Di(2-ethylhexyl) Terephthalate Research Articles

Exposure to organophosphate esters, phthalates, and alternative plasticizers in association with uterine fibroids

Exposure to endocrine disrupting chemicals is suggested to be responsible for the development or progression of uterine fibroids. However, little is known about risks related to emerging chemicals, such as organophosphate esters (OPEs) and alternative plasticizers (APs). A case-control study was conducted to investigate whether exposures to OPEs, APs, and phthalates, were associated with uterine fibroids in women of reproductive age. For this purpose, the cases (n = 32) and the matching controls (n = 79) were chosen based on the results of gynecologic ultrasonography among premenopausal adult women in Korea and measured for metabolites of several OPEs, APs, and major phthalates. Logistic regression models were employed to assess the associations between chemical exposure and disease status. Factor analysis was conducted for multiple chemical exposure assessments as a secondary analysis. Among OPE metabolites, diphenyl phosphate (DPHP), 2-ethylhexyl phenyl phosphate (EHPHP), and 1-hydroxy-2-propyl bis(1-chloro-2-propyl) phosphate (BCIPHIPP) were detected in >80% of the subjects. Among APs, metabolites of di-isononyl phthalate (DINP) and di(2-propylheptyl) phthalate (DPrHpP) were detected in >75% of the urine samples. The odds ratios (ORs) of uterine fibroids were significantly higher among the women with higher exposures to tris(1,3-dichloro-2-propyl) phosphate (TDCIPP) and tris(2-butoxyethyl) phosphate (TBOEP), di(2-ethylhexyl) terephthalate (DEHTP), DPrHpP, and di-(iso-nonyl)-cyclohexane-1,2-dicarboxylate (DINCH). In addition, urinary concentrations of mono(2-ethyl-5-oxohexyl) phthalate (MEOHP), a sum of five di(2-ethylhexyl) phthalate metabolites (∑5DEHP), and mono(4-methyl-7-hydroxyoctyl) phthalate (OH-MINP) were significantly higher in the cases. In factor analysis, a factor heavily loaded with DPrHpP and DEHP was significantly associated with uterine fibroids, supporting the observation from the single chemical regression model. We found for the first time that several metabolites of OPEs and APs are associated with increased risks of uterine fibroids among pre-menopausal women. Further epidemiological and mechanistic studies are warranted to validate the associations observed in the present study.

Development and validation of a liquid chromatography/tandem mass spectrometry method to quantify metabolites of phthalates, including di-2-ethylhexyl terephthalate (DEHTP) and bisphenol A, in human urine.

Several phthalates and bisphenol A are endocrine-disrupting chemicals (EDCs). Recently, their use has been partially restricted and less toxic compounds, such as di-2-ethylhexyl terephthalate (DEHTP), have been placed on the market. The aim of this work was to develop and validate a method for the simultaneous quantitation of bisphenol A and urinary metabolites of phthalates, including DEHTP. An isotopic dilution high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS) method for the determination of bisphenol A (BPA), monobenzyl phthalate (MBzP), mono-2-ethyl-5-carboxypentyl phthalate (MECPP), mono-2-ethyl-5-carboxypentyl terephthalate (MECPTP), mono-2-ethyl-5-hydroxyhexyl terephthalate (MEHHTP), monoethyl phthalate (MEP), and mono-n/i-butyl phthalates (MnBP/MiBP) in human urine was developed. A complete validation was carried out and the method was applied to 36 non-occupationally exposed adults. Limits of quantitation ranged from 0.02 (MECPP) to 1 μg/L (MnBP and MiBP). Relative standard deviations below 10% indicated a suitable precision; accuracy, evaluated using a standard reference material, ranged from 74.3% to 117.5%; isotopically labelled internal standards were suitable for correcting the matrix effect. The accuracy was confirmed by the successful participation in an external verification exercise. However, for terephthalates, the validation was incomplete due to the lack of reference materials and external verification. Levels of the investigated chemicals in subjects were in line with those previously reported. An LC/MS/MS assay for the simultaneous measurement of BPA and phthalate metabolites in human urine was developed and validated; it is useful to investigate exposure in epidemiological studies involving the general population.

