Ethyl esterification of docosahexaenoic acid in an organic solvent-free system with immobilized Candida antarctica lipase

Ethyl docosahexaenoate (EtDHA) is regarded as a potentially useful pharmace utical substance on account of its beneficial physiological activities. We attempted the ethyl esterification of docosahexaenoic acid (DHA) in an orga nic solvent-free system using Candida antarctica lipase, which acts strongl y on DHA and ethanol. Esterification of 88% was attained by shaking a mixtu re of DHA/ethanol (1: 1, mol/mol) and 2 wt% immobilized C antarctica lipase at 30 degreesC for 24 h. However, even in the presence of an excess amount of ethanol, the extent of esterification could not be raised above 90%. To attain a higher level of esterification, a two-step reaction was found to be effective. The first step was performed in a mixture of DHA/ethanol (1:1 , mol/mol), and the reaction mixture was then dehydrated. In the second ste p, the resulting mixture was shaken at 30 degreesC for 24 It with 5 molar e quivalents of ethanol against the remaining DHA using 2 wt% immobilized lip ase. By means of this two-step procedure, 96% esterification was attained. Repetition of the first and second reactions showed that the immobilized li pase was reusable for at least 50 cycles. In addition, DHA remaining in the second-step reaction mixture was removed by a conventional alkali refining process, giving purified EtDHA with a high yield.