Biotransformation and toxicokinetics of 2-phenoxyethanol after oral exposure in humans: a volunteer study.

2-Phenoxyethanol (PhE) is an aromatic glycol ether and is used in a variety of functions and applications, e.g., as preservative in pharmaceuticals, cosmetic and personal care products, as biocide in disinfectants (e.g. human hygiene), or as a solvent in formulations (e.g. coatings, functional fluids). Despite its widespread use, little is yet known on its biotransformation and toxicokinetics in humans. Therefore, a pilot study was conducted with oral administration of PhE (5 mg/kg body weight) to five volunteers. Blood and urine samples were collected and analyzed for PhE and three of its presumed metabolites up to 48 h post-exposure. Additionally, one volunteer was dermally exposed to PhE and monitored until 72 h post-exposure. PhE was rapidly resorbed following both oral and dermal application with tmax-levels in blood of about 1 h and 3 h, respectively. Metabolism of PhE was observed to be rather extensive with phenoxyacetic acid (PhAA) and 4-hydroxyphenoxyacetic acid (4-OH-PhAA) as the main metabolites found in blood and urine following oral and dermal exposure. PhE was excreted rapidly and efficiently via urine mostly in metabolized form: following oral exposure, on average 77% and 12% of the applied dose was excreted within 48 h as PhAA and 4-OH-PhAA, respectively. A similar metabolism pattern was observed following the single dermal exposure experiment. The obtained data on biotransformation and toxicokinetics of PhE in humans provide valuable information on this important chemical and will be highly useful for pharmacokinetic modelling and evaluation of human PhE exposure.

A specific and sensitive GC-MS/MS method for the quantitative determination of 2-phenoxyethanol and selected metabolites in human blood and urine.

2-Phenoxyethanol (PhE) is widely used as a preservative in consumer products such as cosmetics as well as at the workplace as a component of metal-working fluids and hydraulic fluids. Therefore, both industry workers and consumers may potentially be exposed to PhE. An analytical method for the quantification of PhE and three selected metabolites, namely phenoxyacetic acid (PhAA), 4-hydroxyphenoxyacetic acid (4-OH-PhAA), and 4-hydroxyphenoxyethanol (4-OH-PhE) in human urine and blood was developed and validated. The sample preparation includes enzymatic hydrolysis of urine samples or protein precipitation of blood samples, followed by liquid-liquid extraction and silylation of the target analytes. Analyses of the extracts were carried out by gas chromatography with tandem mass spectrometry (GC-MS/MS). 3,4-Hydroxyphenoxyethanol, a probably minor PhE metabolite could not reliable be analyzed due to its instability. The limits of quantification (LOQ) of the analytes ranged between 0.5 and 6.1 µg/L and 2.0 and 3.9 µg/L in urine and blood, respectively. The method was successfully applied to spot urine samples of 50 individuals without occupational exposure to PhE and additionally to blood samples from seven volunteers. In urine, PhAA and 4-OH-PhAA could be quantified in all analyzed samples, whereas 4-OH-PhE and unchanged PhE were found in 36 % and 32 % of the samples, respectively. In blood, PhAA was also found in every sample in levels above the LOQ, whereas PhE itself was detected in three of seven samples only. Neither 4-OH-PhAA nor 4-OH-PhE were found in any of the analyzed blood samples. The developed method promises to be a valuable tool for PhE monitoring of urine and blood samples and may also enable an advanced investigation of PhE biotransformation pathways in humans.

Human Metabolism and Urinary Elimination Kinetics of the Fragrance Geraniol after Oral Dosage.

