Synthesis of DHA/EPA-rich phosphatidylcholine by immobilized phospholipase A1: effect of water addition and vacuum condition.

DHA/EPA-rich phosphatidylcholine (PC) was successfully synthesized by immobilized phospholipase A1 (PLA1)-catalyzed transesterification of PC and DHA/EPA-rich ethyl esters in a solvent-free system. Effects of reaction temperature, water addition and substrate mass ratio on the incorporation of DHA/EPA were evaluated using response surface methods (RSM). Water addition had most significant effect on the incorporation. Reaction temperature and substrate mass ratio, however, had no significant effect on the incorporation. The maximal incorporation was 19.09 % (24 h) under the following conditions: temperature 55.7 °C, water addition 1.1 wt % and substrate mass ratio (ethyl esters/PC) 6.8:1. Furthermore, effects of water addition (from 0 to 1.25 wt %) on DHA/EPA incorporation and the composition of products were further investigated. The immobilized PLA1 was more active when water addition was above 0.5 wt %. By monitoring the reaction processes with different water addition, a possible reaction scheme was proposed for transesterification of PC with DHA/EPA-rich ethyl esters. In summary, PC and sn2-lysophosphatidylocholine (LPC) were predominant in the mixtures at early stages of reaction, whereas sn1-LPC and glycerophosphocholine (GPC) predominant at later stages. The vacuum employed after 24 h significantly increased the incorporation of DHA/EPA and the composition of PC, and the highest incorporation (30.31 %) of DHA/EPA was obtained at 72 h and the yield of PC was 47.2 %.