Cross-cutting studies of per- and polyfluorinated alkyl substances (PFAS) in Arctic wildlife and humans.

This cross-cutting review focuses on the presence and impacts of per- and polyfluoroalkyl substances (PFAS) in the Arctic. Several PFAS undergo long-range transport via atmospheric (volatile polyfluorinated compounds) and oceanic pathways (perfluorinated alkyl acids, PFAAs), causing widespread contamination of the Arctic. Beyond targeting a few well-known PFAS, applying sum parameters, suspect and non-targeted screening are promising approaches to elucidate predominant sources, transport, and pathways of PFAS in the Arctic environment, wildlife, and humans, and establish their time-trends. Across wildlife species, concentrations were dominated by perfluorooctane sulfonic acid (PFOS), followed by perfluorononanoic acid (PFNA); highest concentrations were present in mammalian livers and bird eggs. Time trends were similar for East Greenland ringed seals (Pusa hispida) and polar bears (Ursus maritimus). In polar bears, PFOS concentrations increased from the 1980s to 2006, with a secondary peak in 2014-2021, while PFNA increased regularly in the Canadian and Greenlandic ringed seals and polar bear livers. Human time trends vary regionally (though lacking for the Russian Arctic), and to the extent local Arctic human populations rely on traditional wildlife diets, such as marine mammals. Arctic human cohort studies implied that several PFAAs are immunotoxic, carcinogenic or contribute to carcinogenicity, and affect the reproductive, endocrine and cardiometabolic systems. Physiological, endocrine, and reproductive effects linked to PFAS exposure were largely similar among humans, polar bears, and Arctic seabirds. For most polar bear subpopulations across the Arctic, modeled serum concentrations exceeded PFOS levels in human populations, several of which already exceeded the established immunotoxic thresholds for the most severe risk category. Data is typically limited to the western Arctic region and populations. Monitoring of legacy and novel PFAS across the entire Arctic region, combined with proactive community engagement and international restrictions on PFAS production remain critical to mitigate PFAS exposure and its health impacts in the Arctic.

Distinct profiles of oxylipid mediators in liver, lung, and placenta after maternal nano-TiO2 nanoparticle inhalation exposure.

Nano-titanium dioxide (nano-TiO2) is a widely used nanomaterial found in several industrial and consumer products, including surface coatings, paints, sunscreens and cosmetics, among others. Studies have linked gestational exposure to nano-TiO2 with negative maternal and fetal health outcomes. For example, maternal pulmonary exposure to nano-TiO2 during gestation has been associated not only with maternal, but also fetal microvascular dysfunction in a rat model. One mediator of this altered vascular reactivity and inflammation is oxylipid signaling. Oxylipids are formed from dietary lipids through several enzyme-controlled pathways as well as through oxidation by reactive oxygen species. Oxylipids have been linked to control of vascular tone, inflammation, pain and other physiological and disease processes. In this study, we use a sensitive UPLC-MS/MS based analysis to probe the global oxylipid response in liver, lung, and placenta of pregnant rats exposed to nano-TiO2 aerosols. Each organ presented distinct patterns in oxylipid signaling, as assessed by principal component and hierarchical clustering heatmap analysis. In general, pro-inflammatory mediators, such as 5-hydroxyeicosatetraenoic acid (1.6 fold change) were elevated in the liver, while in the lung, anti-inflammatory and pro-resolving mediators such as 17-hydroxy docosahexaenoic acid (1.4 fold change) were elevated. In the placenta the levels of oxylipid mediators were generally decreased, both inflammatory (e.g. PGE2, 0.52 fold change) and anti-inflammatory (e.g. Leukotriene B4, 0.49 fold change). This study, the first to quantitate the levels of these oxylipids simultaneously after nano-TiO2 exposure, shows the complex interplay of pro- and anti-inflammatory mediators from multiple lipid classes and highlights the limitations of monitoring the levels of oxylipid mediators in isolation.

Targeted and Suspect Screening of Per- and Polyfluoroalkyl Substances in Cosmetics and Personal Care Products.

