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Per- and polyfluoroalkyl substances (PFASs) have been detected in an array of environmental media due to their ubiquitous use in industrial and consumer products as well as potential release from fluorochemical manufacturing facilities. During their manufacture, many fluorotelomer (FT) facilities rely on neutral intermediates in polymer production including the FT-alcohols (FTOHs). These PFAS are known to transform to the terminal acids (perfluoro carboxylic acids; PFCAs) at rates that vary with environmental conditions. In the current study on soils from a FT facility, we employed gas chromatography coupled with conventional- and high-resolution mass spectrometry (GC-MS and GC-HRMS) to investigate the profile of these precursor compounds, the intermediary secondary alcohols (sFTOHs), FT-acrylates (FTAcr), and FT-acetates (FTAce) in soils around the former FT-production facility. Of these precursors, the general trend in detection intensity was [FTOHs] > [sFTOHs] > [FTAcrs], while for the FTOHs, homologue intensities generally were [12:2 FTOH] > [14:2 FTOH] > [16:2 FTOH] > [10:2 FTOH] > [18:2 FTOH] > [20:2 FTOH] > [8:2 FTOH] â¼ [6:2 FTOH]. The corresponding terminal acids were also detected in all soil samples and positively correlated with the precursor concentrations. GC-HRMS confirmed the presence of industrial manufacturing byproducts such as FT-ethers and FT-esters and aided in the tentative identification of previously unreported dimers and other compounds. The application of GC-HRMS to the measurement and identification of precursor PFAS is in its infancy, but the methodologies described here will help refine its use in tentatively identifying these compounds in the environment.
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Fluorocarburos , Contaminantes del Suelo , Suelo , Contaminantes del Suelo/análisis , Suelo/química , Fluorocarburos/análisis , Cromatografía de Gases y Espectrometría de Masas , Monitoreo del Ambiente , Instalaciones Industriales y de FabricaciónRESUMEN
Per- and polyfluoroalkyl substances (PFAS) are a unique class of chemicals synthesized to aid in industrial processes, fire-fighting products, and to benefit consumer products such as clothing, cosmetics, textiles, carpets, and coatings. The widespread use of PFAS and their strong carbon-fluorine bonds has led to their ubiquitous presence throughout the world. Airborne transport of PFAS throughout the atmosphere has also contributed to environmental pollution. Due to the potential environmental and human exposure concerns of some PFAS, research has extensively focused on water, soil, and organismal detection, but the presence of PFAS in the air has become an area of growing concern. Methods to measure polar PFAS in various matrices have been established, while the investigation of polar and nonpolar PFAS in air is still in its early development. This literature review aims to present the last two decades of research characterizing PFAS in outdoor and indoor air, focusing on active and passive air sampling and analytical methods. The PFAS classes targeted and detected in air samples include fluorotelomer alcohols (FTOHs), perfluoroalkane sulfonamides (FASAs), perfluoroalkane sulfonamido ethanols (FASEs), perfluorinated carboxylic acids (PFCAs), and perfluorinated sulfonic acids (PFSAs). Although the manufacturing of perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid (PFOS) has been largely phased out, these two PFAS are still often detected in air samples. Additionally, recent estimates indicate that there are thousands of PFAS that are likely present in the air that are not currently monitored in air methods. Advances in air sampling methods are needed to fully characterize the atmospheric transport of PFAS.
