme synthesis in favor ofthe composition of the above factors as: glycerol as carbon source, yeast extract and peptoneas nitrogen source, initial pH8.0, inoculum5%, medium volume100mL/500mL TriangleBottle, temperature25℃,4hours after the fermentation of lactose added. Identified the basicmedium composition and fermentation conditions. By Plackett-Burman experimental designwas investigated in the medium composition and culture conditions of the eight factors on thedegradation of enzyme synthesis, selected from the three affect the production of the keyfactors degrading enzyme: glycerol concentration, fermentation temperature and lactosecontent. The optimal conditions for other factors: yeast extract15g/L, peptone10g/L,phosphate0.08mol/L, inoculum concentration5%, medium volume100ml/500mL flask,initial pH8.0, lactose Added4hours after fermentation. Further investigated by responsesurface analysis of three important factors on the degreation of enzyme synthesis byregression equation, the optimal value of the above three conditions were: glycerol20.8g/L,fermentation temperature23.4℃, lactose content13.08mmol/L. Therefore, the bestoptimized medium composition and culture conditions: yeast extract15g/L, peptone15g/LK₂HPO₄3H₂O,0.08mol/L, KH₂PO₄0.02mol/L. Appropriate culture conditions: mediumwith initial pH8.0, liquid volume100mL, inoculum concentration5%, temperature37℃,speed,180r/min, engineered bacteria E.coliBL2l （DE3）/pET30a-mpd in this Enzymeproduction under the best time of12h, the strain produced by the optimized enzyme activity7.02U/mL.Degradation enzyme purification, enzyme activity increased29times over. Enzymeproperties and do a preliminary study, the optimal enzyme reaction temperature by30℃pH8.0best in alkaline conditions, can retain stored at room temperature. Metal ions Fe²⁺, Mn²⁺activate the enzyme has a strong effect. With the purified enzyme degradation of chlorpyrifosand malathion, with gas chromatography, have minimal residual chlorpyrifos, malathion wasnot detected, indicating that the two enzymes have very good organic phosphorus pesticidedegradation.