Metabolism Of Di(2-ethylhexyl) Phthalate Research Articles

Di(2-ethylhexyl) phthalate disrupts circadian rhythm associated with changes in metabolites and cytochrome P450 gene expression in Caenorhabditis elegans

The plasticizer di(2-ethylhexyl) phthalate (DEHP) is a widespread environmental pollutant due to its extensive use. While circadian rhythms are inherent in most living organisms, the detrimental effects of DEHP on circadian rhythm and the underlying mechanisms remain largely unknown. This study investigated the influence of early developmental exposure to DEHP on circadian rhythm and explored the possible relationship between circadian disruption and DEHP metabolism in the model organism Caenorhabditis elegans. We observed that DEHP disrupted circadian rhythm in a dose-dependent fashion. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed that DEHP-induced circadian disruption accompanies with altered proportions of DEHP metabolites in C. elegans. RNA sequencing data demonstrated that DEHP-induced circadian rhythm disruption caused differential gene expression. Moreover, DEHP-induced circadian disruption coincided with attenuated inductions of DEHP-induced cytochrome P450 genes, cyp-35A2, cyp-35A3, and cyp-35A4. Notably, cyp-35A2 mRNA exhibited circadian rhythm with entrainment, but DEHP exposure disrupted this rhythm. Our findings suggest that DEHP exposure disrupts circadian rhythm, which is associated with changes in DEHP metabolites and cytochrome P450 gene expression in C. elegans. Given the ubiquitous nature of DEHP pollution and the prevalence of circadian rhythms in living organisms, this study implies a potential negative impact of DEHP on circadian rhythm and DEHP metabolism in organisms.

Phthalate Biomarkers Composition in Relation to Fatty Liver: Evidence from Epidemiologic and in vivo studies

Phthalates, classified as environmental endocrine disruptors, pose potential toxicity risks to human health. Metabolic dysfunction-associated fatty liver disease is one of the most widespread liver diseases globally. Compared to studies focusing on metabolic disorders in relation to pollutants exposure, the impact of individual factors such as fatty liver on the in vivo metabolism of pollutants is always overlooked. Therefore, this study measured concentrations and composition of phthalate monoesters (mPAEs) in human urine samples, particularly those from fatty liver patients. Furthermore, we induced fatty liver in male Wistar rats by formulating a high-fat diet for twelve weeks. After administering a single dose of DEHP at 500 mg/kg bw through gavage, we compared the levels of di-2-ethylhexyl phthalate (DEHP), its metabolites (mDEHPs) and three hepatic metabolic enzymes, namely cytochrome P450 enzymes (CYP450), UDP glucuronosyltransferase 1 (UGT1), and carboxylesterase 1 (CarE1), between the normal and fatty liver rat groups. Compared to healthy individuals (n = 75), fatty liver patients (n = 104) exhibited significantly lower urinary concentrations of ∑mPAEs (median: 106 vs. 166 ng/mL), but with a higher proportion of mono-2-ethylhexyl phthalate in ∑mDEHPs (25.7 % vs. 9.9 %) (p < 0.05). In the animal experiment, we found that fatty liver in rats prolonged the elimination half-life of DEHP (24.61 h vs. 18.89 h) and increased the contents of CYP450, CarE1, and UGT1, implying the common but differentiated metabolism of DEHP as excess lipid accumulation in liver cells. This study provides valuable information on how to distinguish populations in biomonitoring studies across a diverse population and in assigning exposure classifications of phthalates or similar chemicals in epidemiologic studies.

Prenatal exposure to low-dose di-(2-ethylhexyl) phthalate (DEHP) induces potentially hepatic lipid accumulation and fibrotic changes in rat offspring

