Enzymatic Analysis of Positional Distribution of Fatty Acids in Solid Fat by 1,3‐Selective Transesterification with Candida antarctica Lipase B

Abstract

AbstractA protocol for the analysis of the positional distribution of fatty acids (FA) in solid triacylglycerols (TAG) was developed using sn‐1(3) selective alcoholysis catalyzed by immobilized Candida antarctica lipase B (CALB). One part by weight of solid fat and ten parts by weight of ethanol (99.5 %) were warmed to liquefy the fat. After adding 0.44 parts by weight of CALB, the mixture was shaken at 50 °C for 10 min then at 30 °C for 2.8 h. The recovery of 2‐MAG after the 3‐h transesterification reaction was ca. 85 % of the maximum theoretical yield (33 mol%), with the loss of 15 % attributable to the acyl migration from sn‐2 to sn‐1(3). The recovery was similar to that of the solvent‐free alcoholysis of structured lipids, 1,3‐dipalmitoyl, 2‐oleoyl glycerol and 1,3‐dioleoyl, 2‐palmitoyl glycerol, conducted at 30 °C for 3 h. In contrast, the acyl migration from sn‐1(3) to sn‐2 was hardly observed. Because the detected acyl migration was only in the direction of sn‐2 to sn‐1(3), and not vice versa, it is proposed to determine the FA composition of the sn‐2 position of TAG by the gas chromatographic analysis of 2‐MAG fraction recovered from the enzymatic reaction mixture, and the FA composition of sn‐1(3) position by a mass balance using the FA composition of TAG and of the sn‐2 position as inputs. The procedure was successfully applied to palm oil and shea butter, and docosahexaenoic acid (DHA)‐rich single cell oil from Aurantiochytrium sp. KH105 for the first time.