The synthesis of internal standards for the quantitative determination of sphingolipids by tandem mass spectrometry.

Novel internal standards have been synthesised for the quantitative determination by tandem mass spectrometry (MS/MS) of the sphingolipids that accumulate in lysosomal storage diseases. The [d4]C16- and [d47]C24-isoforms of galactosylceramide (CMH), lactosylceramide (CDH), globotriaosylceramide (CTH), cerebroside sulphate, sphingomyelin and G(M1)-, G(M2)- and G(M3)-gangliosides were synthesised by the reaction of their lyso-forms with the acid chlorides of hexadecanoic 5,5,6,6-d4 acid ([d4]-palmitic acid) and tetracosanoic-d47 acid ([d47]-lignoceric acid), respectively. The acid chlorides were formed using oxalyl chloride. The structures of the internal standards were confirmed by MS/MS. The fragmentation pattern of each novel compound was similar to that of the corresponding natural form of the sphingolipid, making it a good internal standard for the quantitative determination of the natural sphingolipid by ESI-MS/MS. Characteristic product ions were identified for each compound.