Production Of L-phenylalanine By Immobilized Cells

Escherichia coli SW0209-52, a recombinant strain with high transaminase activity, was immobilized and was used to prepare L-phenylalanine in packed bed reactor. L-phenylalanine extraction from the biotransformation solution was also studied in the research.The cells of E. coli SW0209-52 were entrapped by ?-carrageenan and were treated by glutaraldehyde after immobilization. The activity yield of the immobilized cells was 73%. Various characteristics of immobilized cells were assessed. The optimal pH for conversion was 8.0, the optimal temperature was 37 oC and the optimal amount of 5'-PLP was 100μmol/L. The storage stability and the operational stability were greatly improved after immobilization. The effect of 5'-PLP and VB₆ on conversion by immobilized cells was compared. As VB₆ can be converted to 5'-PLP with ATP, it can substitute 5'-PLP at a certain extent, but the stability of immobilized cells was decreased.Comparing the effect of H/D on continuous operation, the packed bed reactor with 13.3 H/D ratio was chosen for continuous production. Effects of substrate concentration and residence time were investigated. Considering both conversion yield and volumetric productivity, studies on the operational stability of the reactor were conducted under the condition of 30g/L PPA feeding concentration and 10 h residence time. The system could be operated for 40 d without significant decline and the average productivity was 1.85g/L/h and the conversion yield was above 85% throughout the 40 days.The extraction of L-phenylalanine from biotransformation solution by ion-exchange resin JK006 was also studied. Effects of factors about adsorption, elution and decoloration were investigated. The optimal operation conditions were as following: biotransformation solution pH 2.0, concentration 5g/L, adsorbing temperature 20 oC, eluant 1mol/L NH3. After decoloration by resin D4006, the effluent was concentrated and crystallized. The purity of L-phenylalanine crystal was 96.7% and the yield of the total process was 82.8%.The raw material cost of the entire process was estimated and the biotransformation cost of production with 5'-PLP and VB₆ respectively was compared. Although the cost of immobilization was increased, the whole cost of biotransformation with VB₆ as coenzyme was decreased. In order to bring the process of production of L-phenylalanine with immobilized cells industrialization, 5'-PLP immobilization and methods of improving the stability of immobilized cells in the process of production with VB₆ as coenzyme need to be further studied.