Research On Bio-Transformation Of P-Hydroxybenzylcyaide To P-Hydroxyphenzylacetic Acid By Immobilized Bacillus Subtilis E9 In The Packed Bad Bioreactor

P-hydroxyphenylacetic acid is an organic synthetic intermediate, that is widely used in organic synthesis to prepare some valuable medicine, pesticide and liquid crystal material etc. Our laboratory has got a strain Bacillus subtilis E9 which can catalysis p-hydroxybenzylcyanide to p-hydroxyphenylacetic acid. This dissertation studied the process of bio-transformation of p-hydroxyphenylacetic acid by free cells and immobilized cells in bioreactor.This paper studied the conversion process of p-hydroxyphenylacetic acid by free cells in the bubble bioreactor. The biotransformation conditions were optimized. The substrate dissolved in the 300 mL buffer solution including the free cells was transformed in the 500 mL bubble bioreactor. The aeration rate was 0.6mL/min, which was used to mix the solution. Many gel matrixes were used to embedding resting cells, in which we optimized biotransformation conditions of agar and calcium alginate. The mechanical strength of the immobilized cells was improved by polyethylene imine and glutaraldehyde. The strengthen conditions were optimized by response surface method.The strengthed immobilized cells were filled in the packed bed bioreactor to hydrolyze p-hydroxybenzylcyanide. Compared with free cells, the enzyme activity of immobilized cells was more stable. After 46 batches, 58.84% enzyme activity of the immobilized cells was remained. The enzyme denaturation rate(k_D) was 0.0033, and the enzyme half life was 210h at the temperature of 32℃.