Phthalates and non-phthalate plasticizers in sediment from Korean coastal waters: Occurrence, spatial distribution, and ecological risks

Due to regulations on phthalates, non-phthalate plasticizers (NPPs) are now used as an alternative. Limited studies have been conducted on the occurrence and distribution of NPPs. In this study, sediment samples were collected from 50 locations along the Korean coast to assess the occurrence, distribution, sources, and ecological risks of phthalates and NPPs. Phthalates and NPPs were detected in all sediments, indicating ubiquitous contamination of the coastal environment. Di(2-ethylhexyl)phthalate (DEHP) and di(2-ethylhexyl)terephthalate (DEHT) were dominant, suggesting that DEHT could be an emerging contaminant of concern. The highest concentrations of phthalates and NPPs were found in sediment samples from harbors, implying they are contaminated hotspots. Sedimentary organic carbon was a major factor governing the distribution of phthalates and NPPs. Significant correlations were observed among phthalates and NPPs, suggesting similar sources and geochemical behavior. DEHP concentration exceeded threshold values, indicating potential health risks to benthic organisms in sediments.

Plasticizer migration from children's toys, child care articles, art materials, and school supplies

Dialkyl phthalates, including diisononyl phthalate (DINP), have been used as plasticizers in children's products made from polyvinyl chloride (PVC), such as teethers and toys. Children may be exposed to phthalates when handling or mouthing PVC products because plasticizers are not covalently bound. The Consumer Product Safety Improvement Act of 2008 prohibited certain phthalates from use in child care articles and children's toys. Thus, manufacturers have changed to other plasticizers or non-PVC plastics and there is interest in evaluating the potential health risks of alternative plasticizers. In 2008, CPSC staff purchased 63 children's products comprising 129 individual pieces (articles). Plastics identified FTIR included PVC, polypropylene, polyethylene, and acrylonitrile butadiene styrene. Plasticizers identified by in the 38 PVC articles included acetyltributyl citrate (ATBC) (20); di (2-ethylhexyl) terephthalate (DEHT) (14); 1,2-cyclohexanedicarboxylic acid diisononyl ester (DINX) (13); 2,2,4-trimethyl-1,3-pentanediol diisobutyrate (TPIB) (9); di (2 ethyhexyl) phthalate (DEHP) (1); and DINP (1). Half of the tested articles contained multiple plasticizers. CPSC measured migration rates using the Joint Research Centre method. Migration rates correlated roughly with plasticizer concentration and inversely with the molecular mass of the plasticizer. We then combined the migration rates with data on mouthing duration to estimate children's exposure to plasticizers in toys and child care articles, and estimated margins of exposure. All margins of exposure were >1,000, suggesting a low risk potential. However, the plasticizers in this study have multiple uses. Exposure from other sources and routes of exposure will be considered in future work.

Exposure of Nail Salon Workers to Phthalates, Di(2-ethylhexyl) Terephthalate, and Organophosphate Esters: A Pilot Study.

Relatively little is known about the exposure of nail technicians to semivolatile organic compounds (SVOCs) in nail salons. We collected preshift and postshift urine samples and silicone wrist bands (SWBs) worn on lapels and wrists from 10 female nail technicians in the Boston area in 2016-17. We analyzed samples for phthalates, phthalate alternatives, and organophosphate esters (OPEs) or their metabolites. Postshift urine concentrations were generally higher than preshift concentrations for SVOC metabolites; the greatest change was for a metabolite of the phthalate alternative di(2-ethylhexyl) terephthalate (DEHTP): mono(2-ethyl-5-carboxypentyl) terephthalate (MECPTP) more than tripled from 11.7 to 36.6 μg/g creatinine. DEHTP biomarkers were higher in our study participants' postshift urine compared to 2015-2016 National Health and Nutrition Examination Survey females. Urinary MECPTP and another DEHTP metabolite were moderately correlated (r = 0.37-0.60) with DEHTP on the SWBs, suggesting occupation as a source of exposure. Our results suggest that nail technicians are occupationally exposed to certain phthalates, phthalate alternatives, and OPEs, with metabolites of DEHTP showing the largest increase across a work day. The detection of several of these SVOCs on SWBs suggests that they can be used as a tool for examining potential occupational exposures to SVOCs among nail salon workers.

