Analysis of 3,5‐dichloroaniline as a biomarker of vinclozolin and iprodione in human urine using liquid chromatography/triple quadrupole mass spectrometry

Abstract

The fungicides vinclozolin and iprodione are widely used in agriculture. These pesticides are dicarboximide fungicides containing the common moiety 3,5-dichloroaniline (3,5-DCA). It has been suggested that low-level exposures to such compounds may be associated with adverse health effects such as endocrine disruption.In this study a method using liquid chromatography triple quadrupole mass spectrometry (LC-MS/MS) was developed for the analysis of 3,5-DCA as a biomarker of exposure to these fungicides in human urine. The urine samples were treated by basic hydrolysis to degrade the fungicides, their metabolites and conjugates to 3,5-DCA. The 3,5-DCA was then extracted using toluene and derivatized using pentafluoropropionic anhydride (PFPA). Analysis of the derivative was carried out using selected reaction monitoring (SRM) in the negative ion mode. Quantification of the derivative was performed using [ 13 C 6 ] labeled 3,4-DCA as an internal standard with good precision and linearity in the range 0.1-200 ng/mL urine. The limit of detection was determined to 0.1 ng/mL. The metabolites in urine were found stable at storage in -20°C. To validate 3,5-DCA as a biomarker the method was applied in a human experimental exposure to iprodione and vinclozolin. Two healthy volunteers received 200 μg single oral doses of each pesticide followed by urine sampling during 72-120h post exposure.Between 78-107 % of the dose was recovered as 3,5-DCA in the urine after exposure.The dicarboximide fungicides vinclozolin, iprodione and procymidone are commonly used for the control of diseases on fruits, vegetables, ornamental plants and turf grasses. Human exposure to these fungicides can occur from occupational use but the general public may also be exposed via ingestion of contaminated produce.Vinclozolin has been classified as an endocrine disrupting chemical and it has been suggested that low-level exposures may be associated with adverse health effects. Vinclozolin is an androgen receptor antagonist with effects shown in adult, pubertal and neonatal rodents. [1][2][3][4] In addition, epigenetic transgenerational effects have been observed in rats. 5 The US EPA suggests that vinclozolin-induced malformations of the male reproductive tract are highly plausible to occur in humans. 6 However, only one report on human effects has been published and in that no antiandrogenic or reproductive effects were found after occupational exposure to vinclozolin. 7 Iprodione is chemically similar in structure to vinclozolin but there are few reports on its toxicity. In a study on rats by Gray et al.,8 no maternal or fetal endocrine toxicity was observed. The two fungicides contain the common moiety 3,5-dichloroaniline (3,5-DCA), which also has been found as a metabolite after exposure, this isomer has been shown to be nephrotoxic. 9 A prerequisite for epidemiological studies, and for determination of exposure-response relationships, is an accurate measurement of exposure. There is a lack of human exposure and effect data for fungicides overall, i...

The role of biotransformation and oxidative stress in 3,5-dichloroaniline (3,5-DCA) induced nephrotoxicity in isolated renal cortical cells from male Fischer 344 rats

“…Toxicity arising from exposure to anilines, important chemical intermediates used in the production of agricultural, industrial and pharmaceutical products (Lindh et al, 2007; Unger, 1996), has been well established. Exposure to aniline and its chlorinated derivatives has been associated with hematotoxicity (methemoglobinemia, hemolytic anemia; Chhabra et al, 1990; Guilhermino et al, 1998; Pauluhn et al 2004; Valentovic et al, 1997), splenotoxicity (splenomegaly, elevated erythropoietic activity, hyperpigmentation, fibrosis; Chhabra et al, 1990; Khan et al, 1999; Ma et al, 2008, 2013), hepatotoxicity (hepatomegaly, elevated serum ALT/GPT levels, centralobular necrosis; Valentovic et al, 1995a, 1995b, 1992), and nephrotoxicity (Hong et al, 2000; Lo et al, 1990; Racine et al, 2014; Valentovic et al, 1995a).…”

Efficacy of electrolyzed water, chlorine dioxide and photocatalysis for disinfection and removal of pesticide residues from stone fruit

“…This pesticide is moderately persistent in soil with a half-life time of 7-60 d depending on the environmental conditions (Carmona et al, 2001). In plants, it is quickly degraded once adsorbed by the roots with the formation of 3,5-dichloroaniline as the main metabolite (Athiel et al, 1995, Lindh et al, 2007. This generated metabolite is highly nephrotoxic (Lo et al, 1990) and carcinogenic.…”

Nephrotoxic Potential of Putative 3,5-Dichloroaniline (3,5-DCA) Metabolites and Biotransformation of 3,5-DCA in Isolated Kidney Cells from Fischer 344 Rats

“…Exposure to chloroanilines occurs in occupational settings, via their release or formation during mammalian metabolism of numerous compounds [1][2][3], or by environmental degradation of pesticides [1,[4][5][6]. The detection of chloroanilines in human urine or blood has been used as a biomarker for exposure to chloroaniline-based pesticides [7][8][9][10]. Toxicity associated with exposure to chloroanilines includes hematotoxicity (e.g., anemia or methemoglobinemia) [11][12][13], splenotoxicity [11,14], hepatotoxicity [14][15][16][17], endocrine disruption [18] and nephrotoxicity [16,19,20].…”