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Contaminantes Atmosféricos , Contaminación del Aire Interior , Monitoreo del Ambiente , Fluorocarburos , Fluorocarburos/análisis , Monitoreo del Ambiente/métodos , Contaminantes Atmosféricos/análisis , Contaminación del Aire Interior/análisis , HumanosRESUMEN
The scientific foundation for per- and polyfluoroalkyl substances (PFAS) measurements in water, soils, sediments, biosolids, biota, and outdoor air has rapidly expanded; however, there are limited efforts devoted to developing analytical methods to measure vapor-phase PFAS in indoor air. A gas chromatography-tandem mass spectrometry (GC-MS/MS) method coupled with thermal desorption (TD) sorbent tube analysis was developed to quantify trace levels of fluorotelomer alcohols (FTOHs) emitted from consumer products in the indoor environment. Method evaluation included determination of instrument detection limits (IDLs), quality assurance checks of target standards purchased from different vendors, sample loss during storage, and TD sorbent breakthrough with tubes coupled in-series. The IDLs for TD-GC-MS/MS analyses ranged from 0.07 - 0.09 ng/tube. No significant loss of FTOHs was observed during stability tests over 28 days with relative standard deviations (RSDs) of spiked TD tubes ranging from 3.1 - 7.7% and the RSDs of polypropylene copolymer vial storage of standard solutions ranging from 4.3 - 8.4%. TD tube breakthrough was minimal with recovered FTOHs in the second tubes <1% of the spiked concentrations in the first tubes with carrier gas volume up to 20 L. The method has been applied to determine FTOH emissions from three consumer products in micro-scale chambers. A liquid stone cleaner/sealer product contained the highest levels of 6:2, 8:2, and 10:2 FTOHs, while the mattress pad products contained lower levels of 8:2 and 10:2 FTOHs. The emission parameters, including the initial emission factors and first order decay rate constants, were obtained based on the experimental data. The developed methods are sensitive and specific for analysis of all four target FTOHs (4:2, 6:2, 8:2, 10:2 FTOHs) with chamber testing. The methods can be extended to indoor air sampling and could be applicable to ambient air sampling.
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Fluorocarburos , Espectrometría de Masas en Tándem , Espectrometría de Masas en Tándem/métodos , Cromatografía de Gases y Espectrometría de Masas/métodos , Fluorocarburos/análisis , Aire/análisisRESUMEN
New approach methodologies (NAMs) that make use of in vitro screening and in silico approaches to inform chemical evaluations rely on in vitro toxicokinetic (TK) data to translate in vitro bioactive concentrations to exposure metrics reflective of administered dose. With 1364 per- and polyfluoroalkyl substances (PFAS) identified as of interest under Section 8 of the U.S. Toxic Substances Control Act (TSCA) and concern over the lack of knowledge regarding environmental persistence, human health, and ecological effects, the utility of NAMs to understand potential toxicities and toxicokinetics across these data-poor compounds is being evaluated. To address the TK data deficiency, 71 PFAS selected to span a wide range of functional groups and physico-chemical properties were evaluated for in vitro human plasma protein binding (PPB) by ultracentrifugation with liquid chromatography-mass spectrometry analysis. For the 67 PFAS successfully evaluated by ultracentrifugation, fraction unbound in plasma (fup) ranged from less than 0.0001 (pentadecafluorooctanoyl chloride) to 0.7302 (tetrafluorosuccinic acid), with over half of the PFAS showing PPB exceeding 99.5% (fup < 0.005). Category-based evaluations revealed that perfluoroalkanoyl chlorides and perfluorinated carboxylates (PFCAs) with 6-10 carbons were the highest bound, with similar median values for alkyl, ether, and polyether PFCAs. Interestingly, binding was lower for the PFCAs with a carbon chain length of ≥11. Lower binding also was noted for fluorotelomer carboxylic acids when compared to their carbon-equivalent perfluoroalkyl acids. Comparisons of the fup value derived using two PPB methods, ultracentrifugation or rapid equilibrium dialysis (RED), revealed RED failure for a subset of PFAS of high mass and/or predicted octanol-water partition coefficients exceeding 4 due to failure to achieve equilibrium. Bayesian modeling was used to provide uncertainty bounds around fup point estimates for incorporation into TK modeling. This PFAS PPB evaluation and grouping exercise across 67 structures greatly expand our current knowledge and will aid in PFAS NAM development.
