Toxicokinetics of mono-(2-ethylhexyl) phthalate with low-dose exposure applying fluorescence tracing technique.

Di(2-ethylhexyl) phthalate (DEHP) belongs to environmental endocrine disrupting chemicals (EEDCs) and can be rapidly hydrolyzed into the ultimate toxicant mono-2-ethylhexyl phthalate (MEHP). In this study, we used 5-aminofluorescein modified MEHP (MEHP-AF) as a fluorescence tracer to explore the toxicokinetics, including toxicokinetic parameters, absorption and transport across the intestinal mucosal barrier, distribution and pathological changes of organs. While the dose was as lower than 10 mg/kg by intragastric administration, the toxicokinetic parameters obtained by fluorescence microplate method were similar to those with the literatures by chromatography. MEHP-AF can be rapidly absorbed through the intestinal mucosal barrier in rats. In situ organ distribution in mice showed that MEHP-AF was mainly concentrated in the liver, kidney and testis. Our results suggested that the fluorescence tracing technique had the advantages with easy processing, less time-consuming, higher sensitivity for the quantitative determination, In addition, this technology also avoids the interference of exogenous or endogenous DEHP and MEHP in the experimental system. It also can be utilized to the visualization detection of MEHP in situ localization in the absorption organ and the toxic target organ. The results show that this may be a more feasible MEHP toxicological research method.

The oral bioavailability of di-2-ethylhexyl phthalate (DEHP), di-isononyl phthalate (DiNP) and di-(isononyl)-cyclohexane-1,2-dicarboxylate (DINCH®) in house dust.

Phthalates and other plasticizers are detected in high amounts in the indoor environment and therefore house dust can be an exposure source. Especially children have a relatively high unintended uptake of house dust, thus a higher exposure to plasticizers compared to adults may be possible. As accurate as possible exposure assessment data of the oral bioavailability of these compounds are necessary, however only one in vivo study with piglets is available so far. The aim of this study was to examine the oral bioavailability of phthalates and DINCH® in humans, which occur in typical house dust samples. We focused on the high molecular weight phthalates DEHP and DINP and their substitute DINCH®. Eleven volunteers ingested 6 g of house dust sieved to 2 mm. The urine was collected over a period of 36 h. The excreted plasticizers metabolites were quantified by an LC-MS/MS method. The mean recovery of urine metabolites was 51 % ± 20 % for DEHP, 26 % ± 13 % for DINP and 19 % ± 6% for DINCH® based on the parent compounds administered as dust samples. The metabolites of DEHP, DINP and DINCH® reached their maximum concentration after 2-19 hours post dose in urine. The bioavailability of DEHP was in agreement among the different dust samples. For DEHP, we were able to confirm previous findings from the oral bioavailability study with piglets and we could not observe a significant difference between the dust particle size (65 µm vs 2 mm) and the bioavailability. Considering the observed bioavailability, an estimated dust intake of 50 mg/d for toddlers can substantially contribute to the total plasticizer exposure.

Food emulsifier polysorbate 80 promotes the intestinal absorption of mono-2-ethylhexyl phthalate by disturbing intestinal barrier.

Di-2-ethylhexyl phosphate (DEHP) and its main toxic metabolite mono-2-ethylhexyl phthalate (MEHP) are the typical endocrine disrupting chemicals (EDCs) and widely affect human health. Our previous research reported that synthetic nonionic dietary emulsifier polysorbate 80 (P80, E433) had the promotional effect on the oral absorption of DEHP in rats. The aim of this study was to explore its mechanism of promoting oral absorption, focusing on the mucus barrier and mucosal barrier of the small intestine. A small molecule fluorescent probe 5-aminofluorescein-MEHP (MEHP-AF) was used as a tracker of MEHP in vivo and in vitro. First of all, we verified that P80 promoted the bioavailability of MEHP-AF in the long-term and low-dose exposure of MEHP-AF with P80 as a result of increasing the intestinal absorption of MEHP-AF. Afterwards, experimental results from Western blot, qPCR, immunohistochemistry, and immunofluorescence showed that P80 decreased the expression of proteins (mucus protein mucin-2, tight junction proteins claudin-1 and occludin) related to mucus barrier and mucosal barrier in the intestine, changed the integrity of intestinal epithelial cell, and increased the permeability of intestinal epithelial mucosa. These results indicated that P80 promoted the oral absorption of MEHP-AF by altering the intestinal mucus barrier and mucosal barrier. These findings are of great importance for assessing the safety risks of some food emulsifiers and clarifying the absorption mechanism of chemical pollutants in food, especially for EDCs.