Urinary levels of phthalates and DINCH metabolites in Korean and Thai pregnant women across three trimesters

Phthalates are anti-androgenic chemicals and may cause long-lasting adverse effects on growing fetuses. Understanding their exposure profile during pregnancy, therefore, is of public health importance. Because both behavioral and physiological changes of pregnant women are expected to be substantial, the amount of phthalate exposure is expected to vary significantly over the course of pregnancy. Temporal trend of phthalate exposure during pregnancy, however, is largely unknown, especially in Asian women. The purpose of this study is to investigate the urinary concentrations of metabolites for major phthalates and alternative plasticizers over the course of pregnancy among Korean (n = 81) and Thai women (n = 102). Twenty-four metabolites from 15 plasticizers, such as dimethyl phthalate (DMP), diethyl phthalate (DEP), di-isobutyl phthalate (DiBP), di-n-butyl phthalate (DnBP), benzyl butyl phthalate (BBzP), di(2-ethylhexyl) phthalate (DEHP), dioctyl phthalate (DnOP), diisononyl phthalate (DiNP), diisodecyl phthalate (DiDP), di(2-ethylhexyl) terephthalate (DEHTP), and di-(iso-nonyl)-cyclohexane-1,2-dicarboxylate (DINCH), were measured in urine samples collected in each trimester from pregnant women. While the levels of several phthalate metabolites were significantly different by trimester among Korean women, those of Thai women were relatively consistent. Urinary metabolites of DEP and DnOP were higher in Thai pregnant women compared to Korean pregnant women. The detection frequencies of the DINCH metabolite were 67.4% and 44.9% among Korean and Thai pregnant women, respectively. However, the ratio of DINCH to DEHP metabolites was significantly higher in Thai women. According to risk assessment, 11.9% of Korean and 5.3% of Thai women were considered at risk due to phthalate exposure, and DEHP, DnBP and DiBP were identified as major risk drivers. Considering the vulnerability of growing fetuses, further studies are warranted to identify major sources of exposure to these plasticizers during pregnancy.

Changes in urinary excretion of phthalates, phthalate substitutes, bisphenols and other polychlorinated and phenolic substances in young Danish men; 2009-2017.

During the past two decades human exposure to bisphenol A (BPA) and phthalates such as di-iso-butyl phthalate (DiBP), di-n-butyl phthalate (DnBP), butylbenzyl phthalate (BBzP) and di-(2-ethyl-hexyl) phthalate (DEHP) has received substantial interest due to widespread population exposures and potential endocrine disrupting effects. Therefore, these chemicals have gradually been restricted and phased out through legislation. However, humans are still exposed to a wide range of other less studied phthalates, phthalate substitutes and BPA analogues as well as other polychlorinated and phenolic substances. In this study, we investigated human exposure to these chemicals over the past decade. Three hundred urine samples collected in 2009, 2013 and 2017 (100 samples each year) from young Danish men of the general population, participating in a large on-going cross-sectional study, were selected for the present time trend study. The urinary concentration of metabolites of 15 phthalates, di-2-ethylhexyl terephthalate (DEHTP) and di-iso-nonyl-cyclohexane-1,2-dicarboxylate (DINCH), seven bisphenols including BPA, bisphenol S (BPS) and bisphenol F (BPF), as well as triclosan, triclocarban, benzophenone-3, three chlorophenols and two phenylphenols were analyzed by two new sensitive LC-MS/MS methods developed and validated for the present study. A significant decrease in urinary concentrations over time was observed for the majority of the chemicals. Median concentrations of BPA and the metabolites of DiBP, DnBP, BBzP and DEHP were more than halved from 2009 to 2017. Similar decreases were observed for triclosan and the chloro- and phenylphenols. In contrast, metabolites of the two phthalate substitutes DEHTP and DINCH increased more than 20 and 2 times, respectively. The potential BPA substitutes; BPS and BPF also increased, but only slightly. Despite these new exposure patterns, the exposure to the old well-known chemicals, such as DiBP, DnBP, BBzP, DEHP and BPA was still higher in 2017 compared to the exposure level of the new substitutes such as DEHTP, DINCH, BPS and BPF. A significant decrease in internal exposure to most of the common phthalates and BPA over the past decade was observed, reflecting market changes and regulatory measures implemented in EU. Despite increasing exposures to some of the known phthalate substitutes and BPA analogues, the total amount of each measured chemical group (original and substitute analytes combined) was lower in the more recently collected samples. This indicates only partial direct substitution or substitution by chemicals not covered in this approach, or a general decline in the exposure to these chemical/product groups over the last decade.