Geraniol is a fragrance with a characteristic rose-like smell, naturally occurring in terpene oil and also chemically synthesized on a large scale. Geraniol is widely used in consumer products such as cosmetics, personal care products, and household cleaners and as an additive in foods. An experimental study in human volunteers was carried out to investigate the metabolism and elimination kinetics of geraniol. Three subjects were orally exposed to geraniol in two different dosages (25 or 250 mg). In each case, one pre-exposure urine sample and all urine voids for 72 h after exposure were collected separately. The geraniol metabolites Hildebrandt acid, geranic acid, 3-hydroxycitronellic acid, and 8-carboxygeraniol were analyzed in every sample after enzymatic hydrolysis and liquid-liquid extraction using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Maximum urinary concentrations of the metabolites were measured between 1 and 5 h after oral dosing, and elimination half-lives were determined to be about 2-4 h. The predominant metabolite found in urine was Hildebrandt acid with 34.4 ± 5.6% of the ingested dose, followed by geranic acid (12.7 ± 5.6%), 3-hydroxycitronellic acid (2.2 ± 0.4%), and 8-carboxygeraniol (0.19 ± 0.09%). In total, the four metabolites determined represent 41.7-55.5% of the ingested dose. Only 8-carboxygeraniol is, however, a specific metabolite, while the other three target analytes are also formed from other terpenes like citral. Within this study, conversion factors were calculated, which allow for a rough estimate of the total geraniol uptake by back-calculation from metabolite concentrations of spot urine samples. Taking the conversion factor for all four metabolites into account, a mean daily uptake of geraniol of 1.43 mg was estimated from 41 urine samples of occupationally nonexposed adults. The metabolites Hildebrandt acid, geranic acid, 3-hydroxycitronellic acid, and 8-carboxygeraniol in urine are suitable biomarkers of exposure for geraniol and can be used for human biomonitoring studies.

Human metabolism and excretion kinetics of the surfactant 2,4,7,9-tetramethyl-5-decyne-4,7-diol (TMDD) after oral and dermal administration.

2,4,7,9-Tetramethyl-5-decyne-4,7-diol (TMDD) is a non-ionic surfactant with a wide range of applications. TMDD is considered a high-production chemical and, due to its low biodegradation rate, possesses a potentially high prevalence in the environment. However, despite its widespread use, toxicokinetic data and data on internal exposure to TMDD in the general population are completely lacking. Hence, we developed a human biomonitoring (HBM) method for TMDD. Our approach included a metabolism study with four subjects, who were administered an oral dose of 75 µg TMDD/kg body weight and a dermal dose of 750 µg/kg body weight. Terminal methyl-hydroxylated TMDD (1-OH-TMDD) was previously identified as the main urinary metabolite in our lab. The results of the oral and dermal applications were used to determine the toxicokinetic parameters of 1-OH-TMDD as a biomarker of exposure. Finally, the method was applied to 50 urine samples from non-occupationally exposed volunteers. Results show that TMDD was rapidly metabolized, with an average tmax of 1.7 h and a rapid and almost complete (96%) excretion of 1-OH-TMDD until 12 h after oral dosage. Elimination was bi-phasic, with half-lives of 0.75-1.6 h and 3.4-3.6 h for phases 1 and 2, respectively. The dermal application resulted in a delayed urinary excretion of this metabolite with a tmax of 12 h and complete excretion after about 48 h. The excreted amounts of 1-OH-TMDD represented 18% of the orally administered TMDD dose. The data of the metabolism study demonstrated a fast oral as well as substantial dermal resorption of TMDD. Moreover, the results indicated an effective metabolism of 1-OH-TMDD, which is excreted rapidly and completely via urine. Application of the method to 50 urine samples revealed a quantification rate of 90%, with an average concentration of 0.19 ng/mL (0.97 nmol/g creatinine). With the urinary excretion factor (Fue) derived from the metabolism study, we estimated an average daily intake of 1.65 µg TMDD from environmental and dietary sources. In conclusion, 1-OH-TMDD in urine is a suitable biomarker of exposure to TMDD and can be applied for biomonitoring of the general population.

Determination of a specific metabolite for the non-ionic surfactant 2,4,7,9-Tetramethyl-5-decyne-4,7-diol (TMDD) by UPLC-MS/MS.