Skin transferability of phthalic acid ester plasticizers and other plasticizers using model polyvinyl chloride sheets.

The transferability of phthalic acid esters (PAEs) and other plasticizers, from model polyvinyl chloride (PVC) sheets to the skin of 11 subjects was assessed by measuring the amount of substance transferred using PVC sheets containing PAEs and alternative plasticizers of different types and contents. For all subjects, the transferred amount, from sheets containing 28 wt% PAE or from mixed sheets containing 14 wt% each of di (2-ethylhexyl) phthalate (DEHP) and other PAE, was greater than that from sheets containing 15 wt% each of PAE or alternative plasticizer only. A comparison of the transferability of five types of PAE showed that transfer tended to occur more readily as the n-octanol-water partition coefficient increased, suggesting that PAE hydrophobicity affected its transferability. The transferability of the alternative plasticizers di(2-ethylhexyl) terephthalate and 1,2-cyclohexane dicarboxylic acid diisononyl ester showed a similar trend; however, the transferred amount tended to be higher from model PVC sheets containing 28 wt% PAE or mixed with DEHP. The transferability of PAEs and alternative plasticizers was higher for certain subjects, suggesting individual differences in the transferability of chemicals to the subject's skin surface and is the presence of a group of people comparatively more susceptible to such transfer.

Comparison of aggregated exposure to di(2-ethylhexyl) phthalate from diet and personal care products with urinary concentrations of metabolites using a PBPK model - Results from the Norwegian biomonitoring study in EuroMix.

Phthalates are widely used as plasticisers in flexible plastics and containers for food and personal care products (PCPs) and contaminates foods and PCPs. A human biomonitoring (BM) study was performed to study exposure of chemicals from foods and PCPs. For two 24-h periods, adult volunteers (n = 144) in Norway kept diaries on food eaten and usage of PCPs, and collected 24-h urine. Aggregated exposure to di(2-ethylhexyl) phthalate (DEHP) from dietary and PCPs was estimated by Monte-Carlo simulation using Oracle Crystal Ball©. Simulated urinary concentrations using physiologically based pharmacokinetic (PBPK) models were compared with measured urinary metabolites of DEHP, 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 (MECCP). DEHP exposure from food are approximately 10 times higher than exposure than from PCPs. The main contributors to dietary exposure are dairy, grain, fruits and vegetables, meat and fish. Body lotion contribute most to the exposure of DEHP from PCPs. Forward-dosimetry gives good convergence with 24-h urinary concentrations of simulated and measured BM data. The measured concentration of the MECCP metabolite correlated well with simulated high exposure, while the measured concentrations of MEHP, MEHHP and MEOHP partly overlapped with both simulated low, medium and high metabolite exposure.

Application of a combined aggregate exposure pathway and adverse outcome pathway (AEP-AOP) approach to inform a cumulative risk assessment: A case study with phthalates.

