Enzymatic synthesis of esters in supercritical fluids

Abstract

Lipases are enzymes that are widely used to catalyze a variety of industrially important reactions. However, lipase?catalyzed reactions are heterogeneous and mass?transfer limited. Therefore, in most cases the reaction rates are very low in conventional aqueous and organic media, which limits the applicability of lipase reactions on an industrial scale. Due to their unique properties, supercritical fluids (SCFs) have proven to be efficient media for reactions with mass?transfer limitations. The present study aims at investigating hydrolysis and esterification reactions using commercially available enzymes (crude Hog pancreas lipase, Lipolase 100 T and Novozym 435) in various supercritical fluids (CO?, CH?, C?H?). The enzymatic hydrolysis of p?Nitrophenyl laurate (PNPL) to p?Nitrophenol (PNP) was studied in supercritical carbon dioxide using crude Hog pancreas lipase (HPL), Lipolase 100 T and Novozym 435. All the three enzymes were compared for their optimal temperatures, pressure effects and kinetics. The conversions were around 25%, 23% and 35% for HPL, Lipolase and Novozym respectively. The esterification of fatty acids of different chain lengths (C?–C?) with isoamyl alcohol was studied in solvent?free conditions and in supercritical carbon dioxide. Three different commercial lipases were used for the synthesis of the esters. The results indicate that in supercritical carbon dioxide Novozym gave high conversion (around 40%) for the esterification of all acids, that is, conversion was almost independent of the fatty?acid chain length. On the contrary, HPL and Lipolase were more specific toward the fatty?acid chain length. HPL favored the conversion of higher chain lengths of fatty acids whereas with Lipolase?catalyzed reaction, conversion decreases with increasing chain length. In the case of solvent?free reactions the conversion obtained was almost constant, about 80%, but the fatty acid with odd number of carbon chain gave higher conversion than the fatty acid containing even number of carbon chain. The esterification of palmitic acid with ethanol was investigated at various temperatures (35–70°C) in the presence of three lipases in supercritical carbon dioxide (SC?CO?) and solvent?free conditions. All enzymes showed an optimum temperature of 55°C under both conditions. The effect of water and ethanol addition on the esterification of acid was also investigated and the conversion obtained in SC?CO? and in solvent?free conditions with Novozym at optimal conditions was 74% and 97%, respectively. The esterification reaction of palmitic acid and 2?ethyl?hexyl alcohol (octanol) was carried out in three different supercritical fluids, CO?, CH?, C?H?, with three different lipases. The conditions of enzymatic synthesis of octyl palmitate were optimized in the system of supercritical carbon dioxide (SC?CO?), supercritical methane (SC?CH?) and supercritical ethane (SC?C?H?). Reaction in supercritical methane gave higher conversions (nearly 85%) than the reactions in SC?CO? (nearly 75%) and in SC?C?H? (nearly 80%). Although a higher apparent yield was obtained in the solvent?free condition due to higher substrate and enzyme concentrations, the reactions in supercritical fluids are more viable because of low enzyme loading, higher reaction rates and easier downstream processing. These studies indicate that Novozym 435 is the most efficient enzyme (among the enzymes tested) for the hydrolysis as well as esterification reactions.