Per- and polyfluoroalkyl substances (PFAS) are anthropogenic chemicals reported in cosmetics and personal care products as ingredients, possible impurities in the raw material manufacturing process, or degradation products. The purpose of this study was to further delineate contributions of these varying PFAS sources to these products. Thirty-eight cosmetics and personal care products were selected and analyzed for polyfluoroalkyl phosphates (PAPs), perfluoroalkyl carboxylic acids (PFCAs), fluorotelomer sulfonic acids (FTSAs), and perfluoroalkyl sulfonic acids (PFSAs) using targeted liquid chromatography tandem mass spectrometry (LC-MS/MS). A subset of products was also subjected to suspect screening using LC-high resolution mass spectrometry (HRMS) for >200 compounds. Results of LC-MS/MS and LC-HRMS indicated a predominant and ubiquitous presence of PAPs (detection frequency 99.7%, mean and median ΣPAPs 1 080 000 and 299 ng/g). Total median PFCA and PFSA concentrations were 3 and 38 times lower, respectively. There were significant correlations (Spearman's correlation coefficients = 0.60-0.81, p < 0.05) between 6:2 PAPs and their biotransformation products. Low levels of other PFAS classes were detected, including those previously measured in wastewater and human blood (e.g., hydrido-PFCAs), and five compounds associated with aqueous film-forming foams. Overall, these data highlight that cosmetics and personal care products can contain a breadth of PFAS at extremely high levels, leading to human and environmental exposure.

The human fecal microbiome contributes to the biotransformation of the PFAS surfactant 8:2 monosubstituted polyfluoroalkyl phosphate ester.

Polyfluoroalkyl phosphate esters (PAPs) can be found throughout society due to their numerous commercial applications. However, they also pose an environmental and health concern given their ability to undergo hydrolysis and oxidation to several bioactive and persistent products, including the perfluorocarboxylic acids (PFCAs). The metabolism of PAPs has been shown to occur in mammalian liver and intestine, however metabolism by the gut microbiome has not yet been investigated. In this study, human fecal samples were used to model the microbial population of the colon, to test whether these anaerobic microbes could facilitate 8:2 monosubstituted PAP (monoPAP) transformation. In vitro testing was completed by incubating the fecal samples with 8:2 monoPAP (400-10,000 nM) up to 120 minutes in an anaerobic chamber. Reactions were then terminated and the samples prepared for GC- and LC-MS/MS analysis. Metabolites of interest were the immediate hydrolysis product, the 8:2 fluorotelomer alcohol (FTOH), and 11 additional metabolites previously shown to form from 8:2 FTOH in both oxic and anoxic environments. The kinetics of 8:2 monoPAP transformation by gut microbiota were compared to those in human S9 liver and intestine fractions, both of which have active levels of hydrolyzing and oxidative enzymes that transform 8:2 monoPAP. Transformation rates from 8:2 monoPAP to 8:2 FTOH were highest in liver S9 > intestine S9 > fecal suspensions. The gut microbiome also produced a unique composition of oxidative metabolites, where the following intermediate metabolites were more abundant than terminal PFCAs: 8:2 fluorotelomer unsaturated carboxylic acid (FTUCA) > 8:2 fluorotelomer carboxylic acid (FTCA) > 7:2 Ketone ≈ perfluorohexanoic acid (PFHxA). Hydrolytic and oxidative metabolites contributed up to 30% of the molar balance after microbial 8:2 monoPAP transformation. Together, the results suggest that the gut microbiome can play a notable role in PAP biotransformation.

Emerging investigator series: human CYP2A6 catalyzes the oxidation of 6:2 fluorotelomer alcohol.