2,4,7,9-Tetramethyldec-5-yne-4,7-diol (TMDD) is a non-ionic surfactant commonly used as defoaming agent and numerous other applications. Effluents of wastewater treatment plants have been identified as one of the main sources of TMDD emissions into the environment. Due to its broad application in various fields, TMDD was selected for the development of a biomonitoring method for assessing human exposure within the frame of the cooperation project of the German Federal Ministry for the Environment, Nature Conservation, Building and Nuclear Safety (BMUB) and the German Chemical Industry Association (VCI) in 2020. This study aimed to identify a urinary metabolite for TMDD by UPLC-Q-Orbitrap-MS which can be used as a biomarker of TMDD exposure. Monohydroxylated TMDD (1-OH-TMDD) was deciphered as the most prominent metabolite of TMDD in humans in a series of in vitro and in vivo experiments. In a next step, a quantitative method for the determination of 1-OH-TMDD was developed and validated. Quantification was achieved by isotope dilution using D3-1-OH-TMDD as internal standard. The method is characterized by a simple sample clean-up procedure and an enzymatic hydrolysis of possible metabolite conjugates with ß-glucuronidase. Method validation was performed according to international guidelines for bioanalytical method validation. The method proved its robustness, precision, accuracy and sensitivity for the intended purpose, i.e. the assessment of TMDD exposure in the general population by means of human biomonitoring.

Human metabolism and excretion kinetics of benzotriazole UV stabilizer UV-327 after single oral administration.

UV-327 (2-(5-chloro-benzotriazol-2-yl)-4,6-di-(tert-butyl)phenol) is used as an ultraviolet (UV) absorber in plastic products and coatings. Due to its ubiquitous distribution in the environment, human exposure is conceivable. In the study presented herein, initial information on the human in vivo metabolism of UV-327 was obtained by single oral administration to three volunteers. Urine and blood samples were collected up to 72 h after exposure. One study participant additionally donated plasma samples. Maximum blood and plasma levels of UV-327 and its two monohydroxylated metabolites UV-327-6-mOH and UV-327-4-mOH were reached 6 h post-exposure. Almost the entire amount found in blood and plasma samples was identified as UV-327, whereas the two metabolites each accounted for only 0.04% of the total amount, indicating that UV-327 is well-absorbed from the intestine, but only partially metabolized. Plasma to blood ratios of UV-327, UV-327-6-mOH, and UV-327-4-mOH ranged from 1.5 to 1.6. Maximum urinary excretion rates of UV-327, UV-327-6-mOH, UV-327-4-mOH, and UV-327-4 + 6-diOH were reached 9-14 h post-exposure. However, only about 0.03% of the orally administered dose of UV-327 was recovered as UV-327 and its metabolites in urine, indicating that biliary excretion may be the major route of elimination of UV-327 and its hydroxylated metabolites. The present study complements the insight in the complex absorption, distribution, metabolism, and elimination (ADME) processes of benzotriazole UV stabilizers (BUVSs).

Reliable determination of the main metabolites of 2-phenoxyethanol in human blood and urine using LC-MS/MS analysis.

2-Phenoxyethanol (PhE) is used as a broad-spectrum preservative in several consumer products like cosmetics and cleaning agents. To enable the analysis and assessment of human exposure to PhE, a fast and sensitive LC-MS/MS method for the quantification of two PhE metabolites, namely phenoxyacetic acid (PhAA) and 4-hydroxyphenoxyacetic acid (4-OH-PhAA) in human urine and blood was developed and validated. The method is based on liquid chromatography combined with tandem mass spectrometry (LC-MS/MS). Sample preparation was different for both matrices: either a simple "dilute&shoot"-approach for urine samples or a liquid-liquid-extraction (LLE) for blood samples was used. The limit of quantification (LOQ) is 10 µg L-1 and 6 µg L-1 for PhAA and 20 µg L-1 and 10 µg L-1 for 4-OH-PhAA in urine and blood, respectively. The method was applied to urine samples of 153 persons without occupational exposure to PhE and to blood samples of 7 additional volunteers. In blood, PhAA was detected in 57% of all samples (range:

Human metabolism and kinetics of the UV absorber 2-(2H-benzotriazol-2-yl)-4,6-di-tert-pentylphenol (UV 328) after oral administration.

