Reversed-phase high-performance liquid chromatographic method for the quantitation of endogenous folate catabolites in rat urine.

We describe a reversed-phase high-performance liquid chromatographic procedure for the analysis of rat urine for p-aminobenzoylglutamate (pABGlu) and its acetamido derivative (p-acetamidobenzoylglutamate, apABGlu). These two catabolites arise following the in vivo cleavage of the folate molecule at the C-9-N-10 bond. Known quantities of high-specific-activity tritiated forms of the catabolites are added as internal standards to aliquots of rat urine. Following preliminary sample clean-up on C18 Sep-Pak cartridges, including derivatisation in the case of pABGlu, the urinary extracts are quantitated by HPLC. The present assay makes possible for the first time the determination of endogenous folate breakdown in the rat.