Sex-specific effects of bisphenol S with tissue-specific responsiveness in adult zebrafish: The antiandrogenic and antiestrogenic effects.

This study investigates the adverse effects and the associated underlying mechanism of bisphenol S (BPS) exposure on reproductive endocrine activity in adult zebrafish. Fish were exposed for 21 days to different BPS concentrations (0, 8, 40, and 200 µg/mL) determined via the lowest observed adverse effect level (LOAEL, i.e., < EC15 = 250 µg/mL) for zebrafish embryos. Exposure to 200 µg/mL BPS in female zebrafish in the absence of vitellogenic oocytes or the presence of degenerated oocytes in the ovary significantly decreased the biosynthesis of hepatic vitellogenin (VTG) mRNA, while hepatic VTG mRNA in male fish abundance was significantly elevated (P < 0.05). The levels of gonadal steroids were significantly increased in female zebrafish, while in male zebrafish, the levels of endogenous androgens were reduced (P < 0.05). However, the activities of 17ß-estradiol and aromatase in male zebrafish were significantly elevated in all BPS exposure groups in male zebrafish (P < 0.05). Interestingly, thyroid hormone levels and residual whole-body BPS levels increased in female and male zebrafish with increasing exposure concentrations. A novel finding is that the response to BPS depends on zebrafish sex and tissue-specific responsiveness to the accumulation of BPS, suggesting that BPS may cause long-term environmental problems in adult zebrafish through tissue-specific suppression and hormonal imbalance.

Urinary di(2-ethylhexyl)phthalate metabolite ratios in obese children of South Korea.

Phthalate exposure has been reported to be more associated with obesity in children than in adults. The concentration of di(2-ethylhexyl)phthalate (DEHP) was high temporal variability in spot urine, so additional tools of assessing DEHP exposure were required. Therefore, we used relative metabolite ratios (RMRs) as well as concentrations, and RMRs did not need to be corrected to the creatinine concentration. We aimed to evaluate the levels of urinary DEHP metabolites and their RMRs in obese children in South Korea, and to investigate the potential of RMRs for assessing the risks for childhood obesity. We analyzed the four urinary DEHP metabolites (mono-2-ethylhexyl phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono-(2-ethyl-5-carboxypentyl) phthalate (MECPP)) in 240 children aged 5-16 years, using isotope dilution GC-MS/MS. The children were placed into three groups ("normal weight," "overweight," and "obese") according to body mass index (BMI) percentiles. We statistically compared the concentrations and RMRs of DEHP metabolites among these groups. The obese group had lower MEHP levels, and higher secondary metabolite (MEHHP, MEOHP, and MECPP) levels, than the normal weight group. DEHP metabolite levels did not differ significantly between the normal weight and obese groups, whereas RMRA2 (as the ratio of the molar concentrations of MEOHP to MEHHP) was found to be negatively associated with BMI percentile (ß= -0.236, p <0.01) and weight percentile (ß= -0.282, p<0.001). Therefore, we suggest that RMRs are an additional tool for assessing the health risks of DEHP.

Reproductive dysfunction linked to alteration of endocrine activities in zebrafish exposed to mono-(2-ethylhexyl) phthalate (MEHP).

This study aimed to investigate the effect of mono-(2-ethylhexyl) phthalate (MEHP), one of the major phthalate metabolites that are widespread in aquatic environments, on reproductive dysfunction, particularly on endocrine activity in adult male and female zebrafish. For 21 days, the zebrafish were exposed to test concentrations of MEHP (0, 2, 10, and 50 µg/mL) that were determined based on the effective concentrations (ECx) for zebrafish embryos. Exposure to 50 µg/mL MEHP in female zebrafish significantly decreased the number of ovulated eggs as well as the hepatic VTG mRNA abundance when those of the control group. Meanwhile, in female zebrafish, the biosynthetic concentrations of 17ß-estradiol (E2) and the metabolic ratio of androgen to estrogen were remarkably increased in all MEHP exposed group compared with those in the control group, along with the elevated levels of cortisol. However, no significant difference was observed between these parameters in male zebrafishes. Therefore, exposure to MEHP causes reproductive dysfunction in female zebrafishes and this phenomenon can be attributed to the alteration in endocrine activities. Moreover, the reproductive dysfunction in MEHP-exposed female zebrafishes may be closely associated with stress responses, such as elevated cortisol levels. To further understand the effect of MEHP on the reproductive activities of fish, follow-up studies are required to determine the interactions between endocrine activities and stress responses. Overall, this study provides a response biomarker for assessing reproductive toxicity of endocrine disruptors that can serve as a methodological approach for an alternative to chronic toxicity testing.

Correction: Percentage fractions of urinary di(2-ethylhexyl) phthalate metabolites: Association with obesity and insulin resistance in Korean girls.

[This corrects the article DOI: 10.1371/journal.pone.0208081.].

Percentage fractions of urinary di(2-ethylhexyl) phthalate metabolites: Association with obesity and insulin resistance in Korean girls.

