Biocatalytic enantiomeric resolution of L-menthol from an eight isomeric menthol mixture through transesterification

Abstract The four diastereomers of menthol and their enantiomers, namely dl -menthol, dl -neomenthol, dl -neoisomenthol and dl -isomenthol, were synthesised by the hydrogenation of thymol to yield an eight isomer liquid menthol. A suitably selective lipase was sought to preferentially esterify l- menthol in organic solvent, hence simplifying separation from the diasteromeric mix through distillation. From an initial screen a commercial Pseudomonas fluorescens lipase (Amano AK) was selected, and vinyl acetate was chosen as a suitable irreversible acyl donor for transesterification. The reaction reagent ratios were optimised, and an enantiomeric excess (ee) of l- menthol of greater than 95% was reproducibly achievable at a conversion of 30% dl -menthol (0.68 M) at ≤50 °C. On the basis of the composition of liquid menthol the reaction had a diastereomeric excess (de) of 82%. The enzyme was recycled 150 times in 5 ml batch reactions using liquid menthol and achieving an overall yield of 184.3 g dl -menthol/g commercial enzyme preparation. The by-product acetic acid, formed by hydrolysis of menthyl acetate, was found to cause a high degree of enzyme inactivation. The resolution reaction was scaled up 400 fold to 2 L and the enzyme recycled 38 times with an average conversion of the available l- menthol of 59% and a volumetric productivity of 1.2 g/L/h.