Advancements in measurement and modeling capabilities are providing unprecedented access to estimates of chemical exposure and bioactivity. With this influx of new data, there is a need for frameworks that help organize and disseminate information on chemical hazard and exposure in a manner that is accessible and transparent. A case study approach was used to demonstrate integration of the Adverse Outcome Pathway (AOP) and Aggregate Exposure Pathway (AEP) frameworks to support cumulative risk assessment of co-exposure to two phthalate esters that are ubiquitous in the environment and that are associated with disruption of male sexual development in the rat: di(2-ethylhexyl) phthalate (DEHP) and di-n-butyl phthalate (DnBP). A putative AOP was developed to guide selection of an in vitro assay for derivation of bioactivity values for DEHP and DnBP and their metabolites. AEPs for DEHP and DnBP were used to extract key exposure data as inputs for a physiologically based pharmacokinetic (PBPK) model to predict internal metabolite concentrations. These metabolite concentrations were then combined using in vitro-based relative potency factors for comparison with an internal dose metric, resulting in an estimated margin of safety of ~13,000. This case study provides an adaptable workflow for integrating exposure and toxicity data by coupling AEP and AOP frameworks and using in vitro and in silico methodologies for cumulative risk assessment.

[Experimental Study on Migration Parameters of DEHP in PVC Infusion].

The work explored the DEHP migration parameters in PVC infusion in clinic,based on the previous research on the test model of DEHP migrated from PVC infusion,to assess the safety of PVC infusion.The leaching solution samples in different conditions were evaluated by analysis of the DEHP in leaching solution using GC-MS under simulated clinical transfusion way.The release behavior of DEHP was significantly affected by the storage time,storage temperature,surrounding temperature,dripping speed,sterilization process,volume of the leaching solution,and the property of the leaching solution.

Recovery From a Myocardial Infarction Is Impaired in Male C57bl/6 N Mice Acutely Exposed to the Bisphenols and Phthalates That Escape From Medical Devices Used in Cardiac Surgery.

Bisphenols and phthalates leach from medical devices, and this exposure is likely to increase in postcardiac surgery patients. Previous studies suggest that such chemical exposure may impact recovery and wound healing, yet the direct effects of bisphenols and phthalates are unknown in this context. To study the direct effect of clinically based chemical exposures, we measured the metabolites representative of 6 bisphenols and 10 phthalates in men before and after cardiac surgery and then replicated this exposure in a mouse model of cardiac surgery and assessed survival, cardiac function and inflammation. Bisphenol A (BPA), di-ethyl hexyl phthalate (DEHP), butylbenzyl phthalate, di-isodecyl phthalate, and di-n-butyl phthalate metabolites were increased after surgery. DEHP exposure predominated, was positively correlated with duration on the cardiopulmonary bypass machine and exceeded its tolerable daily intake limit by 37-fold. In vivo, C57bl/6 N male mice treated with BPA+phthalates during recovery from surgery-induced myocardial infarction had reduced survival, greater cardiac dilation, reduced cardiac function and increased infiltration of neutrophils, monocytes and macrophages suggesting impaired recovery. Of interest, genetic ablation or estrogen receptor beta (ERß) antagonism did not improve recovery and replacement of DEHP with tri-octyl trimellitate or removal of BPA from the mixture did not ameliorate these effects. To examine the direct effects on inflammation, treatment of human THP-1 macrophages with BPA and phthalates induced a dysfunctional proinflammatory macrophage phenotype with increased expression of M1-type macrophage polarization markers and MMP9 secretion, yet reduced phagocytic activity. These results suggest that chemicals escape from medical devices and may impair patient recovery.

Human metabolism and kinetics of tri-(2-ethylhexyl) trimellitate (TEHTM) after oral administration.