German Environmental Specimen Bank: 24-hour urine samples from 1999 to 2017 reveal rapid increase in exposure to the para-phthalate plasticizer di(2-ethylhexyl) terephthalate (DEHTP)

The worldwide plasticizer markets are facing constant substitution processes. Many classic ortho-phthalate plasticizers like di(2-ethylhexyl) phthalate (DEHP) are phased out, due to their proven toxicity to reproduction. Assumedly less critical, less regulated plasticizers such as di(2-ethylhexyl) terephthalate (DEHTP) are increasingly applied in consumer near products like toys, food contact materials, and medical devices. With the increasing use of DEHTP, increasing exposures of the general population have to be expected likewise. Human biomonitoring is a well-established tool to determine population exposures. In the present study we investigate the time trend of exposure to DEHTP using 24-hour urine samples of the German Environmental Specimen Bank (ESB) collected from 1999 to 2017. In these samples (60 per odd-numbered year, 600 samples in total) collected from young German adults (20–29 years, equal gender distribution) we determined four specific urinary metabolites as biomarkers of DEHTP exposure. From 1999 to 2009, the main specific urinary metabolite 5cx-MEPTP was quantifiable in <10% of the samples. Thereafter, detection rates and levels constantly increased, in line with rapidly increasing DEHTP consumption volumes. In 2017, all samples had 5cx-MEPTP levels above the limit of quantification (LOQ) with a median concentration of 3.35 μg/L (95th percentile: 12.8 μg/L). The other metabolites were detected less frequently and at lower levels but correlated well with 5cx-MEPTP robustly confirming the increasing DEHTP exposure. All 5cx-MEPTP concentrations were well below the German health based guidance value (HBM-I) of 2800 μg/L for adults. Likewise, the median calculated daily intake, based on 5cx-MEPTP measured in 2017, was 0.74 μg/kg bw∗d (95th percentile: 3.86 μg/kg bw∗d), still well below the tolerable daily intake (TDI) of 1000 μg/kg bw∗d. Based on current toxicological knowledge we can hence conclude that for the population investigated, DEHTP exposure gives no reason for immediate concern. However, the steep ongoing increase of DEHTP exposure warrants further close monitoring in the future, preferably also in sub-populations with known higher exposures to plasticizers, especially children.

Determinants and characterization of exposure to phthalates, DEHTP and DINCH among pregnant women in the PROTECT birth cohort in Puerto Rico.

Background:As a result of evidence suggesting phthalate toxicity, their use has decreased in recent years. However, new phthalates and non-phthalate replacements have emerged in their place, with unknown potential impacts on health.Methods:We measured 15 phthalate, two di(2-ethylhexyl)terephthalate (DEHTP), and two di(isononyl)cyclohexane-1,2-dicarboxylate (DINCH) urinary metabolites, collected up to three times during pregnancy from 994 women in Northern Puerto Rico (2011-2017). We used tests of linear trend to asses changes in concentrations over time and linear mixed models to identify predictors of exposure (sociodemographic characteristics, drinking water sources, diet, product use).Results:Several phthalate metabolites decreased over the study period indicating decreased exposure, while the geometric mean of DEHTP metabolites (molecular sum) increased 3-fold between 2014-2017. Intraclass correlations revealed low to moderate reproducibility of these biomarkers across pregnancy. Several metabolites were associated with maternal age, income, education, pre-pregnancy BMI, drinking public water, use of cleaning and personal care products and ice cream consumption. DINCH metabolite concentrations remained low throughout the study period.Conclusion:Although exposure to some phthalates may be decreasing, exposure to replacements, such as DEHTP, is increasing. Additional studies are needed to further characterize sources of phthalate replacement chemicals and potential exposure-related health effects among vulnerable populations.

Replacing Di(2-ethylhexyl) Terephthalate by Di(2-ethylhexyl) 2,5-Furandicarboxylate for PVC Plasticization: Synthesis, Materials Preparation and Characterization.