OBJECTIVE: We assessed the associations of percentage fractions of urinary di(2-ethylhexyl) phthalate (DEHP) metabolites with obesity and insulin resistance in Korean girls. METHODS: In total, 137 girls, aged 6 to 13 years (65 overweight cases and 72 controls), were recruited. Anthropometric indices and the homeostatic model assessment of insulin resistance (HOMA-IR) index were determined. Four major urinary DEHP metabolites were analyzed in spot urine samples by gas chromatography-tandem mass spectrometry, including mono(2-ethylhexyl) phthalate, mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono(2-ethyl-5-carboxypentyl) phthalate. RESULTS: There were no significant differences in the urinary concentrations of the DEHP metabolites between the overweight and control groups. The percentage fraction of MEHHP (MEHHP%) among all DEHP metabolites was significantly higher in the overweight prepubertal girls than in the controls (P = 0.035). MEHHP% was positively associated with the body mass index percentile, waist circumference, body fat percentage, and HOMA-IR index in the prepubertal girls. After adjusting for covariates, the prepubertal girls in a higher MEHHP% quartile were found to have a higher odds ratio for central obesity than those in a lower quartile (odds ratios: 5.05 for quartile 3; 7.30 for quartile 4). The relative rate of MEHHP oxidation to MEOHP was negatively associated with the body mass index percentile and waist circumference in the prepubertal girls. However, no such association was observed in the pubertal girls. CONCLUSIONS: MEHHP% was positively associated with obesity and insulin resistance in prepubertal girls. Further studies are necessary to elucidate the causal links between altered phthalate metabolism and increased susceptibility to insulin resistance in children.

Effective and sensitive determination of eleven disinfection byproducts in drinking water by DLLME and GC-MS.

The aim of this study was to optimize the dispersive liquid-liquid microextraction (DLLME) parameters for simultaneous analysis through DLLME-gas chromatography-mass spectrometry (GC-MS) of six iodo-trihalomethanes, four haloacetonitriles, and one halonitromethane, which are residual disinfection products found in drinking water. Eleven disinfection by-product (DBPs) remaining in aqueous samples were extracted and concentrated using a simple, rapid, and environmentally friendly DLLME method, and then analyzed simultaneously by GC-MS. The optimized DLLME parameters were a sample volume of 5 mL, 100 µL of dichloromethane as the extraction solvent, 1 mL of methanol as the dispersion solvent, an extraction time of 60 s, and 1.5 g of sodium chloride for the salting out effect. The enrichment factor values obtained using the established DLLME-GC-MS method were 19.8-141.5, and the limit of detection and limit of quantification were 0.22-1.19 µg/L and 0.75-3.98 µg/L, respectively. The calibration curves had correlation coefficients (r2) of 0.9958-0.9992 in the concentration range of 0.5-40 µg/L, and they exhibited good linearity in quantitative analysis. This new method could be useful for analyzing eleven DBPs that remain in drinking water.

Analysis of 3-MCPD and 1,3-DCP in Various Foodstuffs Using GC-MS.

3-Monochloro-1,2-propanediol (3-MCPD) and 1,3-dichloro-2-propanol (1,3-DCP) are not only produced in the manufacturing process of foodstuffs such as hydrolyzed vegetable proteins and soy sauce but are also formed by heat processing in the presence of fat and low water activity. 3-MCPD exists both in free and ester forms, and the ester form has been also detected in various foods. Free 3-MCPD and 1,3-DCP are classified as Group 2B by the International Agency for Research on Cancer. Although there is no data confirming the toxicity of either compound in humans, their toxicity was evidenced in animal experimentation or in vitro. Although few studies have been conducted, free 3-MCPD has been shown to have neurotoxicity, reproductive toxicity, and carcinogenicity. In contrast, 1,3-DCP only has mutagenic activity. The purpose of this study was to analyze 3-MCPD and 1,3-DCP in various foods using gas chromatography -mass spectrometry. 3-MCPD and 1,3-DCP were analyzed using phenyl boronic acid derivatization and the liquid-liquid extraction method, respectively. The analytical method for 3-MCPD and 1,3-DCP was validated in terms of linearity, limit of detection (LOD), limit of quantitation, accuracy and precision. Consequently, the LODs of 3-MCPD and 1,3-DCP in various matrices were identified to be in the ranges of 4.18~10.56 ng/g and 1.06~3.15 ng/g, respectively.

Biomonitoring of urinary di(2-ethylhexyl) phthalate metabolites of mother and child pairs in South Korea.

Di(2-ethylhexyl) phthalate (DEHP) is one of the common phthalate plasticizers used primarily in soft polyvinyl chloride, which is a plastic polymer that makes up the total weight of goods from 1% up to 40% in many consumer products. The aims of this study were to examine the urinary DEHP metabolites in South Korean children and to investigate the correlation between mother and child DEHP urine excretion. Three kinds of urinary DEHP metabolites: mono(2-ethylhexyl) phthalate (MEHP), mono(2-ethyl-5-hydroxyhexyl) phthalate (5-OH-MEHP) and mono(2-ethyl-5-oxohexyl) phthalate (5-oxo-MEHP), were analyzed. The total of 954 samples (nChildren=392, nMothers=265, nAadults=297), including 258 mother and child pairs, were analyzed using isotope dilution gas chromatography-mass spectrometry. Many studies present higher concentration of DEHP metabolites detected from adults in reproductive age than adults in other ages. Therefore, adults who are age-matched to mothers were evaluated to serve as a standard of comparison against mothers. All statistical analysis was made by adjusting detected volume concentrations (µg/L) with respect to creatinine concentrations (mg/dL) since urinary DEHP metabolites were studied using human reference. The difference in median levels of sum of urinary DEHP metabolites was only significant when children were analyzed in relation to region (p-value≤0.005). Among the three DEHP metabolites, only MEHP of children was significantly correlated to that of paired mothers (p-value≤0.01). The present paper defines the relative metabolic rate (RMR) of DEHP metabolism for the first time in study on phthalates. Children had faster RMR than mothers and adults, specifically in the first step of DEHP metabolism (RMR1: MEHP hydroxylation to 5-OH-MEHP), and RMR1 of children between 1 and 24 months was the fastest. The above results may be used to study and assess human health risk from DEHP exposures, especially among mothers and children in Korea.