Unexpected, ubiquitous exposure of pregnant Brazilian women to diisopentyl phthalate, one of the most potent antiandrogenic phthalates.

BACKGROUND: Human exposure to phthalates and other non-persistent chemicals in developing countries is largely unknown. A preliminary analysis of urinary samples from pregnant Brazilian women revealed the presence of metabolites of Diisopentyl phthalate (DiPeP). OBJECTIVES: Reliably quantify DiPeP metabolites in human urine and investigate the potential antiandrogenic activity of this phthalate in rats. METHODS: We initiated a pilot pregnancy cohort in Curitiba, Brazil, to examine phthalate exposure in urine samples collected in early pregnancy (n = 50) or pooled samples from early, mid and late pregnancy (n = 44). Our well established phthalate method was modified to include the primary DiPeP metabolite, monoisopentyl phthalate (MiPeP), and two additional secondary oxidized metabolites, 3OH-MiPeP and 4OH-MiPeP. In a parallel approach, we orally exposed pregnant rats to DiPeP or Di-n-butyl phthalate (DnBP; reference phthalate) at 0, 125, 250, and 500 mg/kg/day from gestation day 14 to 18 and measured ex vivo fetal testis testosterone production. RESULTS: We were able to detect and quantify specific DiPeP metabolites in nearly all (98%) of the early pregnancy urine samples and in all gestational pool samples with a median concentration for MiPeP of 3.65 and 3.15 µg/L, respectively, and for the two oxidized metabolites between 1.00 and 1.70 µg/L. All three urinary DiPeP metabolites were strongly correlated (r = 0.89 to 0.99). In the rat model, the effective dose (mg/kg/day) inhibiting fetal testosterone production by 50% (ED50 [95% confidence interval]) was 93.6 [62.9-139.3] for DiPeP which was significantly lower than for DnBP (220.3 [172.9-280.7]), highlighting the strong antiandrogenic potency of DiPeP within the spectrum of the phthalates. CONCLUSIONS: We unveiled and confirmed the exposure of pregnant Brazilian women to DiPeP via specific urinary metabolites. This unexpected and ubiquitous DiPeP exposure indicates to unique DiPeP exposure sources in Brazil. These exposures spark considerable concern because DiPeP is one of the most potent antiandrogenic phthalates.

Analysis of 3',5'-dichloro-2,3,4-trihydroxy-2-methylbutylanilide (DTMBA) as a new potential biomarker of exposure to vinclozolin in urine.

Vinclozolin (V) is a fungicide with anti-androgenic properties whose metabolism is not fully understood, and data on urinary elimination of either V or its metabolites are limited. Therefore the kinetics of urinary elimination of V and its metabolites, after an oral dose in adult male rats were investigated. A single oral dose of V (100 mg/kg) suspended in corn oil was administered to male adult Wistar rats, and urine was collected at different times after dosing. V and its metabolites were extracted from urine, then enzymatically hydrolyzed using ß-glucuronidase/sulfatase of H. pomatia, and analyzed by HPLC/DAD. Urinary pharmacokinetic parameters were calculated using the analyte concentrations adjusted by creatinine levels. V and its metabolites 3',5'-dichloro-2,3,4-trihydroxy-2-methylbutylanilide (DTMBA, formerly denoted as M5), 2-[[(3,5-dichlorophenyl)-carbamoyl]oxy]-2-methyl-3-butenoic acid (M1), 3,5-dichloroaniline (M3), and 3',5'-dichloro-2-hydroxy-2-methylbut-3-enanilide (M2) were efficiently detected. The mean urine concentrations of V and M1 metabolite were fitted to a two-compartmental model for pharmacokinetic analysis. DTMBA approximately represented 88% of the total excreted metabolites, it was easily detected up to 168 h after dosing and its half-lives were 21.5 and 74.1 h, respectively. M1 was the second most abundant metabolite and was detected up to 144 h after being void. V and M3 were detected before 48 h, and M2 exhibited the lowest levels during the first 8 h after dosing. DTMBA, the most abundant V metabolite is quickly eliminated by urine, it is chemically stable, specific and could represent a useful alternative to be used as a biomarker of exposure to V.