Quantitative determination ofhippuric and m-methylhippuric acids inurine byhigh-speed liquid chromatography

High-speed liquid chromatography employing an ultraviolet photometric detector has beenapplied tothesimultaneous determination ofhippuric andm-methylhippuric acids inurine. Reversed-phase partition chromatography iscarried outon a ,tBondapak C18columnwith methanol-water astheeluent system. Thismethod obviates thenecessity forisolation orreaction of these acids before assay.Theonly pretreatment necessaryisextraction ofsamples withethyl acetate. A linear relationship isobtained between thepeakheights andthehippuric orm-methylhippuric acidconcentrations. Meanrecoveryofhippuric andm-methylhippuric acids inurine is99-8 % and 99.3%respectively. Thedetermination ofhippuric acid bythis methodgives lower concentrations in normal urine thandoesthecolorimetric methodofUmberger andFiorese (1963). Ithasbeenobserved thattheconcentration ofhip- puric andm-methylhippuric acids inurine correlates wellwithexposure ofthesubject totoluene andm- xylene (Pagnotto andLieberman, 1967;Ikedaand Ohtsuji, 1969; Ogataetal., 1970). Manymethods havebeenusedforthedeter- mination ofthese metabolites usingcolorimetry (Gaffney etal., 1954; Umberger andFiorese, 1963), fluorometry (Ellman etal., 1961), andultraviolet spectrophotometry