Determination of dichloroanilines in human urine by gas chromatography/mass spectrometry : validation protocol and establishment of Reference Values in a population group living in central Italy

3,4- and 3,5-Dichloroanilines (DCAs) are common markers of some non-persistent pesticides, e.g. linuron, diuron, vinclozolin, and iprodione. The general population may be exposed to these DCAs and/or their precursors mainly through diet. Since adverse effects on human health, such as endocrine disruption, have been reported, biological monitoring is essential for exposure assessment both of occupationally exposed subjects and of the general population. A highly sensitive and selective gas chromatography/mass spectrometry (GC/MS) method has been developed for the determination of 3,4- and 3,5-DCAs in urine using 4-chloro-2-methylaniline as an internal standard. The selected ion monitoring (SIM) mode was employed for quantitation of the analytes. The sample treatment procedure is simple and fast and no derivatization is required. The overall method was validated including uncertainty measurement. The limit of detection (LOD) and the lower limit of quantitation (LLOQ) were determined to be 0.005 and 0.010 microg/L for both analytes. The method was then applied to the establishment of reference values for a population group living in a rural area of central Italy (Novafeltria, Marche). A total of 151 out of 153 samples were found to be positive for 3,5-DCA, and 81.7% were positive for 3,4-DCA. For this group, 3,4-DCA levels ranged from 0.01 to 6.19 microg/L, while 3,5-DCA urinary concentrations were between 0.02 and 6.71 microg/L.