Tri-(2-ethylhexyl) trimellitate (TEHTM) is a plasticizer for PVC material and is used for medical devices as an alternative to di-(2-ethylhexyl) phthalate. As plasticizers are known to migrate easily into contact liquids, exposure of patients to TEHTM is highly probable. In the present study, human metabolism pathways of TEHTM and its elimination kinetics were investigated. For that purpose, four healthy volunteers were orally exposed to a single dose of TEHTM. TEHTM and its postulated primary metabolites were investigated in blood samples (up to 48 h after exposure), and in urine samples (collected until 72 h after exposure) using liquid chromatography tandem mass spectrometry (LC-MS/MS). TEHTM was found to be regioselectively hydrolyzed to its diesters di-2-(ethylhexyl) trimellitates (1,2-DEHTM, 2,4-DEHTM) with maximum blood concentrations at 3-h post-exposure, and to its monoester isomers mono-2-(ethylhexyl) trimellitates (1-MEHTM, 2-MEHTM) with peak blood concentrations 5-h post-exposure. For the elimination of investigated urinary metabolites, biphasic elimination kinetics was observed. The most dominant urinary biomarker was found to be 2-MEHTM (2-mono-(2-ethylhexyl) trimellitate), followed by several specific secondary metabolites. All in all, approximately 5.8% of the orally administered dose was recovered in urine over a period of 72 h, indicating a comparatively low resorption rate of TEHTM in humans in combination with an apparently rather slow metabolism and excretion rate. In fact, TEHTM and selected metabolites were still detectable in blood and urine 48-h and 72-h post-exposure, respectively. This study is the first to elucidate TEHTM metabolism pathways in humans and to identify metabolites of TEHTM in blood and urine by usage of especially designed human biomonitoring methods. Powerful tools for exposure monitoring and risk assessment of TEHTM are therewith available for future research.

Development of a human physiologically based pharmacokinetic (PBPK) model for phthalate (DEHP) and its metabolites: A bottom up modeling approach.

DEHP exposure to human comes from different sources such as food, diet, cosmetics, toys, medical products, and food wraps. Recently, DEHP was categorized as non-persistent endocrine disrupting compounds (EDCs) by the world health organization (WHO). Rat experimental studies showed that phthalate and its metabolite(s) can cause hepatic, developmental and reproductive toxicity. In human, DEHP rapidly metabolizes into a toxic metabolite MEHP. This MEHP further metabolizes into the different chemical forms of 5OH-MEHP, 5oxo-MEHP, 5cx-MEPP and phthalic acid. A simple DEHP pharmacokinetics model has been developed, but with a limited number of metabolites. A chemical like DEHP which extensively metabolised indicate the need of a detail metabolic kinetics study. A physiological based pharmacokinetics (PBPK) model of the DEHP considering all the major metabolites in human, has not been developed yet. The objective of this study is to develop a detailed human PBPK model for the DEHP and its major metabolites by using a bottom-up modelling approach with the integration of a in vitro metabolic data. This approach uses an in-vitro-in-vivo extrapolation (IVIVE) and a quantitative structure-activity relationship (QSAR) method for the parameterization of the model. Monte Carlo simulations were performed to estimate the impact of parametric uncertainty onto the model predictions. First, the model was calibrated using the control human kinetic study that represents the time course of DEHP metabolites concentration in both the blood and the urine. Then, the model was evaluated against the published independent data on different dosing scenarios. The results of model predictions for the DEHP metabolites in both the blood and the urine were well within the range of experimentally observed data. The model also captured the similar trend of time course profile to the observed data, shows model good predictability power. The current developed PBPK model can futher be used for the prediction of the time course of chemical concentrations for the different exposure scenarios not only in the blood and the urine but also in the other compartments. Moreover, this model can also be used to explore different biomonitoring studies for the human health risk assessment and might be useful for integrative toxicological study in improving exposure-target tissue dose-response relationship.

Fate of four phthalate plasticizers under various wastewater treatment processes.