The worldwide regulatory demand for the elimination of non-phthalate compounds for poly(vinyl chloride) (PVC) plasticization has intensified the search for alternatives. Concomitantly, sustainability concerns have highlighted sugar-based 2,5-furandicarboxylic acid as one key renewable-chemical for the development of several products, namely di(2-ethylhexyl) 2,5-furandicarboxylate (DEHF) plasticizer. This study addresses the use of DEHF under a realistic scenario of the co-existence of both DEHF and entirely fossil-based plasticizers. More precisely, original PVC blends using mixtures of non-toxic DEHF and di(2-ethylhexyl) terephthalate ester (DEHT) were designed. The detailed structural, thermal, and mechanical characterization of these materials showed that they all have a set of interesting properties that are compatible with those of commercial DEHT, namely a low glass transition (19.2–23.8 °C) and enhanced elongation at break (up to 330%). Importantly, migration tests under different daily situations, such as for example exudation from food/beverages packages and medical blood bags, reveal very low weight loss percentages. For example, in both distilled water and phosphate buffered saline (PBS) solution, weight loss does not exceed ca. 0.3% and 0.2%, respectively. Viability tests show, for the first time, that up to 500 μM of DEHF, a promising cytotoxic profile is observed, as well as for DEHT. Overall, this study demonstrates that the combination of DEHF and DEHT plasticizers result in a noticeable plasticized PVC with an increased green content with promising cytotoxic results.

Occurrence and emission of phthalates and non-phthalate plasticizers in sludge from wastewater treatment plants in Korea.

Phthalates are endocrine-disrupting chemicals that cause adverse health effects in wildlife and humans. Due to domestic and global regulations of phthalates in commercial products, non-phthalate plasticizers (NPPs) have been introduced into industrial markets. Few studies have been conducted on the occurrence of phthalates and NPPs in sludge from wastewater treatment plants (WWTPs). In this study, sludge samples were collected from 40 WWTPs in Korea to investigate the occurrence, compositional profiles, and emission flux of phthalates and NPPs. Total concentrations of phthalates and NPPs in sludge ranged from 4.7 to 1400 (mean: 110) μg/g dry weight and from 0.17 to 780 (mean: 28.0) μg/g dry weight, respectively. Di(2-ethylhexyl)phthalate (DEHP) was a predominant compound, suggesting widespread consumption in Korea. Di(2-ethylhexyl)terephthalate (DEHT) was dominant in industrial sludge samples, whereas di-isononyl cyclohexane-1,2-dicarboxylate (DINCH) and trioctyl trimellitate (TOTM) were dominant in domestic sludge. This implies different consumption patterns of phthalate alternatives by industry and domestic activities. Concentrations of NPPs were significantly correlated with those of high-molecular-weight (HMW) phthalates, indicating that HMW phthalates were preferentially replaced by NPPs. The emission fluxes of phthalates via domestic WWTP activities were higher than those measured for industrial WWTPs, while the emission fluxes of NPPs via industrial WWTPs were higher than those found for domestic and industrial WWTPs. This indicates that phthalate emissions are associated with household activities, while NPP emissions are associated with industrial activities.

Changes in Status of Plasticizers Used in Polyvinyl Chloride Toys

Analytical survey of plasticizers used in about 500 polyvinyl chloride (PVC) toys in Japanese market was performed in 2014. The results obtained were compared with those obtained in 2009. Fifteen types of plasticizers including di (2-ethylhexyl) terephthalate (DEHTP) were detected. These plasticizers were also detected in the products in 2009, suggesting that the types of plasticizers used in PVC toys have not been changed. Among these detected plasticizers, the detection ratios of DEHTP were 60.3 and 73.7% in both designated and not-designated toys, respectively, both of which are the highest. These ratios are over 20-points higher than those in 2009. The six types of phthalic acid esters (PAEs) prohibited to use for the designated toys in Japan were not detected in designated toys used in this study. However, four types of PAEs, including three types of prohibited PAEs, were detected in not-designated toys with the detection ratios from 2.8 to 15.5%. These ratios are lower than those in 2009. The content levels of plasticizers used in the toys are lower than those in 2009. These results show that the main plasticizer used in PVC toys is DEHTP, and that the usage levels of plasticizers have been decreased.

In vitro and in silico hormonal activity studies of di-(2-ethylhexyl)terephthalate, a di-(2-ethylhexyl)phthalate substitute used in medical devices, and its metabolites.

Plasticizers added to polyvinylchloride used in medical devices can be released into patients' biological fluids. The substitution of di-(2-ethylhexyl)phthalate (DEHP) by alternative plasticizers is essential but their safety must be demonstrated. DEHP, di-(2-ethylhexyl)terephthalate (DEHT) and their metabolites were investigated using level 2 Organization for Economic Co-operation and Development bioassays to screen for in vitro hormonal changes. Differences between the DEHP and DEHT metabolites were observed. Albeit weak, the hormonal activities of DEHT-derived metabolites, e.g., 5-OH metabolite of mono-(ethylhexyl)terephthalate (5-OH-MEHT), were detected and the results of docking experiments performed on estrogen receptor alpha and androgen receptor agreed with the biological results. A co-stimulation of human estrogen receptor alpha and human androgen receptor was also observed. With regard to steroidogenesis, a 16-fold increase in estrogen synthesis was measured with 5-OH-MEHT. Therefore, even if DEHT remains an interesting alternative to DEHP because of its low migration from medical devices, it seems important to verify that multi-exposed patients in neonatal intensive care units do not have urinary levels of oxidized metabolites, in particular 5-OH-MEHT, suggesting a potential endocrine-disrupting effect.