The biotransformation of 6:2 fluorotelomer alcohol (6:2 FTOH) results in the production of bioactive and persistent metabolites, including perfluorinated carboxylic acids (PFCAs). While the products of 6:2 FTOH metabolism have been elucidated in several animal models, the responsible cytochrome P450 (CYP) isoform(s) have not been reported. Here, we characterized the in vitro oxidation of 6:2 FTOH using human liver microsomes and recombinant human CYPs. Six major xenobiotic metabolizing CYPs were screened for their capacity to catalyze 6:2 FTOH oxidation using chemical inhibitors selective towards CYP isoforms. Of the CYP isoforms investigated, CYP2A6 was the only enzyme capable of catalyzing 6:2 FTOH in human liver microsomes, with KM and Vmax values of 4076 ng mL-1 and 69 ng mL-1 min-1, respectively. We further probed the metabolic mechanism by plotting the 6:2 FTOH kinetic profile and extrapolating data to several possible kinetic models. 6:2 FTOH oxidation followed the typical one-site Michaelis-Menten kinetic model. This study also reports that 6:2 FTOH loss is associated with active CYP2A6 by incubating microsomes with the selective CYP2A6 inhibitor tranylcypromine, which bound competitively to the enzyme as determined by an increased KM (8796 ng mL-1) but unchanged Vmax value. Collectively, these findings provide a mechanistic perspective on the potential importance of CYP2A6 in the metabolic activation and phase I elimination of 6:2 FTOH and indirect human exposure to PFCAs.

Epoxyeicosatrienoic acid (EET)-stimulated angiogenesis is mediated by epoxy hydroxyeicosatrienoic acids (EHETs) formed from COX-2.

Epoxyeicosatrienoic acids (EETs) are formed from the metabolism of arachidonic acid by cytochrome P450s. EETs promote angiogenesis linked to tumor growth in various cancer models that is attenuated in vivo by cyclooxygenase 2 (COX-2) inhibitors. This study further defines a role for COX-2 in mediating endothelial EET metabolism promoting angiogenesis. Using human aortic endothelial cells (HAECs), we quantified 8,9-EET-induced tube formation and cell migration as indicators of angiogenic potential in the presence and absence of a COX-2 inducer [phorbol 12,13-dibutyrate (PDBu)]. The angiogenic response to 8,9-EET in the presence of PDBu was 3-fold that elicited by 8,9-EET stabilized with a soluble epoxide hydrolase inhibitor (t-TUCB). Contributing to this response was the COX-2 metabolite of 8,9-EET, the 11-hydroxy-8,9-EET (8,9,11-EHET), which exogenously enhanced angiogenic responses in HAECs at levels comparable to those elicited by vascular endothelial growth factor (VEGF). In contrast, the 15-hydroxy-8,9-EET isomer was also formed but inactive. The 8,9,11-EHET also promoted expression of the VEGF family of tyrosine kinase receptors. These results indicate that 8,9-EET-stimulated angiogenesis is enhanced by COX-2 metabolism in the endothelium through the formation of 8,9,11-EHET. This alternative pathway for the metabolism of 8,9-EET may be particularly important in regulating angiogenesis under circumstances in which COX-2 is induced, such as in cancer tumor growth and inflammation.

Celecoxib Does Not Protect against Fibrosis and Inflammation in a Carbon Tetrachloride-Induced Model of Liver Injury.

The cyclooxygenase-2 (COX-2) selective inhibitor celecoxib is widely used in the treatment of pain and inflammation. Celecoxib has been explored as a possible treatment of liver fibrosis with contradictory results, depending on the model. The present study reports the effect of celecoxib in a 5-week carbon tetrachloride (CCl4)-induced liver fibrosis mouse model. Celecoxib alone and in combination with inhibitors of the enzyme-soluble epoxide hydrolase (sEH), as well as a dual inhibitor that targets both COX-2 and sEH, were administered via osmotic minipump to mice receiving intraperitoneal injections of CCl4 Collagen deposition was elevated in the mice treated with both celecoxib and CCl4 compared with the control or CCl4-only groups, as assessed by trichrome staining. Histopathology revealed more extensive fibrosis and cell death in the animals treated with both celecoxib and CCl4 compared with all other experimental groups. Although some markers of fibrosis, such as matrix metalloprotease, were unchanged or lowered in the animals treated with both celecoxib and CCl4, overall, hepatic fibrosis was more severe in this group. Cotreatment with celecoxib and an inhibitor of sEH or treatment with a dual inhibitor of COX-2 and sEH decreased the elevated levels of fibrotic markers observed in the group that received both celecoxib and CCl4 Oxylipid analysis revealed that celecoxib reduced the level of prostaglandin E2 relative to the CCl4 only group. Overall, celecoxib treatment did not decrease liver fibrosis in CCl4-treated mice.