Di(2-ethylhexyl) phthalate (DEHP) is a plasticizer that is widely used to enhance the flexibility and durability of various products. As an endocrine disruptor, DEHP can interfere with normal hormonal functions, posing substantial health risks to organisms. Given the critical role of the liver in DEHP metabolism, we investigated potential liver damage in offspring induced by prenatal exposure to low doses of DEHP in Sprague Dawley rats. Pregnant rats were divided into three groups and administered 20 or 200 μg/kg/day of DEHP or corn oil vehicle control via oral gavage from gestation days 0–20. Male rat offspring were euthanized on postnatal day 84, and blood and liver specimens were collected for analysis. We observed fibrotic changes in the livers of the exposed groups, accompanied by the proliferation and activation of hepatic stellate cells and upregulated expression of TGF-B and collagen 1A1. Additionally, an inflammatory response, characterized by increased macrophage infiltration and elevated levels of pro-inflammatory cytokines, was evident. Third, hepatic and serum triglyceride and serum cholesterol were notably increased, along with upregulated expression of lipid metabolism-related proteins, such as sterol regulatory element-binding protein-1c, acetyl-CoA carboxylase, fatty acid synthase, and diacylglycerol O-acyltransferase 1, particularly in the low-dose group. These results suggest that prenatal exposure to DEHP can disrupt lipid metabolism, resulting in hepatic lipid accumulation in the offspring. This exposure may also induce an inflammatory response that contributes to the development of liver fibrosis. Thus, even at relatively low doses, such exposure can precipitate latent liver damage in offspring.

Evaluation of distinct molecular architectures and coordinated regulation of the catabolic pathways of oestrogenic dioctyl phthalate isomers in Gordonia sp.

Bacterial strain GONU, belonging to the genus Gordonia, was isolated from a municipal waste-contaminated soil sample and was capable of utilizing an array of endocrine-disrupting phthalate diesters, including di-n-octyl phthalate (DnOP) and its isomer di(2-ethylhexyl) phthalate (DEHP), as the sole carbon and energy sources. The biochemical pathways of the degradation of DnOP and DEHP were evaluated in strain GONU by using a combination of various chromatographic, spectrometric and enzymatic analyses. Further, the upregulation of three different esterases (estG2, estG3 and estG5), a phthalic acid (PA)-metabolizing pht operon and a protocatechuic acid (PCA)-metabolizing pca operon were revealed based on de novo whole genome sequence information and substrate-induced protein profiling by LC-ESI-MS/MS analysis followed by differential gene expression by real-time PCR. Subsequently, functional characterization of the differentially upregulated esterases on the inducible hydrolytic metabolism of DnOP and DEHP revealed that EstG5 is involved in the hydrolysis of DnOP to PA, whereas EstG2 and EstG3 are involved in the metabolism of DEHP to PA. Finally, gene knockout experiments further validated the role of EstG2 and EstG5, and the present study deciphered the inducible regulation of the specific genes and operons in the assimilation of DOP isomers.

In-depth profiling of di(2-ethylhexyl) phthalate metabolic footprints in rats using click chemistry-mass spectrometry probes

Di(2-ethylhexyl) phthalate (DEHP), the most widely used plasticizers in the world, has been regarded as an endocrine disrupting chemical with serious adverse health outcomes. Accumulating evidence strongly suggests that the undesirable biological effects of DEHP are meditated by its metabolites rather than itself. However, the metabolic footprints of DEHP in vivo are still unclear. Here we developed a click chemistry-assisted mass spectrometry (CC-MS) strategy for in-depth profiling DEHP metabolites in rats. An alkyne-modified DEHP analogue (alkyne-DEHP) was synthesized as a tracer for in vivo tracing, and a pair of MS probes (4-azido-nphenylbenzamide, 4-ANPA, and its deuterated reagent d5-4-ANPA) were prepared to specifically label the alkyne-DEHP metabolites, and prominently improve their detection sensitivity and selectivity. Using the CC-MS strategy, we successfully screened 247 alkyne-DEHP metabolites from rat urine, feces, and serum, including many unrevealed metabolites, such as oxidized phthalate diester metabolites and glucuronides of phthalate monoester metabolites. The discovery of new DEHP metabolites provides additional insights for understanding the metabolism of DEHP, which may be beneficial in exploring the mechanism underlying DEHP induced-toxicity in the future.

Overcompensation of CoA Trapping by Di(2-ethylhexyl) Phthalate (DEHP) Metabolites in Livers of Wistar Rats.

