Partial synthesis and properties of a series of N-acyl sphingomyelins

Abstract

A series of sphingomyelins (SM) with different chain length fatty acids (C14:0, C16:0, C18:0, C20:0, C22:0, and C24:0) N-linked to the primary amino group of sphingosine have been synthesized starting with bovine brain SM. Two different acid hydrolysis procedures, (1a) butanolic HCl (H. Kaller, 1961, Biochem. Z. 334: 451-456) and (1b) methanolic HCl (R. C. Gaver and C.C. Sweeley. 1965. J. Am. Oil Chem. Soc. 42: 294-298), were used and the resultant sphingosylphosphocholine (SPC) was converted to SM using two acylation methods: (2) using fatty acid imidazolide to yield the O-acyl, N-acyl SPC, followed by mild alkaline hydrolysis for selective deacylation at the O-acyl linkage, and (3) selective acylation at the amino group of SPC using the free fatty acid in the presence of dicyclohexylcarbodiimide. Following chromatographic purification, N-acyl SM were obtained in high yield (80-90%), and were characterized by a combination of thin-layer chromatography, high performance liquid chromatography, chemical analysis, optical rotation, circular dichroism, infrared spectroscopy, 13C NMR, and spingosine base analysis. The N-acyl SM were chemically homogeneous with respect to fatty acid composition and the sphingosine base composition resembled that of the starting bovine brain SM. However, as a consequence of the epimerization at C-3 of SPC in both acid hydrolysis procedures, the resulting N-acyl SM consisted of mixtures of D-erythro and L-threo sphingomyelins. By differential scanning calorimetry hydrated C14:0 to C24:0 SM exhibited gel-liquid crystal transitions in the range 30-50°C but the chain length dependence was complex. X-ray diffraction of hydrated C18:0 SM showed that its transition at 45°C was between bilayer gel and bilayer liquid crystalline states.