Effects of triclosan in breast milk on the infant fecal microbiome.

Triclosan is frequently used for its antimicrobial properties and has been detected in human serum, urine, and breast milk. Animal and molecular studies have shown that triclosan exerts a wide range of adverse health effects at both high (ppm) and low (ppb) concentrations. Since triclosan is of growing concern to human and environmental health, there is a need to improve extraction procedures and to study additional effects from triclosan exposure. In this study, we have improved triclosan extraction from breast milk by using salt (MgSO4) to reduce emulsion formation and increase water polarity and water (∼80%) to enhance the overall extraction efficiency (∼3.5 fold). This extraction method was applied to breast milk samples collected from donors who i) recorded their use of triclosan-containing personal care products and ii) provided matching infant stool samples. Of the participants who had detectable amounts of triclosan in their breast milk, nine (75%) of them reported daily use of triclosan-containing personal care products. Levels of triclosan in breast milk were compared to the donor's infant's fecal microbiome. We found that the bacterial diversity in the fecal microbiome of the infants exposed to breast milk with detectable triclosan levels differed compared to their peers exposed to milk containing non-detectable amounts. This finding implies that exogenous chemicals are impacting microbiome diversity.

LC-MS/MS Analysis of the Epoxides and Diols Derived from the Endocannabinoid Arachidonoyl Ethanolamide.

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a useful tool to characterize the behavior of natural lipids within biological matrices. We report a LC-MS/MS method developed specifically to analyze CYP products of the arachidonoyl ethanolamide (anandamide, AEA), the epoxyeicosatrienoic acid ethanolamides (EET-EAs) and their hydrolyzed metabolites, and the dihydroxyeicosatrienoic acid ethanolamides (DHET-EAs). This method was used to measure EET-EA biotransformation to DHET-EAs by two human epoxide hydrolases: the soluble EH (sEH) and the microsomal EH (mEH). In general, sEH and mEH substrate preference was similar, based on kcat/KM. The 14,15-EET-EA and 11,12-EET-EA were the most efficiently hydrolyzed, followed by 8,9-EET-EA and 5,6-EET-EA. The method was also used to detect endogenous levels of these lipids in mouse tissues, although levels were below the instrumental detection limit (0.1-3.4 nM). Because both AEA and EETs are biologically active, the method described herein will be invaluable in revealing the role(s) of EET-EAs in vivo.

Synthesis of cyclooxygenase metabolites of 8,9-epoxyeicosatrienoic acid (EET): 11- and 15-hydroxy 8,9-EETs.

COX metabolites of 8,9-EET, previously observed as potent mitogenic lipid mediators, were synthesized for the first time by using two synthetic approaches. These synthetic materials allow for structural confirmation of COX metabolites of 8,9-EET and further study of their biological roles.

Cyclooxygenase-derived proangiogenic metabolites of epoxyeicosatrienoic acids.

Arachidonic acid (ARA) is metabolized by cyclooxygenase (COX) and cytochrome P450 to produce proangiogenic metabolites. Specifically, epoxyeicosatrienoic acids (EETs) produced from the P450 pathway are angiogenic, inducing cancer tumor growth. A previous study showed that inhibiting soluble epoxide hydrolase (sEH) increased EET concentration and mildly promoted tumor growth. However, inhibiting both sEH and COX led to a dramatic decrease in tumor growth, suggesting that the contribution of EETs to angiogenesis and subsequent tumor growth may be attributed to downstream metabolites formed by COX. This study explores the fate of EETs with COX, the angiogenic activity of the primary metabolites formed, and their subsequent hydrolysis by sEH and microsomal EH. Three EET regioisomers were found to be substrates for COX, based on oxygen consumption and product formation. EET substrate preference for both COX-1 and COX-2 were estimated as 8,9-EET > 5,6-EET > 11,12-EET, whereas 14,15-EET was inactive. The structure of two major products formed from 8,9-EET in this COX pathway were confirmed by chemical synthesis: ct-8,9-epoxy-11-hydroxy-eicosatrienoic acid (ct-8,9-E-11-HET) and ct-8,9-epoxy-15-hydroxy-eicosatrienoic acid (ct-8,9-E-15-HET). ct-8,9-E-11-HET and ct-8,9-E-15-HET are further metabolized by sEH, with ct-8,9-E-11-HET being hydrolyzed much more slowly. Using an s.c. Matrigel assay, we showed that ct-8,9-E-11-HET is proangiogenic, whereas ct-8,9-E-15-HET is not active. This study identifies a functional link between EETs and COX and identifies ct-8,9-E-11-HET as an angiogenic lipid, suggesting a physiological role for COX metabolites of EETs.