Di(2-ethylhexyl) phthalate (DEHP) is commonly used as a plasticizer in various industrial and household plastic products, ensuring widespread human exposures. Its routine detection in human bio-fluids and the propensity of its monoester metabolite to activate peroxisome proliferator activated receptor-α (PPARα) and perturb lipid metabolism implicate it as a metabolic disrupter. In this study we evaluated the effects of DEHP exposure on hepatic levels of free CoA and various CoA esters, while also confirming the metabolic activation to CoA esters and partial β-oxidation of a DEHP metabolite (2-ethyhexanol). Male Wistar rats were exposed via diet to 2% (w/w) DEHP for fourteen-days, following which hepatic levels of free CoA and various CoA esters were identified using liquid chromatography-mass spectrometry. DEHP exposed rats showed significantly elevated free CoA and increased levels of physiological, DEHP-derived and unidentified CoA esters. The physiological CoA ester of malonyl-CoA and DEHP-derived CoA ester of 3-keto-2-ethylhexanoyl-CoA were the most highly elevated, at eighteen- and ninety eight-times respectively. We also detected sixteen unidentified CoA esters which may be derivative of DEHP metabolism or induction of other intermediary metabolism metabolites. Our results demonstrate that DEHP is a metabolic disrupter which affects production and sequestration of CoA, an essential cofactor of oxidative and biosynthetic reactions.

Biodegradation patterns of the endocrine disrupting pollutant di(2-ethyl hexyl) phthalate by Fusarium culmorum

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer, which is considered an endocrine disrupting pollutant. Growth kinetics and esterases activity by biochemical tests and polyacrylamide gel electrophoresis were characterized for Fusarium culmorum grown in DEHP-supplemented (1000 mg/L) medium as the only carbon source and in control medium with glucose. Intermediate compounds of biodegraded DEHP were identified by GC-MS. F. culmorum degraded 92% of DEHP within 36 h. DEHP was degraded to butanol, hexanal, catechol and acetic acid. It is suggested that the first two compounds would transform into butanediol and the last two would enter into the Krebs cycle and would be mineralized to CO2 and H2O. DEHP induced eight esterase isoforms, which were different to those constitutive isoforms produced in the control medium. It is suggested that five enzymes (25.7, 29.5, 31.8, 97.6 and 144.5 kDa) detected during the first 36 h be involved in the primary biodegradation of DEHP. The rest of the enzymes (45.9, 66.6 and 202.9 kDa) might be involved in the final steps for DEHP metabolism. F. culmorum has a promising practical application in the treatment of DEHP-contaminated environments because it can secrete specific esterase to breakdown high concentrations of DEHP in a short period of time. This research represents the first approach for the study of esterase involved in the DEHP degradation by fungi using this phthalate as the sole source of carbon and energy.

Investigation on Metabolism of Di(2-Ethylhexyl) Phthalate in Different Trimesters of Pregnant Women.

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer but shows diverse toxicity. To investigate the time- and maternal age-differences in metabolism process of DEHP in pregnant women, three urine samples were collected from each pregnant woman ( n = 847) at the first (T1, mean 13.04 gestational weeks), the second (T2, mean 23.63 gestational weeks) and the third time point (T3, mean 35.91 gestational weeks), respectively. Four metabolites of DEHP were analyzed in 2541 urine samples (847 × 3) by using ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry. The level of urinary mono(2-ethylhexyl) phthalate (MEHP) had a decreasing trend across the pregnancy periods. The geometric mean concentrations of mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono(2-ethyl-5-oxohexyl) phthalate (MEOHP) and mono(2-ethyl-5-carboxypentyl) phthalate (MECPP) were significantly decreased in T2 than T1, and recovered slightly in T3. The transformation rate of MEHP to MEHHP in T3 was significantly higher than those in other two time points. The transformation rate of MEHHP to MEOHP in T2 and T3 was significantly higher than that in T1, indicating the oxidation was more efficient in late pregnancy compared with early and middle pregnancy. The percentages of oxidation products MEHHP and MECPP were higher in the higher-age group compared with the lower-age group in the second trimester samples. It is therefore concluded that DEHP metabolism may be influenced by the pregnancy stage and maternal age. The findings may benefit the risk assessment and toxicity evaluation of DEHP.

