[Saccharomyces cerevisiae B5 efficiently and stereoselectively reduces 2'-chloroacetophenone to R-2'-chloro-1-phenylethanol in the presence of 5% ethanol].
(R)-chlorprenaline, a selective activator of beta2 receptor and an effective drug for bronchitis and asthma, is industrially prepared from (R)-2'-chloro-1-phenyl-ethanol. In this communication, we describe (1) the identification of Saccharomyces cerevisiae B5 as an effective host for stereoselective reduction of 2'-chloroacetophenone to (R)-2'-chloro-1-phenyl-ethanol; (2) the presence of ethanol enhances the conversion; and (3) the biochemical factors that effect the yield of the product. Among the four yeast strains capable of reduction 2'-chloroacetophenone to (R)-2'-chloro-1-phenyl-ethanol we screened, Saccharomyces cerevisiae B5 showed the highest activity and stereoselectivity, and was used for the subsequent study. The effect of the presence of methanol, ethanol, 2-propanol, 1-butanol, glucose, glycerol and lactic acid was first investigated, as it was previously reported that they increased the yield and stereoselectivity of the reaction. The addition of the co-substrate methanol, ethanol, 2-propanol, 1-butanol, glucose and glycerol favored the formation of the 2'-chloroacetophenone to (R)-2'-chloro-1-phenyl-ethanol. Lactic acid inhibited the enzyme activity. Ethanol is the best co-substrate among the seven co-substrates and under the optimum concentration of 5% , the yield of (R)-2'-chloro-1-phenyl-ethanol was increased from 17% to 74%. The oxidation of ethanol regenerates NADH required for the reduction. The effects of the reaction time, pH, cell concentration, substrate concentration and temperature on the reduction were investigated next. The enantiometric excess of (R)-2'-chloro-1-phenyl-ethanol reached 100% under the optimal condition: pH8.0, 25 degrees C and 5% ethanol. The product yield went up with the increasing Saccharomyces cerevisiae B5 concentration and reached 100% when the cell dry weight was 10.75 mg/mL and 2'-chloroacetophenone was 6.47 mmol/L. The yield of (R)-2'-chloro-1-phenyl-ethanol decreased sharply with the increase of substrate concentration, as the high concentration of substrates is toxic to the cell and inhibits the activity of reductases. The aerobic cultivation of the yeast and shaking during the reaction increased the yield of (R)-2'-chloro-1-phenyl-ethanol. The yeast can be reused up to 15 times. This research paves the way for economical preparation of chiral 2'-chloroacetophenone to R-2'-chloro-1-phenylethanol.
Ou
Zhi-Min
ZM
Institute of Biochemical Engineering, College of Material Science and Chemical Engineering, Zhejiang University, Hangzhou 310027, China. oozzmm@163.com
Wu
Jian-Ping
JP
Yang
Li-Rong
LR
Cen
Pei-Lin
PL
Liu
Lin
L
Qi
Nan
N
D000431:Ethanol / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D010084:Oxidation-Reduction; D010626:Phenylethyl Alcohol / Q000737:chemistry* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D013237:Stereoisomerism; D002721:omega-Chloroacetophenone / Q000737:chemistry* / Q000378:metabolism*
D000431:Ethanol; D002721:omega-Chloroacetophenone; D010626:Phenylethyl Alcohol