Development and validation of a bioanalytical assay based on liquid chromatography-tandem mass spectrometry for measuring biomarkers of exposure of alternative plasticizers in human urine and serum

Alternative plasticizers (APs) have been increasingly used in the last decade to replace conventional phthalate esters, in particular di(2-ethylhexyl) phthalate (DEHP), due to the toxicity of the latter. However, there is currently very little data about the toxicity of and exposure to APs. No method exists so far for the analysis of multiple exposure biomarkers. The objective of this work consisted in developing a simple bioanalytical procedure for the analysis of multiple exposure biomarkers of APs in human urine and serum. Focus was set on metabolites of di(2-propylheptyl) phthalate (DPrHpP), di(isononyl)cyclohexane-1,2-dicarboxylate (DINCH), di(2-ethylhexyl) terephthalate (DEHTP) and di-2-ethylhexyl adipate (DEHA). A sample preparation protocol was developed and optimized using Oasis HLB solid-phase extraction (SPE) cartridges. Subsequently, an instrumental method based on liquid-chromatography coupled to tandem mass spectrometry (LC-MS/MS) was optimized. Following established guidelines, the sample preparation and instrumental methods were validated in terms of recovery, matrix effects, carry-over, linearity, limits of quantification, within- and between-run precision and trueness. Obtained results were satisfactory for all compounds except for one of the metabolites of DEHA (i.e., mono(2-ethylhexyl) adipate (MEHA)). A pilot biomonitoring study was carried out to assess the method's ability to detect and quantify target analytes in human urine and serum. In urine, most analytes could be detected with frequencies ranging from 8% for mono(2-ethyl-5-hydroxyhexyl) adipate (OH-MEHA) and cyclohexane-1,2-dicarboxylic mono hydroxyisononyl ester (OH-MINCH) to 92% for mono(2-ethyl-5-oxohexyl) adipate (oxo-MEHA), whilst most compounds could not be detected in serum, except for mono(2-ethylhexyl) terephthalate (MEHTP) and mono-(2-propyl-6-hydroxyheptyl) phthalate (OH-MPrHpP) which were detected in all samples. The obtained results show that the developed method can be used to simultaneously analyse multiple exposure biomarkers to APs in human urine and serum.

Hair as an alternative matrix to monitor human exposure to plasticizers – Development of a liquid chromatography - tandem mass spectrometry method

The application and production of alternative plasticizers (APs) has been increasing in the last decade in replacement of conventional phthalates due to their toxicity. This calls for the development of non-invasive monitoring approaches to assess human exposure to APs. A method for the simultaneous measurement of exposure biomarkers of di(2‑propylheptyl) phthalate (DPHP), di(isononyl)cyclohexane‑1,2‑dicarboxylate (DINCH), di(2‑ethylhexyl) terephthalate (DEHTP) and di‑2‑ethylhexyl adipate (DEHA) in hair samples was developed and validated in this study. Prior to the analysis, the hair samples were washed in acetone and ultrapure water and pulverized to powder. Further, a solid-liquid and solid-phase extraction, followed by quantification using liquid chromatography - tandem mass spectrometry (LC-MS/MS) was performed. The method was validated in terms of recovery, matrix effects, carry-over, linearity, limits of quantification, within- and between-run precision and trueness. Satisfying results were obtained for all targeted compounds, except for mono(2‑ethylhexyl) adipate (MEHA), which was monitored only qualitatively. The optimized method was implemented in a pilot biomonitoring study with hair samples from 9 healthy volunteers. Detection frequencies of seven metabolites ranged from 11% to 100%. Mono(2‑ethylhexyl) terephthalate (MEHTP) and mono(2‑ethyl‑5‑oxohexyl) adipate (oxo-MEHA) were found in all hair samples. More hydrophobic monoester metabolites were found to be incorporated in hair to a greater extent compared to their oxidized counterparts. Obtained results show that the developed method can detect AP metabolites in hair, supporting the use of this alternative matrix in human biomonitoring.

