Front Matter

14.5.2 Phospholipase A 2 -catalyzed esterification

The phospholipase A 2 -catalyzed esterification of lysophospholipids, especially of

LPC, has been the subject of several studies. Reaction conditions and yields are

summarized in Table 2, but in the early studies, low yields (6–7 %) were obtained

(Pernas et al., 1990). In most of the published methods, the aim has been to introduce

polyunsaturated fatty acids into the glycerophospholipid (Na et al., 1990). Isooctane

was found to be a good solvent for esterification of LPC (Lilja-Hallberg and Ha ̈rro ̈d,

1995), while a high fatty acid concentration and a low water content were found to

favor the esterification reaction, as would be expected from the law of mass action.

The highest yield was 21 %. Reaction temperatures of up to 80 8 C were attempted,

but when these were highest nonenzymatic esterification occurred in addition to

degradation of the polyunsaturated fatty acids.

A high yield in the phospholipase A 2 -catalyzed esterification (35 %) has been

reported in anhydrous benzene (Mingarro et al., 1994). The reaction has also

been attempted in high-pressure reactors (Ha ̈rro ̈d and Elfman, 1995); the best yield

(25 %) was obtained using propane as solvent, though it was possible also to use

carbon dioxide.

One means of achieving a low water activity is to use a water-miscible solvent

with a low water content. Using this approach with glycerol as solvent, a yield of

27 % was obtained for the esterification of lysophosphatidylethanolamine with poly-

unsaturated fatty acids (Hosokawa et al., 1995a).

Recently, the pancreatic phospholipase A 2 -catalyzed esterification of LPC with

fatty acids was studied in some detail (Egger et al., 1997). The reactions were car-

ried out under controlled water activity with toluene as solvent and immobilized

pancreatic phospholipase A 2 as catalyst. Awater activity of at least 0.22 was needed

for the reaction to occur, and although the reaction rate was found to increase with

increasing water activity, the final yield decreased with increasing water activity, as

expected. In order to achieve a high yield in a reasonable time, an approach with

stepwise changes in water activity was used. The water activity was initially

high to provide a high reaction rate, but was then decreased. At each water activ-

ity, the reaction was continued until equilibrium was approached. Finally, when the

water activity was reduced from 0.22 to 0.11, the reaction stopped after some time

because the resulting water activity was too low for that enzyme (Egger et al., 1997).

The yield obtained (60 %) corresponded to the equilibrium position at the lowest

water activity at which the enzyme was active. If enzymes which are active at still

lower water activities can be found, the yield can be further improved.

14.5.3 Possibilities for enzymatic esterification

of glycerophospholipids

Starting from a fully deacylated phospholipid, several hypothetically possible meth-

ods exist of using enzymatic esterification reactions to prepare acylated lipids (Fig-

ure 8). The lipases are normally selective for esterification in thesn-1 position, and

phospholipase A 2 for thesn-2 position. However, if the reactions are carried out

14.5 Esterification reactions for the incorporation of fatty acids into lipids 301