Direct separation of monoacylglycerol isomers by enantioselective high-performance liquid chromatography

An HPLC-based method for separation of isomers of monoacylglycerols (MAGs) was established using an enantioselective column, Chiralcel OF. Three MAG isomers, i.e. sn-1-MAG, sn-2-MAG and sn-3-MAG, having the same acyl groups were baseline separated with resolution factor of more than 1.2 between adjacent peaks. In addition, mixtures of enantiomer pairs of MAG species (sn-1 and sn-3-MAGs) having different acyl groups could be resolved with partial overlaps of some peaks. The sn-1 and sn-3 molecular species were eluted in turn, and the elution order depended on the equivalent carbon number. As a demonstration of the applicability of the established analytical method, stereoselectivity of four microbial lipases, i.e. Candida antarctica lipase B, Rhizomucor miehei lipase, Pseudomonas pseudoalkali lipase, and Thermomyces lanuginosa lipase, in esterification of glycerol with fatty acids was investigated. It was found that the stereopreference of the lipases depended on the types of fatty acids and types of organic solvents.