The fate of four phthalate plasticizers during wastewater treatment processes at six different wastewater treatment plants (WWTPs) was investigated. Concentrations of benzyl butyl phthalate (BBP), di(2-ethylhexyl) phthalate (DEHP), diisononyl phthalate (DiNP), and diisodecyl phthalate (DiDP) were determined prior to either aerobic or anaerobic (conventional and advanced) treatment, after treatment, and in final, dewatered solids. Despite their elevated use worldwide, the fate of DiNP and DiDP during wastewater treatment have not been well characterized. DEHP was readily degraded during aerobic treatments while anaerobic digestion resulted in either no significant change in concentrations or an increase in concentration, in the case of more advanced anaerobic processes (thermal hydrolysis pretreatment and a two-phase acid/gas process). Impacts of the various treatment systems on DiNP, DiDP, and BBP concentrations were more varied - anaerobic digestion led to significant decreases, increases, or no significant change for these compounds, depending on the treatment facility, while aerobic treatment was generally effective at degrading the compounds. Additionally, thermal hydrolysis pretreatment of sludge prior to anaerobic digestion resulted in increases in DiNP, DiDP, and BBP concentrations. The predicted environmental concentrations for all four compounds in soils after a single biosolids application were calculated and the risk quotients for DEHP in soils were determined. The estimated toxicity risk for DEHP in soils treated with a single application of sludge from any of the six studied WWTPs is lower than the level of concern for acute and chronic risk, as defined by the US EPA.

Excretion of Di-2-ethylhexyl phthalate (DEHP) metabolites in urine is related to body mass index because of higher energy intake in the overweight and obese.

A higher body mass index (BMI) has been positively associated with the rate of excretion of di-2-ethylhexyl phthalate (DEHP) metabolites in urine in data from the National Health and Nutrition Examination Survey (NHANES), suggesting an association between DEHP exposure and BMI. The association, however, may be due to the association between body mass maintenance and higher energy intake, with higher energy intake being accompanied by a higher intake of DEHP. To examine this hypothesis, we ran a Monte Carlo simulation with a DEHP physiologically-based pharmacokinetic (PBPK) model for adult humans. A realistic exposure sub-model was used, which included the relation of body weight to energy intake and of energy intake to DEHP intake. The model simulation output, when compared with urinary metabolite data from NHANES, supported good model validity. The distribution of BMI in the simulated population closely resembled that in the NHANES population. This indicated that the simulated subjects and DEHP exposure model were closely aligned with the NHANES population of interest. In the simulated population, the ordinary least squares regression coefficient for log(BMI) as a function of log(DEHP nmol/min) was 0.048 (SE 0.001), as compared with the reported value of 0.019 (SE 0.005). In other words, given our model structure, the higher energy intake in the overweight and obese, and the concomitant higher DEHP exposure, describes the reported relationship between BMI and DEHP.

Prenatal exposure estimation of BPA and DEHP using integrated external and internal dosimetry: A case study.

Prenatal exposure to Endocrine disruptors (EDs), such as Bisphenol A (BPA) and di (2-ethylhexyl) phthalate (DEHP), has been associated with obesity and diabetes diseases in childhood, as well as reproductive, behavioral and neurodevelopment problems. The aim of this study was to estimate the prenatal exposure to BPA and DEHP through food consumption for pregnant women living in Tarragona County (Spain). Probabilistic calculations of prenatal exposure were estimated by integrated external and internal dosimetry modelling, physiologically based pharmacokinetic (PBPK) model, using a Monte-Carlo simulation. Physical characteristic data from the cohort, along with food intake information from the questionnaires (concentrations of BPA and DEHP in different food categories and the range of the different food ratios), were used to estimate the value of the total dietary intake for the Tarragona pregnancy cohort. The major contributors to the total dietary intake of BPA were canned fruits and vegetables, followed by canned meat and meat products. In turn, milk and dairy products, followed by ready to eat food (including canned dinners), were the most important contributors to the total dietary intake of DEHP. Despite the dietary variations among the participants, the intakes of both chemicals were considerably lower than their respective current tolerable daily intake (TDI) values established by the European Food Safety Authority (EFSA). Internal dosimetry estimates suggest that the plasma concentrations of free BPA and the most important DEHP metabolite, mono (2-ethylhexyl) phthalate (MEHP), in pregnant women were characterized by transient peaks (associated with meals) and short half-lives (< 2h). In contrast, fetal exposure was characterized by a low and sustained basal BPA and MEHP concentration due to a lack of metabolic activity in the fetus. Therefore, EDs may have a greater effect on developing organs in young children or in the unborn child.

