DEVELOPMENT OF BIOREACTOR SYSTEM FOR OLIGOPEPTIDE SYNTHESIS COMBINED WITH A SIMULATED MOVING BED ADSORBER

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In this study, we aimed to construct an effective bioreactor for synthesis of aspartame precursor, as a model oligopeptide in an organic solvent, using thermolysin immobilized onto Amberlite XAD-7. First, we investigated the factors affecting the synthesis of the asparatme precursor using an immobilized enzyme. We studied a synthetic rate, particularly in a mixed organic solvent with ethyl acetate and tert-amyl alcohol. Using a mixed solvent of 33% tert-amyl alcohol and 67% ethyl acetate, the activity and stability of the immobilized thermolysin were high enough to conduct a continuous reaction. With increasing the content of tert-amyl alcohol, the stability of the immobilized enzyme was enhanced. The reason for the high activity and stability of the immobilized thermolysin in the presence of tert-amyl alcohol was studied from the viepoint of stability of calcium bound on the enzyme molecule, a stabilizing factor for thermolysin. An optimum water content for the synthesis of asparatame … More precursor was found to be 4-6 % in the mixed organic solvent. The concentration of L-phenyllalanine methyl ester (PheOMe) and N- (benzy loxy carbonyl) -L-as partic acid (Z-Asp) was optimized. The concentration of PheOMe was set to 200 mM and that of Z-Asp was 40 mM.We could also conduct a continuous reaction in a column reactor at 40ﾟC.The conversion at the outlet of the reactor with respect of Z-Asp was around 99% with a space velocity of 4.9 (1/h). Based on this result, we aimed to develop a bioreactor system combined with a simulated moving bed absorber. Namely, at the outlet of the bioreactor, the Z-Asp concentration was negligible low with asparatame precursor Z-Asp PheOMe of around 40 mM and PheOMe of 160 mM.Thus, we could recycle PheOMe to the bioreactor with continuous separation of PheOMe and Z-AspPheOMe using a simulated mooving bed adsorber. We found Dowex MWA-1 as a suitable adsorbent and analyzed adsorption/desorption behaviors of PheOMe and Z-AspPheOMe. We also calculated a separation performance for a simulated moving bed adosorber. We also showed a possible bioreactor system which could be used for an industrial production. Finally, we briefly compared an enzymatic synthesis and chemical synthesis. Less