2-(2H-Benzotriazol-2-yl)-4,6-di-tert-pentylphenol (UV 328; CAS: 25973-55-1) is an ultraviolet light (UV) absorber which belongs to the class of hydroxy phenol benzotriazoles. Therefore, UV 328 is added to plastics and other polymers due to its photostability to prevent discoloration and prolong product stability which may result in an exposure of consumers. However, information about the toxic effects on humans and the human metabolism are still lacking. In the present study, human metabolism pathways of UV 328 and its elimination kinetics were explored. For that purpose, three healthy volunteers were orally exposed to a single dose of 0.3 mg UV 328/kg bodyweight. UV 328 and its metabolites were investigated in blood and urine samples collected until 48 and 72 h after exposure, respectively. Thereby, previously published analytical procedures were applied for the sample analysis using dispersive liquid-liquid microextraction and subsequent measurement via gas chromatography coupled to tandem mass spectrometry with advanced electron ionization. UV 328 was found to be oxidized at its alkyl side chains leading to the formation of hydroxy and/or oxo function with maximum blood concentrations at 8-10 h after exposure for UV 328-6/3-OH, UV 328-4/3-OH and UV 328-4/3-CO. In contrast, a plateau for UV 328-4/3-CO-6/3-OH levels was reached around 10 h post-dosage. The highest blood levels were found for native UV 328 at 8 h after ingestion. Furthermore, biphasic elimination kinetics in blood were revealed for almost all detected metabolites. UV 328 and its metabolites did not occur in blood as conjugates. The renal elimination kinetics were very similar with the kinetics in blood. However, the prominence of the metabolites in urine was somewhat different compared to blood. In contrast, mostly conjugated metabolites occurred for renal elimination. In urine, UV 328-4/3-CO-6/3-OH was found to be the most dominant urinary biomarker followed by UV 328-6/3-OH and UV 328-4/3-OH. In total, approximately 0.1% of the orally administered dose was recovered in urine within 72 h. Although high levels of UV 328 in blood proved good resorption and high systemic availability of the substance in the human body, the urine results revealed a rather low quantitative metabolism and urinary excretion rate. Consequently, biliary excretion as part of the enterohepatic cycle and elimination via feces are assumed to be the preferred pathways instead of renal elimination.

Quantitative determination of urinary metabolites of geraniol by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS).

Geraniol is a fragrance which occurs in natural terpene oil or is chemically synthesized on a large scale. It is used in a wide variety of consumer products such as perfumes, deodorants, household products and cosmetics. Hence, not only industry workers in the production of geraniol, but also consumers can come into contact with the substance. Human biomonitoring (HBM), i.e. the analytical determination of substances and their metabolites in human biological material, is a key element in the analysis and assessment of the distribution and intensity of occupational and environmental exposure of humans. Therefore, a procedure for the quantitative determination of the urinary metabolites Hildebrandt acid, geranic acid, 3-hydroxycitronellic acid and 8-carboxygeraniol as potential biomarkers of geraniol exposure was developed and validated. The method is based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) after enzymatic hydrolysis and liquid-liquid extraction (LLE) of the target analytes. The limit of quantification (LOQ) is 1.5 µg L-1 for 8-carboxygeraniol, 2.7 µg L-1 each for Hildebrandt acid and geranic acid, and 1.8 µg L-1 for 3-hydroxycitronellic acid. The method was applied to urine samples of 41 persons without occupational exposure to geraniol. Hildebrandt acid and geranic acid were detected in all samples, 8-carboxygeraniol in 83% and 3-hydroxycitronellic acid in 81% of the samples. Hildebrandt acid (median: 313 µg L-1, range: 37-1966 µg L-1) was the most abundant metabolite, followed by geranic acid (93 µg L-1; 9-477 µg L-1), 3-hydroxycitronellic acid (18 µg L-1;

Development of a human biomonitoring method for assessing the exposure to ethoxyquin in the general population.

