Lipase-catalyzed reactions in organic and supercritical solvents: application to fat-soluble vitamin determination in milk powder and infant formula

The aim of this study was to thoroughly investigate the possibility of usin g enzyme catalyzed hydrolysis and alcoholysis of ester bonds in vitamin A a nd E esters to facilitate their determination in different food formulas. T wo vitamin esters, retinyl palmitate and alpha -tocopheryl acetate were use d as model compounds and two food formulas, milk powder and infant formula, were used as model matrices. Six lipase preparations and one esterase prep aration were investigated in the solvents di-isopropyl ether, hexane/ethano l and supercritical carbon dioxide containing ethanol, Three of the enzyme preparations, lipases from Candida antarctica (Novozyme 435), Rhizomucor mi ehei (Lipozyme IM) and Pseudomonas cepacia, showed considerably higher acti vity toward retinyl palmitate than the other four enzyme preparations. Ther e was no observed activity with alpha -tocopheryl acetate using any of the enzyme preparations. Novozyme 435 showed highest activity in supercritical fluid and generally larger tolerance to variations of the investigated para meters. Using this enzyme preparation in supercritical carbon dioxide conta ining 3 vol% ethanol and 0.03 vol% water at 366 bar and 80 degreesC, quanti tative conversion of retinyl palmitate to retinol was obtained. These condi tions were then used for simultaneous lipase-catalyzed reaction and extract ion of vitamin A and E from milk powder and infant formula. The developed s upercritical fluid extraction method using immobilized Candida antarctica p reparation seems to be more beneficial to the oxidation prone vitamins A an d E compared to extraction methodologies based on alkaline saponification, resulting in comparatively higher recoveries. (C) 2001 Elsevier Science Inc . All rights reserved.