Immobilization of Candida rugosa Lipase on Different Ionic Supports to Improve the Enantioselectivity on Mandelic Acid Resolution

Abstract

Thiswork describes the improvement in enantioselectivity on the resolution ofmandelic acid catalyzed by the Candidarugosa lipase (CRL) immobilized and stabilized on two different ionicsupports. CRL was quickly immobilized on anionic diethylaminoethyl (DEAE)support and, due to the low content of positive charges on enzyme surface; theenzyme could only be successfully immobilized on cationic carboxymethyl supportafter chemical amination. Immobilized the enzyme presented higher thermalstability and higher stability to pH than the free enzyme. In relation to thefree-enzyme, the DEAE derivative was 2-fold more stable in acid pH, while thecarboxymethyl derivative was 2-fold more stable in alkaline pH. When incubated atpH 7.0 and 50 ºC, the carboxymethyl derivative was more stable retaining 80% ofactivity even after 7 h incubation, and the DEAE derivative presented half-lifeof 6.6 h. Due to this promising characteristics, both CRL derivatives wereevaluated on the hydrolysis of (R,S)-mandelic acid ethyl ester under differentpH. The CRL immobilized on DEAE support presented stereochemical preference forthe R isomer in pH 5.0 and 7.0, while in pH 9.0 the hydrolysis of the S isomerhydrolysis was faster with a higher E-value of 21.2. On the other hand, thecarboxymethyl derivative showed opposite results regarding stereochemicalpreference with higher E-value at pH 5.0 (E > 200) demonstrating excellentenantioselective transesterification towards the S-isomer of mandelic acid witha theoretical 50% conversion yield and a 99.9% enantiomeric excess.