Ethoxyquin (EQ) is commonly used as an antioxidant in animal feeds. Although EQ is not permitted for usage in food products for humans within the EU, residues of EQ and its transformation products could be determined in food of animal origin. Despite its widespread use and concerns on its toxicological profile, no information about the systemic exposure to EQ in the general population is available. Hence, we developed a human biomonitoring (HBM) method for EQ. Our approach included a metabolism study with five subjects, who were administered an oral dose of 0.005 mg EQ/kg body weight. Unchanged EQ and the major metabolite 2,2,4-trimethyl-6(2H)-quinolinone (EQI) were identified as urinary excretion products of EQ. While small amounts of EQ could be determined in high concentrated samples from the metabolism study only, 28.5% of the orally applied EQ dose could be recovered as EQI. Toxicokinetic parameters were determined for EQI, the potential biomarker of exposure. In addition, an analytical method for EQI (LOQ = 0.03 µg/L) in urine based on UHPLC-MS/MS comprising enzymatic glucuronide hydrolysis and salt-assisted liquid-liquid extraction was developed, validated and applied to 53 urine samples from the general population. EQI could be quantified in 11 (21%) of the samples in levels up to 1.7 µg/L urine, proving the suitability of the developed method for the intended purpose.

Identification of in vitro phase I metabolites of benzotriazole UV stabilizer UV-327 using HPLC coupled with mass spectrometry.

The benzotriazole UV stabilizer (BUVS) 2-(5-chloro-benzotriazol-2-yl)-4,6-di-(tert-butyl)phenol (UV-327) is used in various plastic products to protect them against harmful UV radiation. Meanwhile, there are concerns about potential adverse health effects on humans, as residues of UV-327 and other BUVSs have already been detected in various environmental matrices. However, information on the metabolism of UV-327 is not yet available. Therefore, in vitro experiments with human liver microsomes (HLMs) were performed in order to identify phase I metabolites to be used as specific biomarkers of exposure in biomonitoring studies. The samples were analyzed by HPLC coupled with mass spectrometry (HPLC/MS). Potential metabolites, which were formed by hydroxylation and further oxidation to carboxylic acid, were tentatively identified. Special metabolite structures were suspected and custom-synthesized as reference substances for verification. In total, seven phase I metabolites, which may be suitable biomarkers for the assessment of exposure to UV-327, have been identified and quantified. The results of the present study provide initial insights into the metabolic pathway of UV-327, which is essential for further research on its human metabolism.

Human metabolism and urinary excretion kinetics of the UV filter Uvinul A plus® after a single oral or dermal dosage.

Hexyl 2-[4-(diethylamino)-2-hydroxybenzoyl]benzoate, better known under its trading name Uvinul A plus® is a UV filter mainly used in sunscreens, but also present in other cosmetic products with a maximum concentration of 10% (w/w) according to the EU directive. In this study we investigated the human metabolism after a single oral and a single dermal dose of Uvinul A plus®, respectively. Samples collected within 72 h of administration were analyzed with a newly developed UHPLC-MS/MS method. Results of the study revealed three major urinary metabolites, namely 2-(4-amino-2-hydroxybenzoyl)benzoic acid (AHB), 2-(4-(ethylamino)-2-hydroxybenzoyl)benzoic acid (EHB) and 2-(4-(diethylamino)-2-hydroxybenzoyl)benzoic acid (DHB), representing 52% of the administered oral dose. The three major metabolites are further converted into four minor metabolites with an additional hydroxyl group in the aniline moiety. Toxicokinetic parameters (amount excreted, tmax, elimination constant and half-life t1/2) and conversion factors were determined for the three major metabolites. The conversion factors were used to estimate the mean daily exposure to Uvinul A plus® in spot urine samples from 58 volunteers not intentionally exposed to Uvinul A plus® derived from a pilot study. The three major metabolites were quantifiable in 26% of the samples. In 35% of the samples, at least one major metabolite could be quantified. The daily systemic exposure to Uvinul A plus® was estimated to approximately 8.1-9.3 µg/d by applying the combined conversion factor for all three major metabolites. In conclusion, a very low systemic exposure to DHHB was observed with regard to the no observed adverse effect level (NOAEL) as an established threshold for chronic uptake.