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Fluorocarburos , Unión Proteica , Toxicocinética , Contaminantes Químicos del Agua , Humanos , Teorema de Bayes , Proteínas Sanguíneas , Ácidos Carboxílicos/toxicidad , Ácidos Carboxílicos/análisis , Fluorocarburos/química , Unión Proteica/efectos de los fármacos , Contaminantes Químicos del Agua/análisisRESUMEN
Concern over per- and polyfluoroalkyl substances (PFAS) has increased as more is learned about their environmental presence, persistence, and bioaccumulative potential. The limited monitoring, toxicokinetic (TK), and toxicologic data available are inadequate to inform risk across this diverse domain. Here, 73 PFAS were selected for in vitro TK evaluation to expand knowledge across lesser-studied PFAS alcohols, amides, and acrylates. Targeted methods developed using gas chromatography-tandem mass spectrometry (GC-MS/MS) were used to measure human plasma protein binding and hepatocyte clearance. Forty-three PFAS were successfully evaluated in plasma, with fraction unbound (fup) values ranging from 0.004 to 1. With a median fup of 0.09 (i.e., 91% bound), these PFAS are highly bound but exhibit 10-fold lower binding than legacy perfluoroalkyl acids recently evaluated. Thirty PFAS evaluated in the hepatocyte clearance assay showed abiotic loss, with many exceeding 60% loss within 60 min. Metabolic clearance was noted for 11 of the 13 that were successfully evaluated, with rates up to 49.9 µL/(min × million cells). The chemical transformation simulator revealed potential (bio)transformation products to consider. This effort provides critical information to evaluate PFAS for which volatility, metabolism, and other routes of transformation are likely to modulate their environmental fates.
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Per- and polyfluoroalkyl substances (PFAS) are a diverse set of commercial chemicals widely detected in humans and the environment. However, only a limited number of PFAS are associated with epidemiological or experimental data for hazard identification. To provide developmental neurotoxicity (DNT) hazard information, the work herein employed DNT new approach methods (NAMs) to generate in vitro screening data for a set of 160 PFAS. The DNT NAMs battery was comprised of the microelectrode array neuronal network formation assay (NFA) and high-content imaging (HCI) assays to evaluate proliferation, apoptosis, and neurite outgrowth. The majority of PFAS (118/160) were inactive or equivocal in the DNT NAMs, leaving 42 active PFAS that decreased measures of neural network connectivity and neurite length. Analytical quality control indicated 43/118 inactive PFAS samples and 10/42 active PFAS samples were degraded; as such, careful interpretation is required as some negatives may have been due to loss of the parent PFAS, and some actives may have resulted from a mixture of parent and/or degradants of PFAS. PFAS containing a perfluorinated carbon (C) chain length ≥8, a high C:fluorine ratio, or a carboxylic acid moiety were more likely to be bioactive in the DNT NAMs. Of the PFAS positives in DNT NAMs, 85% were also active in other EPA ToxCast assays, whereas 79% of PFAS inactives in the DNT NAMs were active in other assays. These data demonstrate that a subset of PFAS perturb neurodevelopmental processes in vitro and suggest focusing future studies of DNT on PFAS with certain structural feature descriptors.
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Fluorocarburos , Síndromes de Neurotoxicidad , Humanos , Síndromes de Neurotoxicidad/metabolismo , Neuronas/metabolismo , Proyección Neuronal , Apoptosis , Fluorocarburos/toxicidadRESUMEN
Per- and polyfluoroalkyl substances (PFAS) represent a large chemical class lacking hazard, toxicokinetic, and exposure information. To accelerate PFAS hazard evaluation, new approach methodologies (NAMs) comprised of in vitro high-throughput toxicity screening, toxicokinetic data, and computational modeling are being employed in read across strategies to evaluate the larger PFAS landscape. A critical consideration to ensure robust evaluations is a parallel assessment of the quality of the screening stock solutions, where dimethyl sulfoxide (DMSO) is often the diluent of choice. Challenged by the lack of commercially available reference standards for many of the selected PFAS and reliance on mass spectrometry approaches for such an evaluation, we developed a high-throughput framework to evaluate the quality of screening stocks for 205 PFAS selected for these NAM efforts. Using mass spectrometry coupled with either liquid or gas chromatography, a quality scoring system was developed that incorporated observations during mass spectral examination to provide a simple pass or fail notation. Informational flags were used to further describe findings regarding parent analyte presence through accurate mass identification, evidence of contaminants and/or degradation, or further describe characteristics such as isomer presence. Across the PFAS-DMSO stocks tested, 148 unique PFAS received passing quality scores to allow for further in vitro testing whereas 57 received a failing score primarily due to detection issues or confounding effects of DMSO. Principle component analysis indicated vapor pressure and Henry's Law Constant as top indicators for a failed quality score for those analyzed by gas chromatography. Three PFAS in the hexafluoropropylene oxide family failed due to degradation in DMSO. As the PFAS evaluated spanned over 20 different structural categories, additional commentary describes analytical observations across specific groups related to PFAS stock composition, detection, stability, and methodologic considerations that will be useful for informing future analytical assessment and downstream HTS efforts. The high-throughput stock quality scoring workflow presented holds value as a tool to evaluate chemical presence and quality efficiently and for informing data inclusion in PFAS or other NAM screening efforts.
