Quantitative determination of hippuric and m-methylhippuric acids in urine by high-speed liquid chromatography.

High-speed liquid chromatography employing an ultraviolet photometric detector has been applied to the simultaneous determination of hippuric and m-methylhippuric acids in urine. Reversed-phase partition chromatography is carried out on a muBondapak C18 column with methanol-water as the eluent system. This method obviates the necessity for isolation or reaction of these acids before assay. The only pretreatment necessary is extraction of sample with ethyl acetate. A linear relationship is obtained between the peak heights and the hippuric or m-methylhippuric acid concentrations. Mean recovery of hippuric and m-methylhippuric acids in urine is 99.8% and 99.3% respectively. The determination of hippuric acid by this method gives lower concentrations in normal urine than does the colorimetric method of Umberger and Fiorese (1963).