Synthesis and spectral properties of chemically and stereochemically homogeneous sphingomyelin and its analogues

Abstract

2-N-Stearoylsphingosyl-1-phosphocholines of D-erythro(2S,3R) and L-threo(2S,3S) configurations and their phosphorothioyl analogues were obtained by a purely synthetic approach using O-methyl-N,N-di-isopropylaminophosphorochloridite as the phosphitylating reagent for the formation of the phosphodiester linkage. Final products were obtained pure in yields of 70–75%. The structure and the purity of synthesized compounds was determined using ¹H, ¹³C, and ³¹P n.m.r. spectroscopy. With respect to the structure of the sphingosine long-chain base the synthetic D-erythro-SPM was found to be identical with the natural sphingomyelin from bovine brain whereas the semisynthetic N-palmitoylsphingomyelin obtained via a deacylation–reacylation pathway comprises a mixture of D-erythro- and L-threo-SPM. The phosphorothioyl analogues of sphingomyelin synthesized by addition of elemental sulphur to intermediate phosphite were separated into individual diastereoisomers having opposite configuration at phosphorus. The absolute configurations of diastereoisomers at phosphorus were assigned based on the known stereospecificity of phospholipase C in the hydrolysis of phosphorothioyl analogues of phospholipids. ¹ H N.m.r. data of synthesized compounds suggest that the configuration at C-3 of the sphingosine influences the average conformation of sphingomyelin with respect to the angle of rotation about the C(1)–C(2) bond.