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Dimetilsulfóxido , Fluorocarburos , Ensayos Analíticos de Alto Rendimiento , Simulación por Computador , Excipientes , Fluorocarburos/toxicidadRESUMEN
A structurally diverse set of 147 per- and polyfluoroalkyl substances (PFAS) was screened in a panel of 12 human primary cell systems by measuring 148 biomarkers relevant to (patho)physiological pathways to inform hypotheses about potential mechanistic effects of data-poor PFAS in human model systems. This analysis focused on immunosuppressive activity, which was previously reported as an in vivo effect of perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS), by comparing PFAS responses to four pharmacological immunosuppressants. The PFOS response profile had little correlation with reference immunosuppressants, suggesting in vivo activity does not occur by similar mechanisms. The PFOA response profile did share features with the profile of dexamethasone, although some distinct features were lacking. Other PFAS, including 2,2,3,3-tetrafluoropropyl acrylate, demonstrated more similarity to the reference immunosuppressants but with additional activities not found in the reference immunosuppressive drugs. Correlation of PFAS profiles with a database of environmental chemical responses and pharmacological probes identified potential mechanisms of bioactivity for some PFAS, including responses similar to ubiquitin ligase inhibitors, deubiquitylating enzyme (DUB) inhibitors, and thioredoxin reductase inhibitors. Approximately 21% of the 147 PFAS with confirmed sample quality were bioactive at nominal testing concentrations in the 1-60 micromolar range in these human primary cell systems. These data provide new hypotheses for mechanisms of action for a subset of PFAS and may further aid in development of a PFAS categorization strategy useful in safety assessment.
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Ácidos Alcanesulfónicos , Contaminantes Ambientales , Fluorocarburos , Humanos , Ácidos Alcanesulfónicos/toxicidad , Caprilatos , Fluorocarburos/toxicidad , Fluorocarburos/análisisRESUMEN
The destruction of per- and polyfluoroalkyl substances (PFAS) is critical to ensure effective remediation of PFAS contaminated matrices. The destruction of hazardous chemicals within incinerators and other thermal treatment processes has historically been determined by calculating the destruction efficiency (DE) or the destruction and removal efficiency (DRE). While high DEs, >99.99%, are deemed acceptable for most hazardous compounds, many PFAS can be converted to other PFAS at low temperatures resulting in high DEs without full mineralization and the potential release of the remaining fluorocarbon portions to the environment. Many of these products of incomplete combustion (PICs) are greenhouse gases, most have unknown toxicity, and some can react to create new perfluorocarboxylic acids. Experiments using aqueous film forming foam (AFFF) and a pilot-scale research combustor varied the combustion environment to determine if DEs indicate PFAS mineralization. Several operating conditions above 1090 °C resulted in high DEs and few detectable fluorinated PIC emissions. However, several conditions below 1000 °C produced DEs >99.99% for the quantifiable PFAS and mg/m3 emission concentrations of several non-polar PFAS PICs. These results suggest that DE alone may not be the best indication of total PFAS destruction, and additional PIC characterization may be warranted.
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High-throughput (HT) in vitro to in vivo extrapolation (IVIVE) is an integral component in new approach method (NAM)-based risk assessment paradigms, for rapidly translating in vitro toxicity assay results into the context of in vivo exposure. When coupled with rapid exposure predictions, HT-IVIVE supports the use of HT in vitro assays for risk-based chemical prioritization. However, the reliability of prioritization based on HT bioactivity data and HT-IVIVE can be limited as the domain of applicability of current HT-IVIVE is generally restricted to intrinsic clearance measured primarily in pharmaceutical compounds. Further, current approaches only consider parent chemical toxicity. These limitations occur because current state-of-the-art HT prediction tools for clearance and metabolite kinetics do not provide reliable data to support HT-IVIVE. This paper discusses current challenges in implementation of IVIVE for prioritization and risk assessment and recommends a path forward for addressing the most pressing needs and expanding the utility of IVIVE.