Is there a human health risk associated with indirect exposure to perfluoroalkyl carboxylates (PFCAs)?

The production and widespread use of poly- and perfluoroalkyl substances (PFAS) has led to their presence in the environment, wildlife, and humans. Particularly, the perfluoroalkyl carboxylates (PFCAs) are pervasive throughout the world and have been found at ng/mL concentrations in human blood. PFCAs, especially those having longer carbon chain lengths (≥C6), are associated with developmental and hormonal effects, immunotoxicity, and promote tumor growth in rodents through their role as PPARα agonists. Humans are directly exposed to PFCAs primarily through contaminated food, drinking water, and house dust. However, indirect exposure to PFCAs through the biotransformation of fluorotelomer-based substances may also be a significant, yet relatively underappreciated pathway. We are exposed to fluorotelomer-based substances through use of consumer products, ingestion of food, or from inhalation of dust particles, but the risk of this exposure has been largely uncharacterized. Here, we summarize the work that has been done to characterize toxicity of the classes of fluorotelomer-based substances shown to biotransform to PFCAs: the polyfluoroalkyl phosphate esters (PAPs), fluorotelomer alcohols (FTOHs), fluorotelomer iodides (FTIs), and fluorotelomer acrylate monomers (FTAcs). These fluorotelomer-based substances biotranform to yield PFCAs, yet also form bioactive intermediate metabolites, which have been observed to be more toxic than their corresponding PFCAs. We address what is known regarding the toxicity of the fluorotelomer-based substances and their metabolites, with focus on covalent binding to biological nucleophiles, such as glutathione, proteins, and DNA, as a possible mechanism of toxicity that may influence the risk of indirect exposure to PFCAs.

Protein binding associated with exposure to fluorotelomer alcohols (FTOHs) and polyfluoroalkyl phosphate esters (PAPs) in rats.

The biotransformation of fluorotelomer-based compounds such as fluorotelomer alcohols (FTOHs) and polyfluoroalkyl phosphate esters (PAPs) are sources of exposure to perfluorinated carboxylates (PFCAs), leading in part to the observation of significant concentrations of PFCAs in human blood. The biotransformation of FTOHs and PAPs yield intermediate metabolites that have been observed to covalently modify proteins. In the current investigation, the extent of covalent protein binding in Sprague-Dawley rats upon exposure to 8:2 FTOH and the 6:2 polyfluoroalkyl phosphate diester (6:2 diPAP) was quantified. The animals were administered a single dose of 8:2 FTOH or 6:2 diPAP at 100 mg/kg by oral gavage to monitor biotransformation and extent of protein binding within the liver, kidney, and plasma. In the 8:2 FTOH-dosed animals, perfluorooctanoate (PFOA) was produced as the primary PFCA, at 623.13 ± 59.3, 459.5 ± 171.8, and 397.3 ± 133.0 ng/g in the plasma, liver, and kidney, respectively. For the animals exposed to 6:2 diPAPs, perfluorohexanoate (PFHxA) was the primary PFCA produced, with maximum concentrations of 57.4 ± 6.5, 9.0 ± 1.2, and 25.3 ± 1.2 ng/g in the plasma, liver, and kidney, respectively. Protein binding was observed in the plasma, liver, and kidney after 8:2 FTOH and 6:2 diPAP exposure, with the most significant binding occurring in the liver (>100 nmol/g protein). This is the first study to link the exposure and in vivo biotransformation of fluorotelomer-based compounds to covalent protein binding.

Cellular toxicity associated with exposure to perfluorinated carboxylates (PFCAs) and their metabolic precursors.