Fate of di (2‑ethylhexyl) phthalate in different soils and associated bacterial community changes

Di (2‑ethylhexyl) phthalate (DEHP) is a ubiquitous organic pollutant, which has caused considerable pollution in arable soils. In this study, the relationship between DEHP degradation potential and soil properties in 12 agricultural soils (S1-S12) was examined in a microcosm based experiment. Six of these soils were then selected to monitor patterns in bacterial community responses. It was found that DEHP degradation was positively correlated with bacterial counts in the original soils, suggesting a key role for bacteria in degradation. However, DEHP metabolism did not always lead to complete degradation. Its monoester metabolite, mono (2‑ethylhexyl) phthalate (MEHP), was present at appreciable levels in the two acidic soils (S1 and S2) during the incubation period of 35 days. Based on high-throughput sequencing data, we observed a greater impact of DEHP contamination on bacterial community structure in acidic soils than in the other soils. Nocardioides, Ramlibacter and unclassified Sphingomonadaceae were enriched in the two near-neutral soils where degradation was highest (S4 and S7), suggesting that these organisms might be efficient degraders. The relative abundance of Tumibacillus was greatly reduced in 50% of the six soils examined, demonstrating a high sensitivity to DEHP contamination. Furthermore, putative organic-matter decomposing bacteria (including Tumebacillus and other bacteria taxa such as members from Micromonosporaceae) were greatly reduced in the two acidic soils (S1 and S2), possibly due to the accumulation of MEHP. These results suggest a crucial role of soil acidity in determining the fate and impact of DEHP in soil ecosystems, which deserves further investigation. This work contributes to a better understanding of the environmental behavior of DEHP in soil and should facilitate the development of appropriate remediation technologies.

Exposure to phthalates, bisphenol-a, parabens and organophosphates among mother-child pairs

Introduction: Organophosphates, phthalates, parabens and bisphenol-a are chemicals with applications as pesticides, solvents and plasticizers. The long term exposure, even at low doses, has been connected with serious adverse health effects. Early-life exposure is considered as the most critical. Urinary metabolites of the above-mentioned chemicals are used as exposure biomarkers. Methods: Exposure levels were assessed in a mother-child cohort (Crete, Greece) during pregnancy (6th month) and at children (2.5 and 4 years of age) by determining six parabens, bisphenol-a, seven phthalate and six dialkylphosphate metabolites in their urine. Spot samples (N=978) were analyzed with liquid chromatography-tandem mass spectrometry. Phthalates were determined in drinking water and indoor air of houses and car interiors of the study area. Results: Principal component analysis grouped exposure in three main sources: plastic (all phthalates-BPA), personal care products (di-ethyl phthalate-parabens) and food (organophosphates). There were correlations for specific metabolite (2-tailed Spearman, p&lt;0.001) between levels a) at pregnancy and at 2.5 year old children b) and children age/gender. Total daily intake was calculated; in general it was highest in mothers and lowest in 4 years old children. In some cases exceeded ESA and USEPA limits (e.g. 5% of all subjects exceeds USEPA-TDI for di-2ethylhexyl phthalate (DEHP)). Drinking water and indoor air contribution to the total phthalate daily intake was low. Relative rates of DEHP metabolism were calculated. Discussion: Pregnant women and children are exposed to mixtures of the above-mentioned chemicals. Some specific sources (three phthalates and ethyl paraben) are common during pregnancy and at 2.5 years old children. Although levels in our country in most of the studied chemicals are comparable with other studies, exposure is significant. Overall exposure and rate of DEHP metabolism are related with gender and age.

Associations of individual characteristics and lifestyle factors with metabolism of di-2-ethylhexyl phthalate in NHANES 2001–2012

BackgroundPrevious studies suggest that a higher ratio of primary to secondary metabolites of di-2-ethylhexyl phthalate (DEHP), reflective of a slower DEHP conversion rate, is associated with a greater physiologic effect. We examined associations of several individual characteristics and lifestyle factors with the ratio of mono-2-ethylhexyl phthalate to mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHP:MEHHP) and %MEHP (the ratio of MEHP to the sum of the secondary metabolites). MethodsWe used the data from the National Health and Nutrition Examination Survey, 2001–2012. The study included adults with BMI<30 and no diabetes. Pregnant women were excluded. We examined associations of age, race, gender, Body Mass Index, smoking, alcohol and caffeine consumption, medication use, cancer history, and menopausal status and postmenopausal hormone use (in women) with MEHP:MEHHP and %MEHP using multivariable linear regression. The values for %MEHP were log-transformed in the analysis. ResultsIn multivariable analysis, non-Caucasian individuals had higher %MEHP (non-Hispanic Blacks: β=0.114, 95% Confidence interval [CI]: 0.050, 0.177; Hispanic: β=0.089, 95% CI: 0.024, 0.154; other race: β=0.126, 95% CI: 0.033, 0.219). Age was inversely associated with MEHP:MEHHP (β=−0.001, 95% CI: −0.002, −0.001) and %MEHP (β=−0.006, 95% CI: −0.008, −0.004). Overweight individuals had lower MEHP: MEHHP and lower %MEHP (β=−0.035, 95% CI: 0.062, −0.008 and β=−0.104, 95% CI: −0.162, −0.046, respectively). Alcohol consumption was inversely associated with %MEHP among men (p-trend=0.03). ConclusionsIndividual and lifestyle characteristics are associated with differences in DEHP metabolism. Understanding underlying biological mechanisms could help to identify individuals at a greater risk of adverse effects from DEHP exposure.