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Per- and polyfluoroalkyl substances (PFAS) are a broad class of hundreds of fluorinated chemicals with environmental health concerns due to their widespread presence and persistence in the environment. Several of these chemicals have been comprehensively studied for experimental toxicity, environmental fate and exposure, and human epidemiology; however, most chemicals have limited or no data available. To inform methods for prioritizing these data-poor chemicals for detailed toxicity studies, we evaluated 142 PFAS using an in vitro screening platform consisting of two multiplexed transactivation assays encompassing 81 diverse transcription factor activities and tested in concentration-response format ranging from 137â¯nM to 300 µM. Results showed activity for various nuclear receptors, including three known PFAS targets--specifically estrogen receptor alpha and peroxisome proliferator receptors alpha and gamma. We also report activity against the retinoid X receptor beta, the key heterodimeric partner of type II, non-steroidal nuclear receptors. Additional activities were found against the pregnane X receptor, nuclear receptor related-1 protein, and nuclear factor erythroid 2-related factor 2, a sensor of oxidative stress. Using orthogonal assay approaches, we confirmed activity of representative PFAS against several of these targets. Finally, we identified key PFAS structural features associated with nuclear receptor activity that can inform future predictive models for use in prioritizing chemicals for risk assessment and in the design of new structures devoid of biological activity.
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Proliferación Celular/efectos de los fármacos , Fluorocarburos/química , Fluorocarburos/toxicidad , Transducción de Señal/efectos de los fármacos , Proliferación Celular/fisiología , Fluorocarburos/metabolismo , Células Hep G2 , Humanos , Estructura Molecular , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/fisiologíaRESUMEN
Glutathione transferase zeta 1 (GSTZ1), expressed in liver and several extrahepatic tissues, catalyzes dechlorination of dichloroacetate (DCA) to glyoxylate. DCA inactivates GSTZ1, leading to autoinhibition of its metabolism. DCA is an investigational drug for treating several congenital and acquired disorders of mitochondrial energy metabolism, including cancer. The main adverse effect of DCA, reversible peripheral neuropathy, is more common in adults treated long-term than in children, who metabolize DCA more quickly after multiple doses. One dose of DCA to Sprague Dawley rats reduced GSTZ1 expression and activity more in liver than in extrahepatic tissues; however, the effects of multiple doses of DCA that mimic its therapeutic use have not been studied. Here, we examined the expression and activity of GSTZ1 in cytosol and mitochondria of liver, kidney, heart, and brain 24 hours after completion of 8-day oral dosing of 100 mg/kg per day sodium DCA to juvenile and adult Sprague Dawley rats. Activity was measured with DCA and with 1,2-epoxy-3-(4-nitrophenoxy)propane (EPNPP), reported to be a GSTZ1-selective substrate. In DCA-treated rats, liver retained higher expression and activity of GSTZ1 with DCA than other tissues, irrespective of rodent age. DCA-treated juvenile rats retained more GSTZ1 activity with DCA than adults. Consistent with this finding, there was less measurable DCA in tissues of juvenile than adult rats. DCA-treated rats retained activity with EPNPP, despite losing over 98% of GSTZ1 protein. These data provide insight into the differences between children and adults in DCA elimination under a therapeutic regimen and confirm that the liver contributes more to DCA metabolism than other tissues. SIGNIFICANCE STATEMENT: Dichloroacetate (DCA) is one of few drugs exhibiting higher clearance from children than adults, after repeated doses, for reasons that are unclear. We hypothesized that juveniles retain more glutathione transferase zeta 1 (GSTZ1) than adults in tissues after multiple DCA doses and found this was the case for liver and kidney, with rat as a model to assess GSTZ1 protein expression and activity with DCA. Although 1,2-epoxy-3-(4-nitrophenoxy)propane was reported to be a selective GSTZ1 substrate, its activity was not reduced in concert with GSTZ1 protein.
