Lipase-catalyzed acidolysis and phospholipase D-catalyzed transphosphatidylation of phosphocholine

Two approaches on enzymatic phospholipid modification were studied: (1) transphosphatidylation of the 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC) and ethanolamine in biphasic and anhydrous organic solvent systems by phospholipase D (PLD) and (2) incorporation of oleic acid into the sn1-position of DLPC in organic solvents with different immobilized lipases at controlled water activity. First, DLPC was chemically synthesized from glycerophosphocholine and lauric acid. Next, PLD-catalyzed head group exchange of DLPC with ethanolamine was studied using an enzyme from Streptomyces antibioticus expressed recombinantly in E coli. A comparison of the free PLD with the biocatalyst activated by a salt-activation technique using KCl showed that the salt-activated enzyme (PLD-KCl) was 10-12 folds more active based on the amount of protein used. Thus, DLPC was quantitatively converted to 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine in an anhydrous solvent system within 12 h at 60 degreesC. For the acidolysis of DLPC with oleic acid, among the four lipases studied (CAL-B, Lipozyme TL IM, Lipozyme RM IM and lipase D immobilized on Accurel EP-100), Lipozyme TL IM showed the highest activity and incorporation of oleic acid. A quantitative incorporation was achieved at 40 degreesC using a 8-fold molar excess of oleic acid in n-hexane at a water activity of 0.11.