Purification Of Bromoperoxidase And Its Application In Enzyme Membrane Reactor

Vanadium-bromoperoxidase is an important class of haloperoxidase, rich in marine algae. The enzyme has good thermal stability and organic solvent tolerance. It can catalyze a variety of reactions, such as epoxidation of olefins and oxidation, sulfoxidations of organic compounds, amino acid oxidation and so on,showing an excellent industial applications value. In this research, vanadium-bromoperoxidase was purified from Coralline officinalis in Da lian, and its properties were studied. Meanwhile, V-BPO's catalytic function in biphasic enzyme membrane bioreactor (EMBR) was also studied.A novel, simple and highly efficient process for purifying vanadium- bromoperoxidase from Corallina officinalis was developed. Crude enzyme extraction after being added 1.0 mmol/L Na₃VO₄ was heated at 70 oC for 2h, and then was ultrafiltered with Poly(vinylidene fluoride) (PVDF) membrane, which MWCO is 100 kDa. The V-BPO was purified with a total activity yield of 96% and a specific activity of 310 U/ mg, which is 2.1 times of commercial V-BPO from Sigma.The enzyme characteristics of purified V-BPO were investigated. The optimum pH is 6.0 and each subunit has a molecular weight of 66 Kda. The Km values for MCD, KBr and H₂O₂ were 3.74μmol/L, 0.019 mmol/Land 0.016 mmol/L respectively. It has good tolerance to H₂O₂. Besides, its activity and thermal stability can be enhanced by vanadium.The catalytic traits of V-BPO purified from Corallina officinalis in a biphasic enzyme membrane bioreactor (EMBR) were reasearched. In this subject, we used membrane raector instead of one-pot reactor. Effect of operating variables on the performance of this EMBR was investigated with the catalytic oxidation of cyclohexene, such as immobilized methodes, membrane aperture, reaction temprature, the total of enzyme, substrate concentration and so on. It was found that, under the optimum condition, the eventually concentration of epoxy cyclohexane was about 19.87 mmol/L. This reaction system avoided some problems that were separation and purification of the products. Meanwhile, the separation of aqueous and organic phase was realized, and greatly simplified the process, reduced the cost of production, which can supply some reference for V-BPO in EMBR preliminary application.