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Ácido Dicloroacético/farmacocinética , Glutatión Transferasa/antagonistas & inhibidores , Hígado/efectos de los fármacos , Adulto , Factores de Edad , Animales , Niño , Ácido Dicloroacético/administración & dosificación , Relación Dosis-Respuesta a Droga , Metabolismo Energético/efectos de los fármacos , Compuestos Epoxi/farmacocinética , Femenino , Glutatión Transferasa/metabolismo , Humanos , Hígado/metabolismo , Masculino , Enfermedades Mitocondriales/tratamiento farmacológico , Modelos Animales , Nitrofenoles/farmacocinética , RatasRESUMEN
Chemical risk assessment relies on toxicity tests that require significant numbers of animals, time and costs. For the >30,000 chemicals in commerce, the current scale of animal testing is insufficient to address chemical safety concerns as regulatory and product stewardship considerations evolve to require more comprehensive understanding of potential biological effects, conditions of use, and associated exposures. We demonstrate the use of a multi-level new approach methodology (NAMs) strategy for hazard- and risk-based prioritization to reduce animal testing. A Level 1/2 chemical prioritization based on estrogen receptor (ER) activity and metabolic activation using ToxCast data was used to select 112 chemicals for testing in a Level 3 human uterine cell estrogen response assay (IKA assay). The Level 3 data were coupled with quantitative in vitro to in vivo extrapolation (Q-IVIVE) to support bioactivity determination (as a surrogate for hazard) in a tissue-specific context. Assay AC50s and Q-IVIVE were used to estimate human equivalent doses (HEDs), and HEDs were compared to rodent uterotrophic assay in vivo-derived points of departure (PODs). For substances active both in vitro and in vivo, IKA assay-derived HEDs were lower or equivalent to in vivo PODs for 19/23 compounds (83%). Activity exposure relationships were calculated, and the IKA assay was as or more protective of human health than the rodent uterotrophic assay for all IKA-positive compounds. This study demonstrates the utility of biologically relevant fit-for-purpose assays and supports the use of a multi-level strategy for chemical risk assessment.
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Alternativas al Uso de Animales/métodos , Disruptores Endocrinos/toxicidad , Ensayos Analíticos de Alto Rendimiento/métodos , Pruebas de Toxicidad/métodos , Útero/efectos de los fármacos , Animales , Bioensayo/métodos , Técnicas de Cultivo de Célula , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Simulación por Computador , Estudios de Factibilidad , Femenino , Humanos , Modelos Biológicos , Ratas , Medición de Riesgo/métodos , Útero/citologíaRESUMEN
Dichloroacetate (DCA) has potential for treating mitochondrial disorders and cancer by activating the mitochondrial pyruvate dehydrogenase complex. Repeated dosing of DCA results in reduced drug clearance due to inactivation of glutathione transferase ζ1 (GSTZ1), its metabolizing enzyme. We investigated the time-course of inactivation of GSTZ1 in hepatic cytosol and mitochondria after one oral dose of 100 mg/kg DCA to female Sprague-Dawley rats aged 4 weeks (young) and 52 weeks (adult) as models for children and adults, respectively. GSTZ1 activity with both DCA and an endogenous substrate, maleylacetone (MA), as well as GSTZ1 protein expression were rapidly reduced in cytosol from both ages following DCA treatment. In mitochondria, loss of GSTZ1 protein and activity with DCA were even more rapid. The cytosolic in vivo half-lives of the loss of GSTZ1 activity with DCA were 1.05 ± 0.03 and 0.82 ± 0.02 h (mean ± S.D., n = 6) for young and adult rats, respectively, with inactivation significantly more rapid in adult rats, p < 0.001. The mitochondrial inactivation half-lives were similar in young (0.57 ± 0.02 h) and adult rats (0.54 ± 0.02 h) and were significantly (p < 0.0001) shorter than cytosolic inactivation half-lives. By 24 h after DCA administration, activity and expression remained at 10% or less than control values. The in vitro GSTZ1 inactivation half-lives following incubation with 2 mM DCA in the presence of physiological chloride (Cl-) concentrations (cytosol = 44 mM, mitochondria = 1-2 mM) exhibited marked differences between subcellular fractions, being 3 times longer in the cytosol than in the mitochondria, regardless of age, suggesting that the lower Cl- concentration in mitochondria explained the faster degradation of GSTZ1. These results demonstrate for the first time that rat mitochondrial GSTZ1 is more readily inactivated by DCA than cytosolic GSTZ1, and cytosolic GSTZ1 is inactivated more rapidly in adult than young rats.