German Environmental Specimen Bank: Urine Samples from 1999-2017 Document Rapid Increase in Exposure to the Para-Phthalate Plasticizer DEHTP

Di(2-ethylhexyl) terephthalate (DEHTP) is increasingly used as an alternative to reprotoxic ortho-phthalate plasticizers, e.g. di(2-ethylhexyl) phthalate (DEHP), in everyday products such as food contact materials, toys and medical devices. In Western Europe, DEHTP consumption rose from 2,000 t in 2002, to 100,000 t in 2014 and a predicted 135,000 t in 2019.To investigate for possible increases of DEHTP exposure in the German population over time, we analyzed 600 24-hour urine samples of the German Environmental Specimen Bank for specific DEHTP metabolites as exposure biomarkers. Samples were collected in the years 1999-2017 (ten sampling years, 30 male and 30 female volunteers each, aged 20-29). The samples were blinded and randomized before analysis. Four urinary DEHTP metabolites (side-chain oxidized monoesters) were determined by online SPE LC-MS/MS with isotope dilution quantification. The limits of quantification were 0.2 µg/L (5cx-MEPTP and 5oxo-MEHTP), 0.3 µg/L (5OH-MEHTP) and 0.4 µg/L (2cx-MMHTP).From 1999 until 2009 the main oxidized DEHTP metabolite 5cx-MEPTP was rarely detected. Afterwards, detection rates increased significantly (8% in 2009, 42% in 2011, 95% in 2013, 98% in 2015, and 100% in 2017). The median concentration for 5cx-MEPTP increased from &amp;lt;LOQ in 1999 to 3.35 µg/L in 2017. Detection rates and concentrations for the other metabolites were lower, as expected from human metabolism studies, but strongly correlated with 5cx-MEPTP. Overall, urinary 5cx-MEPTP concentrations were well below the health based guidance value (HBM-I) of 2800 µg/L, and calculated daily intakes were well below the tolerable daily intake of 1 mg/kg bw*d.For the first time, we document the increasing exposure to DEHTP of the German population in a robust set of 600 24-hour urine samples collected between 1999 and 2017. Although considered uncritical for now, the steep increase in DEHTP exposure calls for a continued exposure monitoring.

LC-HRMS Biomarker Suspect Screening in Low Dose Studies

Exposure biomarker development is a time consuming procedure, starting with identification of presumably appropriate metabolites of the xenobiotic in question. Investing into unsuitable biomarker candidates (i.e. metabolites which are formed in low proportion or are not formed at all) can be costly. Accordingly, efficient strategies for the identification of promising biomarker candidates are needed. For this purpose, liquid chromatography coupled with high resolution mass spectrometry (LC-HRMS) is often applied for metabolite screening in in vitro and animal in vivo screening approaches, typically involving high doses/concentrations of the xenobiotic.Given the limitations of in vitro models and interspecies differences in metabolism, we sought to improve the certainty of metabolite screening by analyzing urine samples from human studies with low oral dosing (i.e. toxicologically uncritical). Metabolite screening was performed by LC-Q-Orbitrap-MS combined with a commercial data mining software. Identification criteria were plausible excretion kinetics, product ion spectra, 13C isotopomer ratios, and retention times.We present results of a pilot study, which identified nine metabolites of the plasticizer di (2-ethylhexyl) terephthalate (DEHTP). Three of them (5OH-MEHTP, 5Oxo-MEHTP, and 5cx-MEPTP) are already established biomarkers of DEHTP exposure. Five additional, specific metabolites have not been described previously. In another example we successfully identified several tentative metabolites of the UV filter avobenzone. However, we also report examples, in which metabolite identification did not succeed by this approach (e.g. the plasticizer DEHA) and we highlight influencing factors (such as mobile phase pH, in source adduct formation and fragmentation) possibly leading to wrong conclusions.The presented approach is a valuable but not universal tool for biomarker development. In some cases, a more sensitive triple quadrupole screening can be superior.

