The effect of the reducing power on the reduction of methyl-2-chlorobenzoylformate was evaluated by using carbon substrates with different reducing powers. Glucose, sorbitol, and gluconate regenerated 2, 3, and 1 NAD(P)H during its conversion to pyruvate, respectively. When sorbitol was used as the carbon substrate, complete conversion was achieved in 8 h while it took 12 h and 19 h when glucose and gluconate were used, respectively. The enantiomeric excess (ee) value was 96.7% when sorbitol was used.