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Citosol/enzimología , Ácido Dicloroacético/farmacología , Ácido Dicloroacético/toxicidad , Glutatión Transferasa/antagonistas & inhibidores , Hígado/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Animales , Ácido Dicloroacético/administración & dosificación , Femenino , Glutatión Transferasa/metabolismo , Hígado/metabolismo , Mitocondrias/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Glutathione transferase zeta1 (GSTZ1) catalyzes glutathione (GSH)-dependent dechlorination of dichloroacetate (DCA), an investigational drug with therapeutic potential in metabolic disorders and cancer. GSTZ1 is expressed in both hepatic cytosol and mitochondria. Here, we examined the ontogeny and characterized the properties of human mitochondrial GSTZ1. GSTZ1 expression and activity with DCA were determined in 103 human hepatic mitochondrial samples prepared from livers of donors aged 1 day to 84 years. DNA from each sample was genotyped for three common GSTZ1 functional single nucleotide polymorphisms. Expression of mitochondrial GSTZ1 protein increased in an age-dependent manner to a plateau after age 21 years. Activity with DCA correlated with expression, after taking into account the somewhat higher activity of samples that were homo- or heterozygous for GSTZ1A. In samples from livers with the GSTZ1C variant, apparent enzyme kinetic constants for DCA and GSH were similar for mitochondria and cytosol after correcting for the loss of GSH observed in mitochondrial incubations. In the presence of 38 mM chloride, mitochondrial GSTZ1 exhibited shorter half-lives of inactivation compared with the cytosolic enzyme (P = 0.017). GSTZ1 protein isolated from mitochondria was shown by mass spectrometry to be identical to cytosolic GSTZ1 protein in the covered primary protein sequence. In summary, we report age-related development in the expression and activity of human hepatic mitochondrial GSTZ1 does not have the same pattern as that reported for cytosolic GSTZ1. Some properties of cytosolic and mitochondrial GSTZ1 differed, but these were not related to differences in amino acid sequence or post-translationally modified residues.
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Glutatión Transferasa/genética , Hígado/metabolismo , Mitocondrias/genética , Mitocondrias/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Secuencia de Aminoácidos , Niño , Preescolar , Citosol/metabolismo , Ácido Dicloroacético/metabolismo , Drogas en Investigación/metabolismo , Femenino , Glutatión Transferasa/metabolismo , Humanos , Lactante , Cinética , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple/genética , Adulto JovenRESUMEN
Biotransformation of dichloroacetate (DCA) to glyoxylate by hepatic glutathione transferase zeta 1 (GSTZ1) is considered the principal determinant of the rate of plasma clearance of the drug. However, several other organismal and subcellular factors are also known to influence DCA metabolism. We utilized a female rat model to study these poorly understood processes. Rats aged 4â¯weeks (young) and 42-52â¯weeks (adult) were used to model children and adults, respectively. Hepatic chloride concentrations, which influence the rate of GSTZ1 inactivation by DCA, were lower in rat than in human tissues and rats did not show the age dependence previously seen in humans. We found GSTZ1 expression and activity in rat brain, heart, and kidney cell-free homogenates that were age-dependent. GSTZ1 expression in brain was higher in young rats than adult rats, whereas cardiac and renal GSTZ1 expression levels were higher in adult than young rats. GSTZ1 activity with DCA could not be measured accurately in kidney cell-free homogenates due to rapid depletion of glutathione by γ-glutamyl transpeptidase. Following oral administration of DCA, 100â¯mg/kg, to rats, GSTZ1 expression and activity were reduced in all rat tissues, but chloride concentrations were not affected. Together, these data extend our understanding of factors that determine the in vivo kinetics of DCA.