The biotransformation of fluorotelomer based compounds yields saturated and unsaturated fluorotelomer aldehydes (FTALs and FTUALs, respectively) and carboxylic acids (FTCAs and FTUCAs, respectively) as intermediate metabolites that subsequently transform to perfluorinated carboxylic acids (PFCAs). Previous studies have demonstrated that the FTCAs and FTUCAs are 1 to 5 orders of magnitude more toxic than PFCAs after exposure to aquatic organisms. Additionally, FTUALs have demonstrated reactivity with proteins, which may be associated with toxicity through the inhibition of protein function. The purpose of this study was to carry out a comprehensive assessment of the relative toxicity between PFCAs and their intermediate precursor metabolites: the FTALs, FTUALs, FTCAs, and FTUCAs. Analytes were separately incubated with human liver epithelial (THLE-2) cells to assess how varying the functional group and the fluorinated chain length affects cell viability. For each analyte, dose-response EC50 values were calculated. The EC50 values for FTUCAs and FTCAs were similar, with values ranging from 22 ± 9 and 24 ± 9 µM for the 10:2 congeners to 1004 ± 20 and 1004 ± 24 µM for the 4:2 congeners, respectively. The EC50 values for the PFCAs ranged from 65 ± 41 (PFDA) to 1361 ± 146 (PFBA) µM. The range of toxicity between PFCAs and their acid precursors were similar. However, the comparative toxicity between the 6:2 and 8:2 congeners and their corresponding PFCA had toxicity thresholds that varied depending on the functional headgroup, where FTUALs ≥ FTALs > FTUCAs ≥ FTCAs > PFCAs. For all PFCAs and acid precursors, toxicity depended on the length of the fluorinated chain, where the longer chain lengths yielded greater bioaccumulation and enhanced toxicity, results which agreed with those previously reported. By contrast, FTALs and FTUALs were the most toxic of all the analytes examined, where toxicity was enhanced at shorter chain lengths, with EC50 values of 7 ± 1 µM (6:2 FTUAL) and 8.6 ± 0.8 µM (6:2 FTAL). DNA adducts were not detectable for the aldehyde precursors, using a quantitative long-range PCR method. Our data provide the first evidence that aldehyde intermediates have demonstrated toxicity in cellular systems that is more significant than PFCAs and their corresponding acid intermediates.

Covalent binding of fluorotelomer unsaturated aldehydes (FTUALs) and carboxylic acids (FTUCAs) to proteins.

Fluorotelomer unsaturated carboxylic acids and aldehydes (FTUCAs and FTUALs) are intermediate compounds that form from the biotransformation of fluorotelomer-based compounds. Previous evidence that FTUCAs and FTUALs bind to biological nucleophiles has indicated that protein binding might give rise to toxicity resulting from protein function disruption. The current study assesses the reactivity of FTUALs and FTUCAs by probing the covalent interactions of FTUALs and FTUCAs with proteins present in rat liver microsomes and bovine blood plasma. The FTUALs exhibited significant levels of protein covalent binding, with binding levels ranging from 20.1 (±2.8)% to 71.3 (±19.5)% in microsomes and 24.0 (±1.5)% to 82.5 (±14.0)% in blood plasma. By contrast, the FTUCAs did not exhibit any apparent covalent binding. Bovine serum albumin (BSA) was extracted and isolated from the plasma after incubation of 8:2 FTUAL (5-100 µM). Electrospray ionization mass spectrometry (ESI-MS) was used to investigate the stoichiometry of 8:2 FTUAL covalently bound to BSA; three measurable FTUAL adducts were formed with BSA. To compare the percent binding results from the FTUAL microsome incubation experiments, 8:2 FTOH was incubated in microsomes to determine the protein binding associated with the biotransformation of 8:2 FTOH. Results from this study showed that the biotransformation of 8:2 FTOH yielded 26.1 (±3.0)% binding, and was statistically similar to the percent binding associated with 8:2 FTUAL exposure (p > 0.05), indicating that the binding of 8:2 FTUAL to proteins might be a primary fate in the biotransformation of 8:2 FTOH.

In vitro interactions of biological nucleophiles with fluorotelomer unsaturated acids and aldehydes: fate and consequences.