A validated UPLC-MS/MS method for the determination of urinary metabolites of Uvinul® A plus.

Uvinul® A plus (DHHB) is a synthetic benzophenone derivative mainly used in sunscreens, and also in other skin care products. The compound is authorized by the EU as UV filter and a maximum concentration of 10% in consumer products is permitted. Despite its high production volume and usage in consumer products,to date, no information about the systemic exposure to Uvinul® A plus in humans is available. Therefore, we developed a human biomonitoring method which allows the simultaneous determination of three major metabolites of Uvinul® A plus in human urine samples. Furthermore, three minor metabolites of Uvinul® A plus were identified by ion trap experiments. Urine samples were enzymatically hydrolyzed, extracted via liquid-liquid extraction with ethyl acetate, and analyzed by means of UPLC-MS/MS. The final method was validated according to FDA guidelines and applied to 58 urine samples retrieved from the general German population. The three major and specific metabolites of Uvinul® A plus were found in about 36% of the samples, proving the suitability of the method for future human biomonitoring studies.

Derivation of safe exposure levels for potential migration of formaldehyde into food.

Polyoxymethylene (POM) is a polymer of formaldehyde used inter alia for kitchenware and food processing machines. By migration into food, consumers may be exposed to small additional amounts of formaldehyde in food. In order to address such potential exposures, Specific Migration Limits are derived using all studies with oral exposure in mammals and birds. The assessment is not only based on local irritation observed in a 2-year rat study that has previously served to calculate acceptable exposure levels, but also on systemic effects, namely on effects on the kidney in adult rats and testes in birds before sexual maturity. At the relatively high oral exposure levels (up to 2000 ppm in drinking water) long-term effects caused by formic acid, the first step metabolite of formaldehyde, such as acidosis, cannot be excluded. The lowest Specific Migration Limit of 2.74 mg/dm2, corresponding to 16.5 mg formaldehyde/kg food, is based upon kidney effects in rats, leading to potential exposures that range between 2900 and 4400 times below the endogenous turnover of formaldehyde. Lastly, a recent migration study with POM showed that migration of formaldehyde into food simulants is over an order of magnitude below the lowest Specific Migration Limit derived herein.

Oxidative phase I metabolism of the UV absorber 2-(2H-benzotriazol-2-yl)-4,6-di-tert-pentylphenol (UV 328) in an in vitro model with human liver microsomes.

2-(2H-Benzotriazol-2-yl)-4,6-di-tert-pentylphenol (UV 328, CAS: 25973-55-1) is an ultraviolet light (UV) absorber which is used as an additive for plastics and other polymeric substances to prevent the host material from light induced degradation reactions. However, no information about human exposure, metabolism and kinetics is available for this substance so far. Therefore, in vitro experiments with human liver microsomes were performed to derive oxidative phase I metabolites of UV 328 in an explorative approach using liquid-chromatography coupled with tandem mass spectrometry. Initially, a suspect screening mode was applied to the incubated samples. Six metabolites with hydroxy or oxo groups as well as a metabolite carrying both hydroxy and carbonyl moieties at the alkyl side chains were postulated and custom synthesized as reference standards. Afterwards, the results were verified in a target screening approach. Thereby, five of the six investigated analyte structures were confirmed. Quantitative estimations of the generated transformation products revealed 2-(2H-benzotriazol-2-yl)-6-(3-hydroxy-2-methylbutan-2-yl)-4-(tert-pentyl)phenol (UV 328-6/3-OH), 2-(2H-benzotriazol-2-yl)-4-(3-hydroxy-2-methylbutan-2-yl)-6-(tert-pentyl)phenol (UV 328-4/3-OH) and 2-(2H-benzotriazol-2-yl)-4-(2-methylbutan-3-on-2-yl)-6-(3-hydroxy-2-methylbutan-2-yl)phenol (UV 328-4/3-CO-6/3-OH) as most promising parameters. In summary, oxidation of both alkyl side chains at the phenol moiety was proven, but no metabolic transformations at the benzotriazole moiety were observed.