Safety evaluation of dermal exposure to phthalates: Metabolism-dependent percutaneous absorption.

Phthalates, known as reproductive toxicants and endocrine disruptors, are widely used as plasticizers in polyvinyl chloride products. The present study was conducted for risk identification of dermal exposure to phthalates. When dibutyl phthalate was applied to the skin of hairless rats and humans, only monobutyl phthalate appeared through the skin, and the permeability of the skin was higher than that after the application of the monoester directly. The inhibition of skin esterases made the skin impermeable to the metabolite following dermal exposure to dibutyl ester, whereas removal of the stratum corneum from the skin did not change the skin permeation behavior. Similar phenomena were observed for benzyl butyl phthalate. The skin permeability of monobenzyl phthalate was higher than that of monobutyl phthalate in humans, although the reverse was observed in rats. Species difference in skin permeation profile corresponded to the esterase activity of the skin homogenate. Di(2-ethylhexyl) phthalate, which was not metabolized by esterases in the skin, was not transported across the skin. These results suggest that highly lipophilic phthalates may be transported easily across the stratum corneum lipids. The water-rich viable layer may become permeable to these phthalates by their metabolism into monoesters, which are relatively hydrophilic. Skin metabolism is essential to the percutaneous absorption of phthalates. Because esterase activity has large inter-individual differences, further study will be needed for individual risk identification of dermal exposure to phthalates.

Glucuronidation of mono(2-ethylhexyl) phthalate in humans: roles of hepatic and intestinal UDP-glucuronosyltransferases.

Mono(2-ethylhexyl) phthalate (MEHP) is an active metabolite of di(2-ethylhexyl) phthalate (DEHP), which is an endocrine-disrupting chemical. In the present study, MEHP glucuronidation in humans was studied using recombinant UDP-glucuronosyltransferases (UGTs) and microsomes of the liver and intestine. Among the recombinant UGTs examined, UGT1A3, UGT1A7, UGT1A8, UGT1A9, UGT1A10, UGT2B4, and UGT2B7 glucuronidated MEHP. The kinetics of MEHP glucuronidation by UGT1A3, UGT1A7, UGT1A8, UGT1A10, UGT2B4, and UGT2B7 followed the Michaelis-Menten model, whereas that by UGT1A9 fit the negative allosteric model. CLint values were in the order of UGT1A9 > UGT2B7 > UGT1A7 > UGT1A8 ≥ UGT1A10 > UGT1A3 > UGT2B4. The kinetics of MEHP glucuronidation by liver microsomes followed the Michaelis-Menten model. Diclofenac (20 µM) and raloxifene (20 µM) decreased CLint values to 43 and 36 % that of native microsomes, respectively. The kinetics of MEHP glucuronidation by intestine microsomes fit the biphasic model. Diclofenac (150 and 450 µM) decreased CLint values to 32 and 13 % that of native microsomes for the high-affinity phase, and to 28 and 21 % for the low-affinity phase, respectively. Raloxifene (2.5 and 7.0 µM) decreased CLint values to 35 and 4.1 % that of native microsomes for the high-affinity phase, and to 48 and 53 % for the low-affinity phase, respectively. These results suggest that MEHP glucuronidation in humans is catalyzed by UGT1A3, UGT1A9, UGT2B4, and/or UGT2B7 in the liver, and by UGT1A7, UGT1A8, UGT1A9, UGT1A10, and/or UGT2B7 in the intestine, and also that these UGT isoforms play important and characteristic roles in the detoxification of DEHP.

Kinetics of the phthalate metabolites mono-2-ethylhexyl phthalate (MEHP) and mono-n-butyl phthalate (MnBP) in male subjects after a single oral dose.

