AbstractThe enzyme‐catalyzed sequential esterification of (±)‐2‐methylpentanedioic anhydride (rac‐1) with 2‐methylpropanol in diethyl ether afforded a 1:1 mixture of 1‐(2‐methylpropyl) 5‐hydrogen (S)‐4‐methylpentanedioate (ent‐2) with an enantiomeric excess of 88% and 1‐(2‐methylpropyl) 5‐hydrogen (R)‐2‐methylpentanedioate (3) with an e.e. of 99% besides a considerable amount of the diesterent‐4. Exploiting the ability of the enzymes Novozym and SP 382 to differentiate between the two enantiomers and to catalyze the alcoholysis of both enantiomers with opposite regioselectivity, we reduced the resolution ofrac‐1to a simple chromatographic separation of enantiomerically enriched regioisomeric monoesters. The high optical purity of monoester3could be achieved due to the fast conversion of its enantiomer into the diesterent‐4. Via reduction with lithium borohydride the monoestersent‐2and3were converted into the