Analysis of inositol by high-performance liquid chromatography

A high-performance liquid chromatographic (HPLC) method has been developed for identification and quantification of inositol isomers and monosaccharides in inositol-containing glycans. The method, which can determine 10 pmol of inositol, utilizes an Aminex HPX-87C column packed with an 8% crosslinked cation-exchange resin in the calcium form eluted with deionized water at 50 degrees C. NaOH solution is added to the column effluent through a postcolumn tee to increase the pH (pH greater than 11.6) before entering a pulsed amperometric detector which is highly sensitive for polyhydroxylated compounds. Samples in which inositol is linked to sugar through a glycosidic bond are hydrolyzed with 5.5 N trifluoroacetic acid, 100 degrees C, 4 h, and then reduced with NaBH4. Samples in which inositol is linked via a phosphate ester are hydrolyzed with 6 N HCl, 110 degrees C, 24 h. This method has been applied to the analysis of inositol in the hamster prion proteins (PrP) PrP27-30, and PrPSc.