Urinary metabolites of the UV filter octocrylene in humans as biomarkers of exposure.

Octocrylene (OC) is a UV filter used in sun screens and other personal care products, but also in polymers and food contact materials for stabilization. In this study, we investigate human OC metabolism and urinary excretion after oral dosage of approx. 5 mg OC [≙ 61.8-89.5 µg/(kg body weight)] in three male volunteers. In a screening approach, we tentatively identified six urinary OC metabolites. For three, renal elimination kinetics was quantitatively investigated using authentic standards: the sidechain oxidation product 2-ethyl-5-hydroxyhexyl 2-cyano-3,3-diphenyl acrylate (5OH-OC), the beta-oxidation product 2-(carboxymethyl)butyl 2-cyano-3,3-diphenyl acrylate (dinor OC carboxylic acid; DOCCA), and the ester hydrolysis product 2-cyano-3,3-diphenylacrylic acid (CPAA). CPAA was the major urinary metabolite, representing 45% (range 40-50%) of the OC dose. 5OH-OC and DOCCA were only minor metabolites with low, but highly consistent renal conversion factors of 0.008% (0.005-0.011%) and 0.13% (0.11-0.16%), respectively. Peak urinary metabolite concentrations were observed between 3.2 h and 4.2 h postdose. All three metabolites were excreted with biphasic elimination kinetics, with considerably longer elimination half-lives for DOCCA (1st phase: 3.0 h; 2nd phase: 16 h) and CPAA (5.7 h and 16 h) compared to 5OH-OC (1.3 h and 6.4 h). 99% of all 5OH-OC was excreted within 24 h compared to 82% of DOCCA and 77% of CPAA. After dermal exposure, we detected the same metabolites with similar ratios in urine, however, at much lower concentrations and with considerably delayed elimination.

Risk assessment for migration of styrene oligomers into food from polystyrene food containers.

Regulation EU 10/2011 requires a risk assessment of Non Intentionally Added Substances (NIAS) migrating into food for food contact plastics within the EU. Styrene oligomers are important potential components of NIAS in polystyrene used for food packaging and so far only dimers and trimers have been identified. They are not genotoxic in vitro, and there is good evidence that they are not endocrine disruptors. Hazard characterization to establish "safe" exposure levels is based on 1. The No Adverse Effect Level (NOAEL) of 1 mg/kg bw/d in an oral rat study during pregnancy and lactation and 2. The concept of Threshold of Toxicological Concern (TTC). Likely human exposure is derived from 1. the concentrations of dimers and trimers in food simulants or 2. in food and 3. the probabilistic FACET exposure estimation based on dimer and trimer concentrations in polystyrene and their potential for migration. The Margin of Safety as the relation of potential consumer exposure and the "safe" exposure level was always above 1 (apart from migration with 95% ethanol which is no longer recommended as an official food simulant for overall migration into fatty food) demonstrating that dimers and trimers in PS food packaging present a low risk for consumers.

Oligomers of styrene are not endocrine disruptors.

Oligomers of styrene have been identified in polystyrene (PS) polymer samples intended for food packaging. Such oligomers contribute to nonintentionally added substances (NIAS) that may migrate into food or food simulants and therefore have to be assessed for the potential risk to health. No oligomers larger than dimers and trimers of styrene have been found to be present in PS. Some in vivo and in vitro information indicative of an endocrine activity for some specific oligomers suggest concerns for their potential for endocrine disruption in humans. Data on endocrine activity available from in vitro and in vivo screening approaches and from non-guideline studies in experimental animals were evaluated. The different test methods were classified according to the OECD Conceptual Framework for Testing and Assessment of Endocrine Disruptors (OECD) and the ranking system of Borgert et al. proposed in 2014. The quality and reliability of each study is further assessed by professional judgment. The integration of the total information supports the conclusion that neither specific oligomers, nor their mixtures, potentially migrating into food are endocrine disruptors according to the definition of EFSA and WHO/IPCS.

