(626d) Improvement of L(+)-Lactic Acid Production by In Vivo Regulation of Alcohol Dehydrogenase In Rhizopus Oryzae

Sitanan Thitiprasert^a, Sarintip Sooksai^b, and Nuttha Thongchul^b

^aBiotechnology Program, Faculty of Science, Chulalongkorn University, Bangkok 10330 , Thailand.

^bInstitute of Biotechnology and Genetic Engineering, Chulalongkorn University, Bangkok 10330, Thailand.

Lactic acid production by Rhizopus oryzae is now becoming more attractive since this fungus produces an optically pure L-lactic acid. However, the filamentous fungal fermentation has low product yield and production rate due to the formation of other by-products, including ethanol and fumaric acid. Many methods including cell immobilization have been used to improve lactic acid production; however, none of them successfully inhibited ethanol production. In this study, prevention of ethanol production by regulating alcohol dehydrogenase (ADH) during fermentation by R. oryzae was attempted. Changes in cell growth, product formation, and enzyme activity including lactate dehydrogenase (LDH) and ADH when the different potential inhibitors were observed during the shake flask cultivation of immobilized R. oryzae on a cotton matrix. Lower ethanol but higher lactic acid yields were found from the fermentation with the media containing pyrazole and 2,2,2-trifluoroethanol. Growth delay and morphology alteration were observed from the cultivation of R. oryzae in the media containing p-chloromercuricbenzoate, N-ethylmaieimide, and 4,4´-dithiodipyridine. Dramatic increasing ADH and LDH activities but slight changes in product yields might be explained by the inhibitors governed the enzyme activities at the pyruvate branch point. This revealed that in living R. oryzae, the inhibitors regulated the flux through the related pathways.

Keywords       Rhizopus oryzae, lactic acid, ethanol, in vivo inhibition, alcohol dehydrogenase