Fluorotelomer unsaturated aldehydes and acids (FTUALs and FTUCAs) are intermediate metabolites that form from the biotransformation of fluorotelomer-based chemicals. FTUALs and FTUCAs have been previously suggested to contribute to the toxicity associated with human exposure to fluorotelomer compounds by covalently binding to biological nucleophiles. However, the extent of their reactivity has only been assessed with glutathione. The purpose of the present study was to assess the reactivity of these intermediate metabolites with a series of nucleophilic amino acids and model proteins. In vitro experiments were carried out in an aqueous buffer system to determine the reactivity of nucleophilic amino acids with FTUCAs and FTUALs having varying fluorinated chain lengths. Using (19)F NMR spectroscopy to monitor the disappearance of the FTUCAs and FTUAL signals and the production of a fluoride signal, reaction rate constants were determined under pseudo-first-order conditions. The FTUCAs reacted only with cysteine with the following second order rate constants: 3.63 (± 1.37) × 10(-5) min(-1) mM(-1) (4:2 FTUCA), 1.19 (± 0.91) × 10(-5) min(-1) mM(-1) (6:2 FTUCA), and 4.56 (± 0.94) × 10(-5) min(-1) mM(-1) (8:2 FTUCA). The FTUALs were significantly more reactive than any of the FTUCAs with reactivity decreasing in the following order: cysteine >> histidine > lysine >> arginine. The following second-order rate constants were obtained: 5.7 (± 4.2) × 10(-4) min(-1) mM(-1) (histidine), 4.3 (± 1.4) × 10(-4) min(-1) mM(-1) (lysine), and 1.4 (± 0.73) × 10(-4) min(-1) mM(-1) (arginine). FTUCAs and FTUALs were also reacted with model proteins to assess their potential for forming covalent adducts. Electrospray ionization mass spectrometry (ESI-MS) was used to investigate the stoichiometry of FTUCAs and FTUALs covalently bound to apomyoglobin (ApoMg) and human serum albumin (HSA). FTUCAs were not reactive, whereas two measurable FTUAL adducts were formed with both ApoMg and HSA at each of the FTUAL chain lengths (6:2, 8:2, and 10:2). This is the first study to probe the reactivity of FTUALs and FTUCAs with nucleophiles other than glutathione, further elucidating possible FTUAL and FTUCA fate within biological systems.

Assessing the structure-activity relationships of fluorotelomer unsaturated acids and aldehydes with glutathione. Reactivity of glutathione with fluorotelomer unsaturated acids and aldehydes.

Fluorotelomer alcohols (FTOHs) have been shown to degrade via abiotic and biotic mechanisms to perfluorocarboxylates (PFCAs) which are environmentally persistent and bioaccumulate in humans and wildlife depending on their chain length. Fluorotelomer unsaturated aldehydes (FTUALs) and acids (FTUCAs) are intermediate metabolites that form from the degradation of FTOHs. Their potential for toxicity is not yet defined and may be more significant compared to PFCAs. Past studies have shown that these intermediates form adducts with glutathione (GSH). The purpose of this study was to further assess the reactivity of these intermediate compounds. In vitro experiments were carried out in an aqueous buffer system (pH 7.4) where FTUCAs and FTUALs of varying chain lengths were reacted with GSH. To quantify the reactivity of FTUCAs and FTUALs, unreacted free GSH was derivatized with 5,5'-dithiobis(2-nitrobenzoic acid), its absorbance measured at 412 nm, and the percentage of unconjugated free GSH evaluated over time. EC50 values were obtained for the reactions of GSH with acrolein and methyl methacrylate to assess the accuracy of the method, as well as for acrylic acid, FTUCAs, and FTUALs. The results of this study indicated that α,ß-unsaturated aldehydes are comparatively the most reactive and reaction with GSH may be influenced by the length of the fluorinated tail. This is the first study to examine the relationship of FTUCAs and FTUALs with biological nucleophiles by quantifying their intrinsic reactivity.

Perfluorinated carboxylic acids in directly fluorinated high-density polyethylene material.

