Biotransformation of 2,4,6-tris(2,4,6-tribromophenoxy)-1,3,5-triazine (TTBP-TAZ) can contribute to high levels of 2,4,6-tribromophenol (2,4,6-TBP) in humans

Abstract

2,4,6-Tribromophenol (2,4,6-TBP) is a brominated flame retardant that accumulates in human tissues and is a potential toxicant. Previous studies found 2,4,6-TBP levels in human tissues were significantly higher than those of brominated flame retardants measured in the same samples. In contrast, the levels of 2,4,6-TBP in the environment and foodstuff are not elevated, suggesting a low potential for direct intake through environmental exposure or diet. Here, we hypothesized that high levels of 2,4,6-TBP in human tissues are partially from the indirect exposure sources, such as biotransformation of highly brominated substances. We conducted in vitro assays utilizing human and rat liver microsomes to compare the biotransformation rates of four highly brominated flame retardants, which could potentially transform to 2,4,6-TBP, including decabromodiphenyl ethane (DBDPE), 2,4,6-tris-(2,4,6-tribromophenoxy)-1,3,5-triazine (TTBP-TAZ), 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), and tetrabromobisphenol A (TBBPA). Our results show that TTBP-TAZ rapidly metabolizes in both human and rat liver microsomes with a half-life of 1.1 and 2.2 h, respectively, suggesting that TTBP-TAZ is a potential precursor of 2,4,6-TBP. In contrast, 2,4,6-TBP was not formed as a result of biotransformation of TBBPA, BTBPE, and DBDPE in both human and rat liver microsomes. We applied suspect and target screening to explore the metabolic pathways of TTBP-TAZ and identified 2,4,6-TBP as a major metabolite of TTBP-TAZ accounting for 87% of all formed metabolites. These in vitro results were further tested by an in vivo experiment in which 2,4,6-TBP was detected in the rat blood and liver at concentrations of 270 ± 110 and 50 ± 14 μg/g lipid weight, respectively, after being exposed to 250 mg/kg body weight/day of TTBP-TAZ for a week. The hepatic mRNA expression demonstrated that TTBP-TAZ significantly activates the aryl hydrocarbon receptor (AhR) and promotes fatty degeneration (18 and 28-fold change compared to control, respectively) in rats.

Keywords: 2,4,6-TBP; Biotransformation; In vitro; In vivo; TTBP-TAZ.

Figure 1.

The extracted chromatograms of m/z 329.7 for 2,4,6-TBP standard, the control sample and the dosed sample isolated from TTBP-TAZ incubations (A); mass spectra and proposed structures of the metabolite in the 2,4,6-TBP standard and in the dosed sample (B).

Figure 2.

Proposed pathways of TTBP-TAZ hepatic biotransformation. Solid arrows demonstrate pathways determined in this study and dashed arrows demonstrate pathways reported in (Ho et al., 2015).

Figure 3.

Concentrations of 2,4,6-TBP in rat serum and liver (μg/g lw) of the control group, 2,4,6-TBP exposure group, and TTBP-TAZ exposure group (mean ± standard errors [SE]; n = 5). The bars for the control group are not shown because 2,4,6-TBP was not detected in this group.

Figure 4.

mRNA expression (mean ± standard errors [SE]; n = 5) of genes encoding for thyroid functions, nuclear receptors, oxidative stress, angiogenesis, inflammation, and lipid metabolism in rat livers of the control group, 2,4,6-TBP exposure group, and TTBP-TAZ exposure group. * indicates a statistical difference from control and # indicates a statistical difference from 2,4,6 TBP at p <0.05 (determined using ANOVA followed by Dunnett’s post-hoc test).