SYNTHESIS OF CHIRAL LIPIDS. APPLICATIONS TO THE SPECIFICITIES OF LIPOLYTIC ENZYMES.

Abstract

In this study, synthetic routes to certain short acyl chain phospholipids were developed. Their inhibitory or substrate properties for phospholipase A₂ were then examined. An improved method for the acylation of glycerophosphocholine is described using a mixed fatty acid - trifluoroacetic acid anhydride. This partial synthetic route is particularly suitable for obtaining short acyl chain phosphatidylcholines. A new pathway for constructing the unnatural sn-1-phosphatidylcholines is also described, starting from L-arabinose. This is converted first to a key intermediate, 2,3-O-isopropylidene-sn-glycerol, which is then phosphorylated and transformed into sn-glycero-1-phosphocholine. This can be acylated as above to give sn-1-phosphatidylcholines. Routes to the short chain phosphatidylethanolamines were investigated and discussed. A procedure, using phospholipase D, was used to convert L-diC₆ PC into L-diC₆ PE in a transphosphatidylation reaction. Failed attempts to obtain shorter chain homologs by this and other methods are also detailed. The kinetics of inhibition of the phospholipase A₂ hydrolysis of L-diC₆ PC by the D-isomer are also reported for the monomeric concentration range. It was found that the D- enantiomer did not behave as a pure competitive inhibitor, and that an enzyme-substrate-inhibitor complex can exist. The implications of these results with regard to PLA₂ hydrolysis of mixed micelles is discussed. The PLA₂ substrate properties of both the anionic and zwitterionic diC₆ PE's were also studied. It was established that the anionic diC₆ PE is either a very poor substrate relative to the zwitterionic diC₆ PE, or acts as a competitive inhibitor towards its hydrolysis. Similarly, the rates of base-catalyzed acyl ester hydrolysis are about 18 times greater for the zwitterionic than for the anionic diC₆ PE. The importance of a protonated amino group in both these hydrolyses studies is noted. In addition, certain physical properties of diC₆ PE, such as its critical micelle concentration and carbon-13 NMR spectrum, are also given.