Perfluorinated carboxylic acids (PFCAs) are ubiquitous in the environment and have been detected in human blood worldwide. One potential route is direct exposure to PFCAs through contact with polymers that have been fluorinated through a process referred to as direct fluorination. PFCAs are hypothesized to be reaction byproducts of direct fluorination when trace amounts of oxygen are present. The objective of this research was to investigate whether PFCAs could be measured in directly fluorinated high-density polyethylene (HDPE) bottles. PFCAs were quantified using Soxhlet extraction with methanol, followed by LC-MS/MS analysis. Total concentrations of PFCAs ranged from 8.5 ± 0.53 to 113 ± 2.5 ng/bottle (1 L), with the short-chain PFCAs, perfluoropropanoic, perfluorobutanoic, perfluoropentanoic, and perfluorohexanoic acids, being the dominant congeners observed. Relative PFCA concentrations varied depending on fluorination level. Structural isomers were detected using (19)F NMR and are hypothesized to have formed during the fluorination process; NMR data revealed the linear isomer typically comprised 55% of the examined sample. Internally branched, isopropyl branched, and t-butyl PFCA isomers of varying chain length were also identified. Electrochemical fluorination was previously thought to be the only source of branched PFCA isomers. The observation here of branched isomers suggests direct fluorination may be an additional source of exposure to these chemicals. The purpose of this study was to measure PFCAs in directly fluorinated material, serving as a previously unidentified source contributing to the environmental load of PFCAs, with potential for human exposure.

Polypyrroles as antioxidants: kinetic studies on reactions of bilirubin and biliverdin dimethyl esters and synthetic model compounds with peroxyl radicals in solution. Chemical calculations on selected typical structures.

[reaction: see text] Rate constants for hydrogen-atom transfer (HAT) from bilirubin dimethyl ester (BRDE) and biliverdin dimethyl ester (BVDE) to peroxyl radicals during inhibited autoxidation of styrene initiated by azo-bisisobutyronitrile (AIBN) were k(inh)(BRDE) = 22.5 x 10(4) and k(inh)(BVDE) = 10.2 x 10(4) M(-1) s(-1), and the stoichiometric factors (n) were 2.0 and 2.7, respectively. A synthetic tetrapyrrole (bis(dipyrromethene)) containing the alpha-central (2,2') CH2 linkage gave k(inh) = 39.9 x 10(4) M(-1) s(-1) and n = 2.3, whereas the beta-linked (3,3') isomer was not an active antioxidant. Several dipyrrinones were synthesized as mimics of the two outer heterocyclic rings of bilirubin and biliverdin. The dipyrrinones containing N-H groups in each ring were active antioxidants, whereas those lacking two such "free" N-H groups, such as N-CH3 dipyrrinones and dipyrromethenes, did not exhibit antioxidant activity. Overall, the relative k(inh) values compared to those of phenolic antioxidants, 2,6-di-tert-butyl-4-methoxyphenol (DBHA) and 2,6-di-tert-butyl-4-methylphenol (BHT), were 2,2'-bis(dipyrromethene) > BRDE > DBHA > dipyrrinones > BVDE > BHT. This general trend in antioxidant activities was also observed for the inhibited autoxidation of cumene initiated by AIBN. Chemical calculations of the N-H bond dissociation enthalpies (BDEs) of the typical structures support a HAT mechanism from N-H groups to trap peroxyl radicals. Intramolecular hydrogen bonding of intermediate nitrogen radicals has a major influence on the antioxidant activities of all compounds studied. Indeed, chemical calculations showed that the initial nitrogen radical from a dipyrrinone is stabilized by 9.0 kcal/mol because of H-bonding between the N-H remaining on one ring and the ground-state pyrrolyl radical of the adjacent ring in the natural zusammen structure. The calculated minimum structure of bilirubin shows strong intramolecular H-bonding of the N-H groups with carbonyl groups resulting in the known "ridge-tile" structure which is not an active HAT antioxidant. The calculated minimum structure of biliverdin is planar. BRDE is readily converted into BVDE by reaction with the electron-deficient DPPH* radical under argon in chlorobenzene. An electron-transfer mechanism is proposed for the initiating step in this reaction, and this is supported by the relatively low ionizing potential of a model dipyrrole representing the two central rings of bilirubin.