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Cloruros/metabolismo , Ácido Dicloroacético/metabolismo , Glutatión Transferasa/metabolismo , Hígado/metabolismo , Animales , Encéfalo/metabolismo , Femenino , Regulación Enzimológica de la Expresión Génica , Glutatión , Glutatión Transferasa/genética , Riñón/metabolismo , Mitocondrias/metabolismo , Miocardio/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Following the 2010 Gulf of Mexico oil spill, concerns were raised regarding exposure of fish to crude oil components, particularly polycyclic aromatic hydrocarbons (PAHs). This three year study examined hepatic enzymes in post-mitochondrial supernatant fractions from red snapper (Lutjanus campechanus) and gray triggerfish (Balistes capriscus) collected in the north central Gulf of Mexico between 2011 and 2014. Biomarker activities evaluated included benzo(a)pyrene hydroxylase (AHH), ethoxyresorufin O-deethylase (EROD), glutathione transferase (GST), and glutathione peroxidase (GPx). Mean EROD activity was higher in gray triggerfish (12.97 ± 7.15 pmol/min/mg protein [mean ± SD], n = 115) than red snapper (2.75 ± 1.92 pmol/min/mg protein, n = 194), p < 0.0001. In both species, EROD declined over time between 2011 and 2014. Declines in GST and GPx activities were also noted over this time period for both species. Gray triggerfish liver was fatty, and heptane extracts of the liver fat contained fluorescent substances with properties similar to known PAHs, however the origin of these PAHs is unknown.
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Monitoreo del Ambiente , Hígado/metabolismo , Contaminación por Petróleo , Petróleo/toxicidad , Contaminantes Químicos del Agua/toxicidad , Animales , Biomarcadores/metabolismo , Citocromo P-450 CYP1A1/metabolismo , Peces , Glutatión Transferasa/metabolismo , Golfo de México , Hidrocarburos Policíclicos Aromáticos/toxicidadRESUMEN
Dichloroacetate (DCA) has several therapeutic applications based on its pharmacological property of inhibiting pyruvate dehydrogenase kinase. DCA has been used to treat inherited mitochondrial disorders that result in lactic acidosis, as well as pulmonary hypertension and several different solid tumors, the latter through its ability to reverse the Warburg effect in cancer cells and restore aerobic glycolysis. The main clinically limiting toxicity is reversible peripheral neuropathy. Although administration of high doses to rodents can result in liver cancer, there is no evidence that DCA is a human carcinogen. In all studied species, including humans, DCA has the interesting property of inhibiting its own metabolism upon repeat dosing, resulting in alteration of its pharmacokinetics. The first step in DCA metabolism is conversion to glyoxylate catalyzed by glutathione transferase zeta 1 (GSTZ1), for which DCA is a mechanism-based inactivator. The rate of GSTZ1 inactivation by DCA is influenced by age, GSTZ1 haplotype and cellular concentrations of chloride. The effect of DCA on its own metabolism complicates the selection of an effective dose with minimal side effects.
Asunto(s)
Ácido Dicloroacético/administración & dosificación , Glutatión Transferasa/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Ácido Dicloroacético/efectos adversos , Ácido Dicloroacético/farmacología , Relación Dosis-Respuesta a Droga , Humanos , Hipertensión Pulmonar/tratamiento farmacológico , Enfermedades Mitocondriales/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Piruvato Deshidrogenasa Quinasa Acetil-TransferidoraRESUMEN
Glutathione transferase zeta 1 (GSTZ1-1) is a homodimeric enzyme found in the cytosol and mitochondrial matrix of the liver and other tissues. It catalyzes the glutathione-dependent isomerization of maleylacetoacetate to fumarylacetoacetate in the tyrosine catabolic pathway and can metabolize small halogenated carboxylic acids. GSTZ1a-1a crystals diffracted to a resolution of 3.1 Å and belonged to space group P1, with unit-cell parameters a = 42.0, b = 49.6, c = 54.6 Å, α = 82.9, ß = 69.9, γ = 73.4°, with a calculated Matthews coefficient of 2.1 Å(3) Da(-1) assuming a dimer in the crystallographic asymmetric unit. Refinement of the structure is currently in progress.