Simultaneous Determination of Six Urinary Metabolites of Aromatic Compounds by HPLC

KeyWord:HPLC  urine  metabolite  liquid-liquid extraction

Abstract:

      A high performance liquid chromatographic(HPLC) method was established for the simultaneous determination of trans,trans-muconic acid(tt-MA)£¬hippuric acid(HA)£¬mandelic acid(MA)£¬phenylglyoxy acid(PGA)£¬p-aminophenol(PAP)and p-nitrophenol(PNP)in urine£®The chromatographic separation and the detection conditions were optimized£¬and a programmed gradient which the wavelength was changed with the time was applied for the detection£®The optimal pretreatment conditions for the sample were as follows:extraction solvent: 5 mL dichloromethane-isopropanol(7¡Ã3)£¬sample volume:5 mL£¬NaCl:0.75 g£¬extraction time:2 min£®The sample was firstly extracted under acidic condition£¬and repeated once under the neutral condition£¬then the two extraction liquids were merged£®The extract liquor was volatilized to dryness in water bath with the volatilization temperature less than 70 ¡æ£¬then redissolved with 1.0 mL mobile phase.Under the optimal conditions£¬ the linear ranges of the six components were up to 103£¬and the correlation coefficients(r) were not less than 0.999 3£®The detection limits(S/N=3) were in the range of 3-30 ¦Ìg/L£®The spiked recoveries were in the range of 86%- 104% with relative standard deviations less than 5%£®The method was sensitive£¬accurate and rapid£¬and could meet the needs of biological monitoring on the metabolites of harmful substances in urine.