tion of glycerophosphate, is a liquid at room temperature and can thus function both
as substrate and solvent in the reaction. The best results were obtained without any
additional solvent. Vinyl esters are activated substrates, and their hydrolysis is an
important side reaction. In the case of glycerophosphate acylation, the water content
in the reactors was low at the start of the reaction and, due to hydrolysis of the vinyl
ester, was decreased to very low values. It has been shown previously that lipases, in
contrast to most other types of enzymes, can demonstrate activity at very low water
activity (Valivety et al., 1992). Even under these very dry conditions, the lipase can
continue to function, and because of the low water activity the equilibrium is in favor
of the acylated products.
TheRhizopus arrhizuslipase is specific for the 1-position in glycerophospholi-
pids; thus the first acylation product was 1-acyl-rac-glycero-3-phosphate (lysophos-
phatidic acid, Figure 7) (Virto et al., 1999). This would have been the sole product if
acyl migration had not occurred. However, due to acyl migration the initial product
isomerized to 2-acyl-rac-glycero-3-phosphate. Only relatively low amounts of this
product are expected to be formed since lipids with the acyl group on a primary
hydroxyl of the glycerol backbone are usually thermodynamically more favored.
However, 2-acyl-rac-glycero-3-phosphate is a substrate of the lipase and can be
acylated to form 1,2-diacyl-rac-glycero-3-phosphate (phosphatidic acid; Figure
7). Depending on the reaction conditions, the main product was either lysophospha-
tidic acid or phosphatidic acid. Reactions were carried out in atmospheres with fixed
water activity in order to control the water activity in the reaction mixtures. Such
control was ineffective during the period when hydrolysis of the vinyl ester occurred
at a high rate. During this period, water activity control was too slow, and water
activity in the reaction mixture was very low. However, towards the end of the re-
actions, water activity control influenced the product distribution. A low water ac-
tivity favored a relatively high amount of phosphatidic acid (due to relatively fast
acyl migration and equilibrium position in favor of acylated products). A practical
298 14 Enzymatic Conversions of Glycerophospholipids
Figure 7. Acylation ofD,L-glycero-3-phosphate with vinyl esters in a reaction catalyzed byRhizopus
arrhizuslipase. Due to limited acyl migration in the product of the first reaction step (lysophosphatidic
acid, 1-acyl-rac-glycero-3-phosphate), a second acylation reaction could occur yielding phosphatidic
acid. (From Virto et al., 1999.)