An improved procedure for the quantitative determination and characterization of sulfatides in rat kidney and brain by high-performance liquid chromatography.

A significant improvement has been made in the desulfation step of our previously published HPLC determination of cerebrosides, sulfatides, and monogalactosyl diacylglycerols (Nonaka, G. and Kishimoto, Y., Biochim. Biophys. Acta 572 (1978) 423-431). Instead of the original two-phase reaction, a solution of trifluoroacetic acid in ethyl acetate is used for the solvolysis in the new method. The revised method was used to determine the levels of cerebrosides and sulfatides in rat kidney. Among four individual glycosphingolipids studied, hydroxysulfatide was present at the highest level (0.7-1.3 nmol/mg of dry tissue), followed by nonhydroxysulfatide (0.3-0.8 nmol/mg of dry tissue). Hydroxycerebroside (0.09-0.16 nmol/mg of dry tissue) and nonhydroxycerebroside (0.03-0.09 nmol/mg of dry tissue) were present in smaller quantities. There appear to be no significant differences between male and female animals of different ages (30-120 days), although the amounts decreased slightly in older animals and there was a higher concentration in female than in male kidney. Tissue size was significantly smaller in females. The homolog composition of rat kidney sulfatide was studied by reverse-phase HPLC, and was found to be significantly different from that reported in human kidney. Rat sulfatides contained fatty acids with a higher degree of saturation and longer chain length. Preliminary studies indicated that rat kidney contained unusually large quantities of C25:1 and C27:1 fatty acids and also that there was more C26:1 than C24:1 acid. In brain of the same animals the ratio of nonhydroxy to hydroxysulfatide decreased with age (1.5:1 in 30-day-old brain; 1:1 at 90 days).