Exposure of Nail Salon Workers to Phthalates and Organophosphate Esters

Nail salons are one of the fastest growing industries in the USA; there is concern about health impacts associated with nail salon exposures in a vulnerable, largely immigrant worker population. Nail technicians are exposed to semi volatile organic compounds including phthalates and organophosphate esters (OPEs). In this pilot study we collected spot urine samples during one work shift from 10 female nail technicians in the Boston (USA) area in 2016-17. We assessed occupational exposures using urinary biomarkers in pre- and post-work shift samples; we compared post-shift concentrations with the general US female population from the National Health and Nutrition Examination Survey (NHANES) 2011-12 for phthalates, 2013-14 for OPEs. We detected 12 phthalate and 4 OPE urinary biomarkers in &amp;#8805; 60% of participants&amp;#8217; samples. After creatinine correction, geometric mean (GM) post-shift concentrations were higher than pre-shift for all but one phthalate and one OPE biomarker. Triphenyl phosphate is used as a phthalate replacement in some nail polishes; the GM of its urinary metabolite increased slightly (23%) from pre- to post-shift; post-shift concentrations were slightly higher than NHANES females (17%). The greatest pre- to post-shift change was for mono-2-ethyl-5-carboxypentyl terephthalate (MECPTP), which more than tripled from 11.7 to 36.6 µg/g creatinine. Phthalate biomarkers reported in both studies were lower in our study participants than in NHANES females. NHANES did not report MECPTP, a recent study found increasing trends from 2000-2016 of MECPTP in urine from a convenience sample of US adults. Di-2-ethylhexyl terephthalate (DEHTP), the parent compound of MECPTP, is a plasticizer used as a phthalate substitute in consumer products such as food packaging, toys, and perhaps nail polish. Our results suggest that nail technicians are occupationally exposed to certain phthalates and OPEs, with a urine metabolite of DEHTP showing the largest increase across a work day.

De-novo identification of specific exposure biomarkers of the alternative plasticizer di(2-ethylhexyl) terephthalate (DEHTP) after low oral dosage to male volunteers by HPLC-Q-Orbitrap-MS

Context: Human exposure biomonitoring relies on the availability of specific, sensitive biomarkers. For emerging chemicals, the identification (prediction, synthesis, verification) of such biomarkers is time and cost intensive.Objective: This study aimed to further elucidate the urinary metabolic profile of the plasticizer di(2-ethylhexyl) terephthalate (DEHTP) in search of probably additional biomarkers of exposure.Materials and methods: Urine samples of an oral low-dose volunteer study were analysed by HPLC-Q-Orbitrap-MS combined with a commercial data mining software. Metabolite identification was based on isotopic pattern, accurate masses of product ions and excretion profiles.Results: Nine phase I metabolites of DEHTP were tentatively identified by HPLC-Q-Orbitrap-MS. Four previously described, side chain oxidized monoester metabolites were confirmed in all samples. In addition, five previously unknown downstream metabolites were tentatively identified.Discussion and conclusion: The excretion profiles obtained by HPLC-Q-Orbitrap-MS were in good agreement with quantitative HPLC-QqQ-MS data. For the newly discovered metabolites, plausible excretion profiles, similar to the ones of the known metabolites, were obtained. The presented approach proved to be successful for metabolite screening in urine samples after low-dose exposure and will be applied in future human metabolism studies for a fast, reliable and cost effective identification of specific biomarkers of exposure.

Studies on the development of antibodies for the highly hydrophobic plasticizers DINCH and DEHT

Diisononylcyclohexane-1,2-dicarboxylate (DINCH) and di-2-ethylhexyl terephthalate (DEHT), two of the most important substitutes for phthalate plasticizers, are used for a wide range of applications. Consequently, an increasing occurrence in urine and environmental samples is reported. Reliable and fast analytical methods for the quantification of these plasticizers are needed. So far, mainly GC-MS or LC-MS methods are used. We aimed to develop the first antibodies and immunoassays allowing for high-throughput analysis of samples. We designed two DINCH hapten structures and one DEHT hapten structure and employed hapten-protein conjugates for the immunization of rabbits. Sensitive competitive enzyme-linked immunosorbent assays (ELISAs) against each hapten using the produced polyclonal antibodies were established. Yet, binding of DINCH to the respective antibodies was not observed in neither direct nor indirect assay formats, even when using protein conjugates with the heterologous haptens and different carrier proteins in the indirect format. The use of surfactants and solvents in the sample buffer did not result in recognition of the plasticizers. Also, no binding of DEHT in ELISA employing the respective antibodies was detected. We speculate that the production of antibodies against these highly hydrophobic molecules is not possible via our route, however a different hapten design could overcome this obstacle.