Lentinula edodes Grown on Di(2-ethylhexyl) Phthalate-Containing Media: Mycelial Growth and Enzyme Activities

Di(2-ethylhexyl) phthalate (DEHP) is a plasticizer that is widely used to soften plastic products. It is an endocrine-disrupting compound, widely considered an environmental contaminant that is difficult to remove or degrade. The radial growth rate (ur), mycelial biomass, intracellular and extracellular activities of laccase and esterase, intracellular and extracellular contents of protein and glycogen, and the contents of soluble and insoluble glucans were evaluated in colonies of Lentinula edodes, grown in various concentrations of DEHP (0, 750, 1200, and 1500 mg/L) on agar. Glucose and yeast extracts were added to all media. The highest ur was shown in medium lacking DEHP, followed by that shown in the medium containing 1500 mg of DEHP/L. The greatest protein extracellular contents were observed in medium with added DEHP. The largest extracellular glycogen contents and mycelial biomass were found in media containing 1200 and 1500 mg of DEHP/L, respectively. These results suggest that DEHP was degraded and used as a substrate by L. edodes during diauxic growth (glucose metabolized first, followed by DEHP), and laccases were more important than esterases in the metabolism of DEHP. L. edodes can be used to remove DEHP from phthalate-polluted environments.

Phthalate esters, parabens and bisphenol-A exposure among mothers and their children in Greece (Rhea cohort)

Exposure to endocrine disruptors, used as additives, preservatives, plasticisers and solvents in numerous consumer products, might cause adverse health effects. Humans exposed to these chemicals, metabolise and excrete them mostly via urine. Urinary metabolite concentrations are used as biomarkers of exposure. We evaluated the exposure of 4-month pregnant women and their children at 2years of age to phthalates, parabens and bisphenol-A. Concentrations of eight phthalate metabolites, six parabens and bisphenol-A were measured in 239 mother–child pairs of the “Rhea” cohort in Greece. Concentration levels in mother and children were comparable with corresponding concentrations in other countries worldwide. Low Spearman correlation coefficients (CC 0.1–0.2, p-value<0.01) were observed for di-ethyl phthalate (DEP), di-n-butyl phthalate (DnBP), butyl-benzyl phthalate (BBP) and ethyl paraben (EPB) between mothers and their children. We observed higher median daily intake (DIu) for mothers (e.g. di-ethyl phthalate 6.9μgd−1kg−1) than for their children (1.4μgd−1kg−1) for all examined compounds, except for di-2-ethylhexyl phthalate (DEHP) and bisphenol-A. Principal component analysis (PCA) indicated two main sources of exposure (plastic related and personal care-hygiene products) for phthalates, parabens and bisphenol-A. Differences in DEHP metabolism were observed among mothers–children and female–male children.

Di-(2-ethylhexyl) phthalate metabolites in urine show age-related changes and associations with adiposity and parameters of insulin sensitivity in childhood.

