Enhancement effect of water activity on enzymatic synthesis of cephalexin

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Abstract
The effect of water activity (aw) of the reaction medium on the enzymatic synthesis of cephalexin (CEX) from 7‐amino‐3‐deacetoxycephalosporanic acid (7‐ADCA) and D‐α‐phenylglycine methyl ester (PGM) was investigated using the α‐amino acid ester hydrolase enzyme from Xanthomonas citri. It was found that the synthetic activity of the enzyme and the conversion yield were markedly improved when the aw of the reaction medium was lowered to about 0.97. The water activity depressing agents evaluated were glycerol, sucrose, and sorbitol, and the conversion yields were improved up to 170% with 15% glycerol, 230% with 30% sucrose, and 270% with 20% sorbitol, respectively. The extent of favorable effect of aw on the conversion yield was not the same among the aw depressors, probably due to other unknown interactions between the enzyme and depressors. However, optimal aw values corresponding to the maximum conversion yield coincided for all aw depressors used. The conversion yield of CEX showed an increasing trend with increasing aw up to the optimal aw value (0.96–0.97) which corresponds to the maximum conversion yield and a decreasing trend beyond the optimal aw. There appears to be a delicate balance between the hydrolytic reaction of PGM and synthetic reaction of CEX. The increasing aw–[E · PGM] complex and the branched reaction pathway fluxes from [E · PGM] to PG (D‐α‐phenyl glycine) and CEX are balanced in such a way that the maximum CEX conversion yield is obtained at aw value of 0.96–0.97. The aw depressors stabilized the enzyme somewhat, but this positive effect was considered to be only a minor contribution to the substantial yield enhancement. The aw depressor effect on viscosity and in turn the mass transfer rate limitation was ruled out since the change in conversion due to the viscosity change was found to be insignificant. © 1993 John Wiley & Sons, Inc.