Humans have been exposed to dialkyl ortho-phthalates for decades. Due to degradation the phthalate monoesters, responsible for the toxic effects, are additionally found in environmental media as well as food samples. Nevertheless, the toxicokinetic properties of the monoesters are not known. Therefore, metabolism of the phthalate monoesters mono-2-ethylhexyl phthalate (MEHP) and mono-n-butyl phthalate (MnBP) was studied in four male volunteers (23-58 years of age) after ingestion of a single dose of 50µg/kg bw D4-MEHP or 10µg/kg bw D4-MnBP. The main metabolites in urine were determined up to 46h after administration. In the MEHP-study, more than 90% of each metabolite appeared in the urine within the first 22h, and the average excreted amount of D4-MEHP and its four secondary metabolites was 62% of the administered dose. The highest value of 15% was observed for mono-2-ethyl-5-carboxy-pentyl phthalate (D4-5cx-MEPP). The mean elimination half-life of D4-MEHP was estimated to be 3.5±1.4h. In the MnBP-study, the total recovered values of D4-MnBP and its secondary metabolites ranged from 52% to 130%. The monoester itself, with a half-life of 1.9±0.5h, accounted for the majority of the ingested dose (92%), while the secondary metabolites D4-mono-3-hydroxy-n-butyl phthalate (D4-3OH-MnBP) and D4-3-carboxy-mono-propyl phthalate (D4-3cx-MPP) represented only 7.1% and 1.0% of the ingested dose, respectively. Overall, this study determined that the kinetics of the phthalate monoesters MEHP and MnBP after oral dosage are comparable to the properties of their diesters.

Mono(2-ethylhexyl)phthalate accumulation disturbs energy metabolism of fat cells.

Phthalates are lipophilic and tend to accumulate in adipose tissue, an important regulator of energy balance and glucose homeostasis. The study aimed to determine whether cellular phthalate accumulation influenced fat cell energy metabolism. Following a 3-day treatment with adipogenesis-inducing medium and a 2-day treatment with adipogenesis-maintaining medium, 3T3-L1 cells differentiated into adipocytes in the presence of a phthalate at a clinically relevant concentration (30-300 µM) for another 6 days. Two phthalates, di(2-ethylhexyl)phthalate and di-n-butylphthalate, and their metabolites, mono(2-ethylhexyl)phthalate (MEHP) and mono-n-butylphthalate, were used here. The phthalate treatments caused no marked effect on cytotoxicity and adipogenesis. Only the MEHP-treated adipocytes were found having smaller lipid droplets; MEHP accumulated in cells in a dose- and time-dependent manner. The MEHP-treated adipocytes exhibited significant increases in lipolysis and glucose uptake; quantitative real-time polymerase chain reaction (qPCR) analysis revealed correlated changes in expression of marker genes involved in adipogenesis, lipid metabolism, and glucose uptake. Analysis of oxygen consumption rate (a mitochondrial respiration indicator) and extracellular acidification rate (a glycolysis indicator) indicated a higher energy metabolism in the adipocytes. qPCR analysis of critical genes involved in mitochondrial biogenesis and/or energy metabolism showed that expression of peroxisome proliferator-activated receptor γ coactivator-1α, sirtuin 3, and protein kinase A were significantly enhanced in the MEHP-treated adipocytes. In vitro evidence of MEHP impacts on lipolysis, glucose uptake/glycolysis, and mitochondrial respiration/biogenesis demonstrates that MEHP accumulation disturbs energy metabolism of fat cells.

Uptake and Metabolism of Phthalate Esters by Edible Plants.