ObjectivesPhthalates might be implicated with obesity and insulin sensitivity. We evaluated the levels of primary and secondary metabolites of Di-(2-ethylhexyl) phthalate (DEHP) in urine in obese and normal-weight subjects both before and during puberty, and investigated their relationships with auxological parameters and indexes of insulin sensitivity.Design and MethodsDEHP metabolites (MEHP, 6-OH-MEHP, 5-oxo-MEHP, 5-OH-MEHP, and 5-CX-MEHP), were measured in urine by RP-HPLC-ESI-MS. Traditional statistical analysis and a data mining analysis using the Auto-CM analysis were able to offer an insight into the complex biological connections between the studied variables.ResultsThe data showed changes in DEHP metabolites in urine related with obesity, puberty, and presence of insulin resistance. Changes in urine metabolites were related with age, height and weight, waist circumference and waist to height ratio, thus to fat distribution. In addition, clear relationships in both obese and normal-weight subjects were detected among MEHP, its products of oxidation and measurements of insulin sensitivity.ConclusionIt remains to be elucidated whether exposure to phthalates per se is actually the risk factor or if the ability of the body to metabolize phthalates is actually the key point. Further studies that span from conception to elderly subjects besides further understanding of DEHP metabolism are warranted to clarify these aspects.

Skin permeation and metabolism of di(2-ethylhexyl) phthalate (DEHP)

Phthalates are suspected to be endocrine disruptors. Di(2-ethylhexyl) phthalate (DEHP) is assumed to have low dermal absorption; however, previous in vitro skin permeation studies have shown large permeation differences. Our aims were to determine DEHP permeation parameters and assess extent of skin DEHP metabolism among workers highly exposed to these lipophilic, low volatile substances.Surgically removed skin from patients undergoing abdominoplasty was immediately dermatomed (800μm) and mounted on flow-through diffusion cells (1.77cm2) operating at 32°C with cell culture media (aqueous solution) as the reservoir liquid. The cells were dosed either with neat DEHP or emulsified in aqueous solution (166μg/ml). Samples were analysed by HPLC–MS/MS.DEHP permeated human viable skin only as the metabolite MEHP (100%) after 8h of exposure. Human skin was able to further oxidize MEHP to 5-oxo-MEHP. Neat DEHP applied to the skin hardly permeated skin while the aqueous solution readily permeated skin measured in both cases as concentration of MEHP in the receptor liquid.DEHP pass through human skin, detected as MEHP only when emulsified in aqueous solution, and to a far lesser degree when applied neat to the skin. Using results from older in vitro skin permeation studies with non-viable skin may underestimate skin exposures. Our results are in overall agreement with newer phthalate skin permeation studies.

Species and inter-individual differences in metabolic capacity of di(2-ethylhexyl)phthalate (DEHP) between human and mouse livers

This study was conducted to assess inter-species and inter-individual differences in the metabolism of di(2-ethylhexyl)phthalate (DEHP) in humans and mice. The activities of four DEHP-metabolizing enzymes [lipase, UDP-glucuronocyltransferase (UGT), alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH)] were measured in the livers of 38 human subjects of various ages and in eight 129/Sv male mice. Microsomal lipase activity was significantly lower in humans than in mice. The V max/K m value in humans was one-seventh of that in mice, microsomal UGT activity in humans was a sixth of that in mice, and cytosolic ALDH activity for 2-ethylhexanal in humans was one-half of that in mice. In contrast, ADH activity for 2-ethylhexanol was twofold higher in humans than in mice. The total amount of DEHP urinary metabolites and the concentration of mono(2-ethylhexyl)phthalate (MEHP) were much higher in intact mice than in the U.S. general population based on data reported elsewhere, regardless of the similar estimated DEHP intake between these mice and the human reference population. However, mono(2-ethyl-5-oxo-hexyl)phthalate (5oxo-MEHP) and mono(2-ethyl-5-carboxypentyl)phthalate (5cx-MEPP) levels were higher in the latter than in the former. Of note, inter-subject variability in the activities of all enzymes measured was 10-26-fold. The inter-individual variation in the metabolism of DEHP in humans may be greater than the difference between mice and humans (inter-species variation), and both may affects the risk assessment of DEHP.

In vitro intestinal and hepatic metabolism of Di(2-ethylhexyl) phthalate (DEHP) in human and rat