Hexamoll® DINCH: Lack of in vivo evidence for obesogenic properties.

Hexamoll® DINCH is an important alternative to phthalate plasticizers. Although regulatory reviews have not identified any potential hazards even in sensitive populations, an in vitro study by Campioli et al. (2015) suggested Hexamoll® DINCH might alter fat storage in adipocytes resulting in obesity. To evaluate this hypothesis, data from studies with Hexamoll® DINCH were reviewed for evidence of deposition in fat, changes in body weight, or changes in serum chemistry reflecting altered metabolic status. Body weights of F1 and F2 pups in a two-generation study did not differ from controls even at 1000 mg Hexamoll® DINCH/kg body weight. Mean relative liver weights from the 1000 and 300 mg/kg bw groups were increased, but without histopathologic changes. Triglyceride and cholesterol levels in serum were not affected. In addition, subchronic and chronic studies in rats did not give evidence of an obesogenic effect. Radioactivity from 20 or 1000 mg/kg bw 14C-labelled Hexamoll® DINCH dosed orally remained 2-3 times longer in adipose tissue than in well-perfused tissues; however, levels were 20-500% below other tissues at 1 and 8 h post dosing. Radioactivity concentrations in organs and tissues excluding the GI tract declined rapidly and continuously, and decreased in parallel to the concentration in plasma during the following 20 h. Both, initial and terminal half-lives of radioactivity concentration do not indicate a potential for accumulation. Furthermore, a metabolomic comparison of Hexamoll® DINCH with DEHP and other phthalates shows complete separation of the metabolomic profile of these two chemical classes, meaning that their effects on the body and the body's reaction to the substance are different. Hence, comprehensive in vivo data do not show any evidence of Hexamoll® DINCH altering fat metabolism or having obesogenic properties.

Human metabolism and excretion kinetics of the fragrance 7-hydroxycitronellal after a single oral or dermal dosage.

7-Hydroxy-3,7-dimethyl-1-octanal, also known as 7-hydroxycitronellal (7-HC, CAS No. 107-75-5) is a synthetic fragrance widely used in cosmetic and hygiene products. Due to its large scope, 7-HC was selected for the development of a biomonitoring method suitable for the general population within the frame of the cooperation project between the German Federal Ministry for the Environment (BMUB) and the German Chemical Industry Association (VCI). In a human study with 5 healthy subjects who received single dermal and oral doses 7-HC, suitable metabolites and their urinary excretion kinetics was investigated. Two metabolites of 7-hydroxycitronellal were identified in urinary fractions after dermal and oral dosing: The alcohol 7-hydroxycitronellol (7-HCO) and the corresponding acid 7-hydroxycitronellylic acid (7-HCA). Only 7-HCA proved to be a suitable biomarker of exposure to 7-HC, since 7-HCO was quantifiable in only a minority of urine samples collected from the general population. Quantification of 7-HCA was conducted by means of a newly developed UPLC-MS/MS (ultra-high pressure liquid chromatography combined with tandem mass spectrometry) method. Peak excretion of 7-HCA occurred between 3 and 5h after oral application and about 10h after dermal administration. Due to the limited skin absorption of 7-HC, 7-HCA concentrations after dermal application were much lower than levels after oral application. After 24h, about 9% and 50% of the dermally and orally applied dose, respectively, were excreted as 7- HCA. With the conversion factors derived from the controlled human study, we estimated median exposure doses in a group of 40 human volunteers from the general population of approximately 93µg 7-HC per day. In conclusion, the 7-HC metabolite 7-HCA in urine is a suitable biomarker of exposure and can be applied for biomonitoring of the general population.