Phthalate esters (PAEs) are large-volume chemicals and are found ubiquitously in soil as a result of widespread plasticulture and waste disposal. Food plants such as vegetables may take up and accumulate PAEs from soil, potentially imposing human health risks through dietary intake. In this study, we carried out a cultivation study using lettuce, strawberry, and carrot plants to determine the potential of plant uptake, translocation, and metabolism of di-n-butyl phthalate (DnBP) and di(2-ethylhexyl) phthalate (DEHP) and their primary metabolites mono-n-butyl phthalate (MnBP) and mono(2-ethylhexyl) phthalate (MEHP). All four compounds were detected in the plant tissues, with the bioconcentration factors (BCFs) ranging from 0.16 ± 0.01 to 4.78 ± 0.59. However, the test compounds were poorly translocated from roots to leaves, with a translocation factor below 1. Further, PAEs were readily transformed to their monoesters following uptake. Incubation of PAEs and monoalkyl phthalate esters (MPEs) in carrot cell culture showed that DnBP was hydrolyzed more rapidly than DEHP, while the monoesters were transformed more quickly than their parent precursors. Given the extensive metabolism of PAEs to monoesters in both whole plants and plant cells, metabolism intermediates such as MPEs should be considered when assessing human exposure via dietary intake of food produced from PAE-contaminated soils.

Human biofluid concentrations of mono(2-ethylhexyl)phthalate extrapolated from pharmacokinetics in chimeric mice with humanized liver administered with di(2-ethylhexyl)phthalate and physiologically based pharmacokinetic modeling.

Di(2-ethylhexyl)phthalate (DEHP) is a reproductive toxicant in male rodents. The aim of the current study was to extrapolate the pharmacokinetics and toxicokinetics of mono(2-ethylhexyl)phthalate (MEHP, a primary metabolite of DEHP) in humans by using data from oral administration of DEHP to chimeric mice transplanted with human hepatocytes. MEHP and its glucuronide were detected in plasma from control mice and chimeric mice after single oral doses of 250mg DEHP/kg body weight. Biphasic plasma concentration-time curves of MEHP and its glucuronide were seen only in control mice. MEHP and its glucuronide were extensively excreted in urine within 24h in mice with humanized liver. In contrast, fecal excretion levels of MEHP glucuronide were high in control mice compared with those with humanized liver. Adjusted animal biomonitoring equivalents from chimeric mice studies were scaled to human biomonitoring equivalents using known species allometric scaling factors and in vitro metabolic clearance data with a simple physiologically based pharmacokinetic (PBPK) model. Estimated urine MEHP concentrations in humans were consistent with reported concentrations. This research illustrates how chimeric mice transplanted with human hepatocytes in combination with a simple PBPK model can assist evaluations of pharmacokinetics or toxicokinetics of the primary or secondary metabolites of DEHP.

Effects of Potamogeton crispus L.-bacteria interactions on the removal of phthalate acid esters from surface water.

To investigate the mechanism of submerged macrophyte-bacteria interactions on the removal of phthalic acid esters from surface water, experiments with and without Potamogeton crispus L. were performed. A two-compartment (i.e., water and plant) kinetic model was developed. The model adequately described the variation of dibutyl phthalate (DBP) and di-2-ethylhexyl phthalate (DEHP) in the plant-water system by providing the first-order rate constants of plant uptake (k1) and release (k2), microbial degradation in water (k3) and plant degradation (k4). During 10-d incubation, the presence of P. crispus enhanced the removal of DBP and DEHP from water by 6.3% and 22.4%. Compared with the experiment without P. crispus, biodegradation of DBP in water with P. crispus decreased by 8.3% because of plant uptake even though k3 increased by 30%. 21.4% of DBP transferred from water to plants, of which only small amount (5.1%) retained in the plant and the rest (94.9%) was degraded. Different from DBP, biodegradation of DEHP in water with P. crispus was a slightly higher than that without P. crispus. 25.5% of DEHP transferred from water to plants, of which a large portion (73.3%) retained in the plant and the rest (26.7%) was degraded. This finding reveals that the enhancement of DBP removal from surface water is mainly related to faster degradation in the plant, whereas it is mainly related to higher plant accumulation for DEHP.