Species and organ differences in the intrinsic clearance and the enzymes involved in the metabolism of DEHP were examined in subcellular fractions of the intestine and liver as well as by recombinant cytochrome P450 (CYP) isoforms of human and rat. Estimated clearance (CLint) of DEHP via esterase-mediated pathway in human intestine was 2.4-fold greater than that in human liver while its value in rat intestine was 1.7-fold less than that in rat liver. Ranks of CLint for CYP-mediated oxidation/dealkylation of MEHP were human liver>rat liver>human intestine>rat intestine. Estimates of CLint for the production of mono(2-ethyl-5-hydroxyhexyl) phthalate and mono(2-ethyl-5-oxohexyl) phthalate by human CYP2C9*1 were 4.2- and 2.6-fold greater than those by rat CYP2C6, respectively. Total CLint via hCYP2C9*3-mediated oxidation was 1.9- and 2.6-fold less than those by hCYP2C9*2 and 2C9*1, respectively. Estimated CLint for phthalic acid production by hCYP3A4 was 24.5μLnmolCYP−1min−1 while it was continuously produced by rCYP2C6 and 3A2 via passive mechanism. These species/organ differences in major metabolic pathway and CYP isoforms should be considered for appraisal of the potential adverse health effects of DEHP.

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 24months 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.

In vitro metabolism of di(2-ethylhexyl) phthalate (DEHP) by various tissues and cytochrome P450s of human and rat

In vitro metabolism of DEHP by subcellular fractions of human brain, intestine, kidney, liver, lung, skin, testis, rat liver and recombinant CYP isoforms of human and rat was investigated using LC–MS/MS. DEHP was rapidly hydrolyzed to mono(2-ethylhexyl) phthalate (MEHP) in 12 microsomal/cytosolic fractions of selected 7 human organs and rat liver but not in microsomal fractions of human brain and human female skin. MEHP was metabolized to CYP-mediated oxidative and dealkylated metabolites in human and rat liver and at a lower rate in human intestine. Measurable amounts of mono(2-ethyl-5-hydroxyhexyl) phthalate (5-OH MEHP), mono(2-ethyl-5-oxohexyl) phthalate (5-Oxo MEHP), mono(2-ethyl-5-carboxypentyl) phthalate (5-carboxy MEPP), mono(2-carboxymethyl-hexyl) phthalate (2-carboxy MMHP) and phthalic acid (PA) were formed by human liver fractions. Human CYP2C9∗1, CYP2C19 and rat CYP2C6 were the major CYP isoforms producing 5-OH MEHP and 5-Oxo MEHP metabolites; however, only human CYP2C9∗1 and 2C9∗2 produced 5-carboxy MEPP from MEHP. Additionally, human CYP3A4 and rat CYP3A2 were the primary enzymes for PA production via heteroatom dealkylation of MEHP. Percent total normalized rates (%TNR) by CYP2C9∗1 in human liver microsomes (HLM) were 94%, 98% and 100%, respectively, for 5-OH MEHP, 5-Oxo MEHP, 5-carboxy MEPP, and 76% for PA production by CYP3A4.

Urinary Metabolites of Di(2‐ethylhexyl) Phthalate Are Associated With Decreased Steroid Hormone Levels in Adult Men

Experimental animal studies have demonstrated that exposure to some phthalates may be associated with altered endocrine function and adverse effects on male reproductive development and function, but human studies are limited. In the present study, urine and serum samples were collected from 425 men recruited through a US infertility clinic. Urinary concentrations of mono(2-ethylhexyl) phthalate (MEHP), the hydrolytic metabolite of di(2-ethylhexyl) phthalate (DEHP), and other phthalate monoester metabolites were measured, along with serum levels of testosterone, estradiol, sex hormone-binding globulin (SHBG), follicle-stimulating hormone, luteinizing hormone, inhibin B, and prolactin. Two oxidized urinary metabolites of DEHP were also measured in urine from 221 of the men. In multiple regression models adjusted for potential confounders, MEHP was inversely associated with testosterone, estradiol, and free androgen index (FAI). An interquartile range increase in MEHP was associated with 3.7% (95% confidence interval [CI], -6.8% to -0.5%) and 6.8% (95% CI, -11.2% to -2.4%) declines in testosterone and estradiol, respectively, relative to the population median hormone levels. There was limited evidence for effect modification of the inverse association between MEHP and FAI by the proportion of DEHP metabolites in the urine measured as MEHP (MEHP%), which is considered a phenotypic marker of less efficient metabolism of DEHP to its oxidized metabolites. Finally, the ratio of testosterone to estradiol was positively associated with MEHP (P = .07) and MEHP% (P = .007), suggesting potential relationships with aromatase suppression. In conclusion, these results suggest that urinary metabolites of DEHP are inversely associated with circulating steroid hormone levels in adult men. However, additional research is needed to confirm these findings.