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).

Development and Validation of a DLLME-GC-MS-MS Method for the Determination of Benzotriazole UV Stabilizer UV-327 and Its Metabolites in Human Blood.

2-(5-Chloro-benzotriazol-2-yl)-4,6-di-(tert-butyl)phenol (UV-327) is used as an ultraviolet (UV) absorber in plastic materials and coatings. To investigate its metabolism and to assess human exposure, analytical methods are necessary for the determination of UV-327 and its metabolites in human biological specimens. The method thus presented targets the determination of UV-327 and several of its predicted metabolites in blood using protein precipitation, dispersive liquid-liquid microextraction (DLLME) and derivatization. The trimethylsilylated analytes and internal standards are separated by gas chromatography and analyzed with tandem mass spectrometry. The DLLME procedure was optimized with respect to the type and volume of disperser and extraction solvents, the pH value of the sample solution and the addition of salt. During method development, an effective ex vivo lactone/hydroxyl carboxylic acid interconversion was observed for two metabolites, each containing a carboxyl group adjacent to the phenolic hydroxyl group. The analytes resulting from interconversion enabled a more sensitive and reliable determination of the metabolites compared to their native structures. Method validation revealed limits of detection between 0.02 and 0.36 µg/L. The mean relative recovery rates ranged from 91% to 118%. Precision and repeatability were demonstrated by relative standard deviations in the range of 0.6-14.2% and 1.1-13.7%, respectively. The presently described procedure enables the sensitive and robust analysis of UV-327 and its metabolites in human blood and allows the elucidation of the human UV-327 metabolism as well as the assessment of exposure in potentially exposed individuals.

An agenda for future Social Sciences and Humanities research on energy efficiency: 100 priority research questions.

Decades of techno-economic energy policymaking and research have meant evidence from the Social Sciences and Humanities (SSH)-including critical reflections on what changing a society's relation to energy (efficiency) even means-have been underutilised. In particular, (i) the SSH have too often been sidelined and/or narrowly pigeonholed by policymakers, funders, and other decision-makers when driving research agendas, and (ii) the setting of SSH-focused research agendas has not historically embedded inclusive and deliberative processes. The aim of this paper is to address these gaps through the production of a research agenda outlining future SSH research priorities for energy efficiency. A Horizon Scanning exercise was run, which sought to identify 100 priority SSH questions for energy efficiency research. This exercise included 152 researchers with prior SSH expertise on energy efficiency, who together spanned 62 (sub-)disciplines of SSH, 23 countries, and a full range of career stages. The resultant questions were inductively clustered into seven themes as follows: (1) Citizenship, engagement and knowledge exchange in relation to energy efficiency; (2) Energy efficiency in relation to equity, justice, poverty and vulnerability; (3) Energy efficiency in relation to everyday life and practices of energy consumption and production; (4) Framing, defining and measuring energy efficiency; (5) Governance, policy and political issues around energy efficiency; (6) Roles of economic systems, supply chains and financial mechanisms in improving energy efficiency; and (7) The interactions, unintended consequences and rebound effects of energy efficiency interventions. Given the consistent centrality of energy efficiency in policy programmes, this paper highlights that well-developed SSH approaches are ready to be mobilised to contribute to the development, and/or to understand the implications, of energy efficiency measures and governance solutions. Implicitly, it also emphasises the heterogeneity of SSH policy evidence that can be produced. The agenda will be of use for both (1) those new to the energy-SSH field (including policyworkers), for learnings on the capabilities and capacities of energy-SSH, and (2) established energy-SSH researchers, for insights on the collectively held futures of energy-SSH research.

ATF2 loss promotes tumor invasion in colorectal cancer cells via upregulation of cancer driver TROP2.

In cancer, the activating transcription factor 2 (ATF2) has pleiotropic functions in cellular responses to growth stimuli, damage, or inflammation. Due to only limited studies, the significance of ATF2 in colorectal cancer (CRC) is not well understood. We report that low ATF2 levels correlated with worse prognosis and tumor aggressiveness in CRC patients. NanoString gene expression and ChIP analysis confirmed trophoblast cell surface antigen 2 (TROP2) as a novel inhibitory ATF2 target gene. This inverse correlation was further observed in primary human tumor tissues. Immunostainings revealed that high intratumoral heterogeneity for ATF2 and TROP2 expression was sustained also in liver metastasis. Mechanistically, our in vitro data of CRISPR/Cas9-generated ATF2 knockout (KO) clones revealed that high TROP2 levels were critical for cell de-adhesion and increased cell migration without triggering EMT. TROP2 was enriched in filopodia and displaced Paxillin from adherens junctions. In vivo imaging, micro-computer tomography, and immunostainings verified that an ATF2KO/TROP2high status triggered tumor invasiveness in in vivo mouse and chicken xenograft models. In silico analysis provided direct support that ATF2low/TROP2high expression status defined high-risk CRC patients. Finally, our data demonstrate that ATF2 acts as a tumor suppressor by inhibiting the cancer driver TROP2. Therapeutic TROP2 targeting might prevent particularly the first steps in metastasis, i.e., the de-adhesion and invasion of colon cancer cells.

Determination of UV-327 and its metabolites in human urine using dispersive liquid-liquid microextraction and gas chromatography-tandem mass spectrometry.

The benzotriazole UV stabilizer (BUVS) 2-(5-chloro-benzotriazol-2-yl)-4,6-di-(tert-butyl)phenol (UV-327) is added to plastic materials for UV protection. The compound is known to be ubiquitously distributed in the environment. We developed the first analytical method for the determination of UV-327 and seven metabolites, which were identified in vitro, in urine to be able to investigate the in vivo metabolism of UV-327 and to assess potential human exposure to the compound. Enzymatic hydrolysis of phase II conjugates is followed by sample purification with dispersive liquid-liquid microextraction (DLLME). The analytes are extracted from the urine samples after acidification with hydrochloric acid solution and addition of sodium chloride solution. Isopropyl alcohol and chloroform are used as disperser solvent and extraction solvent, respectively. After derivatization, the trimethylsilylated analytes are chromatographically separated and detected by gas chromatography coupled with tandem mass spectrometry (GC-MS/MS). To achieve maximum extraction of the analytes from the sample solution, the DLLME procedure was optimized with respect to the type and volume of disperser and extraction solvent, the pH value of the sample solution, the addition of salt, and the duration of vortex-mixing. Subsequent method validation demonstrated high sensitivity and reliability, with limits of detection (LODs) between 0.05 and 0.1 µg l-1 and mean relative recovery rates ranging from 88 to 112%. Precision and repeatability were proven by relative standard deviations ranging from 1 to 13% and from 5 to 14%, respectively.

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.