Determination of trans,trans-muconic acid in urine: validation of a high performance liquid chromatographic method

Abstract

The main purpose of biological monitoring is to protect the workers's health, preventing the toxic effects that can occur from occupational exposure to chemicals. Trans, trans-muconic acid (ttMA), a benzene metabolite, has been recommended as a sensitive bioindicator in the biological monitoring of workers exposed to this solvent. This work was developed in order to validate a method for ttMA analysis in urine aiming its application on biomonitoring activities. The chosen technique was the high pressure liquid chromatography with reverse-phase column, Lichrosorb RP 18, and UV detection. A linear relationship (r²= 0.9943) was observed in the range from 0.2 to 5.0 mg/L. The detection and quantification limits were respectively 0.1 and 0.2 mg/L. The average recovery was 77.1% and the inaccuracy (bias) was 27.9%. Precision, evaluated by variation coefficient, were 7.7% (intra-assay) and 10.6% (inter-assay). The ttMA remained stable in the matrix during a period of six weeks for the 0.2 mg/L samples and fifteen weeks for the 2.0 mg/L samples, in both cases when stored at -20 ºC. In 0.2, 2.0 and 5.0 spiked samples no significant differences were found when conserved at 4 ºC for ten days. When using the method to analyse post shift samples from benzene handling workers, the mean and median results found were, respectively, 0.8 and 0.6 mg ttMA/g creatinine.