Validation of putative biomarkers of furan exposure through quantitative analysis of furan metabolites in urine of F344 rats exposed to stable isotope labeled furan.

Humans are chronically exposed to furan, a potent liver toxicant and carcinogen that occurs in a variety of heat-processed foods. Assessment of human exposure based on the furan content in foods is, however, subject to some uncertainty due to the high volatility of furan. Biomarker monitoring is thus considered an alternative or complementary approach to furan exposure assessment. Previous work suggested that urinary furan metabolites derived from the reaction of cis-2-butene-1,4-dial (BDA), the reactive intermediate of furan, with glutathione (GSH) or amino acids may serve as potential biomarkers of furan exposure. However, some metabolites were also reported to occur in urine of untreated animals, indicating either background contamination via animal feed or endogenous sources, which may limit their suitability as biomarkers of exposure. The overall aim of the present study was to accurately establish the correlation between external dose and concentration of furan metabolites in urine over time and to discriminate against endogenous formation and furan intake via feed. To this end, the furan metabolites GSH-BDA (N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-L-cysteinylglycine), NAcLys-BDA (R-2-(acetylamino)-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid), NAcCys-BDA-NAcLys (N-acetyl-S-[1-[5-(acetylamino)-5-carboxypentyl]-1H-pyrrol-3-yl]-L-cysteine) and NAcCys-BDA-NAcLys sulfoxide (N-acetyl-S-[1-[5-(acetylamino)-5-carboxypentyl]-1H-pyrrol-3-yl]-L-cysteine sulfoxide) were simultaneously analyzed by stable isotope dilution ESI-LC-MS/MS as unlabeled and [13C4]-furan dependent metabolites following oral administration of a single oral dose of isotopically labelled [13C4]-furan (0.1, 1, 10, 100 and 1000 µg/kg bw) to male and female F344/DuCrl rats. Although a linear correlation between urinary excretion of [13C4]-furan-dependent metabolites was observed, analysis of unlabeled NAcLys-BDA, NAcCys-BDA-NAcLys and NAcCys-BDA-NAcLys sulfoxide revealed substantial, fairly constant urinary background levels throughout the course of the study. Analysis of furan in animal feed excluded feed as a source for these background levels. GSH-BDA was identified as the only furan metabolite without background occurrence, suggesting that it may present a specific biomarker to monitor external furan exposure. Studies in humans are now needed to establish if analysis of urinary GSH-BDA may provide reliable exposure estimates.

Removal of Remdesivir's Metabolite GS-441524 by Hemodialysis in a Double Lung Transplant Recipient with COVID-19.

Remdesivir has reported efficacy against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro and in vivo Drug-drug interactions limit therapeutic options in transplant patients. Remdesivir and its metabolite GS-441524 are excreted principally in urine. In intensive care unit (ICU) settings, in which multiple-organ dysfunctions can occur rapidly, hemodialysis may be a viable option for maintaining remdesivir treatment, while improving tolerance, by removing both remdesivir's metabolite (GS-441524) and sulfobutylether ß-cyclodextrin sodium (SEBCD). Additional studies may prove informative, particularly in the evaluations of therapeutic options for coronavirus disease 2019 (COVID-19).

A New UPLC-qTOF Approach for Elucidating Furan and 2-Methylfuran Metabolites in Human Urine Samples after Coffee Consumption.

We have investigated urine samples after coffee consumption using targeted and untargeted approaches to identify furan and 2-methylfuran metabolites in urine samples by UPLC-qToF. The aim was to establish a fast, robust, and time-saving method involving ultra-performance liquid chromatography-quantitative time-of-flight tandem mass spectrometry (UPLC-qToF-MS/MS). The developed method detected previously reported metabolites, such as Lys-BDA, and others that had not been previously identified, or only detected in animal or in vitro studies. The developed UPLC-qToF method detected previously reported metabolites, such as lysine-cis-2-butene-1,4-dial (Lys-BDA) adducts, and others that had not been previously identified, or only detected in animal and in vitro studies. In sum, the UPLC-qToF approach provides additional information that may be valuable in future human or animal intervention studies.

Role of mitochondria-derived reactive oxygen species in microvascular dysfunction in chronic kidney disease.

Cardiovascular disease is the leading cause of mortality in chronic kidney disease (CKD). Mitochondrial dysfunction secondary to CKD is a potential source of oxidative stress that may impair vascular function. This study sought to determine if mitochondria-derived reactive oxygen species contribute to microvascular dysfunction in stage 3-5 CKD. Cutaneous vasodilation in response to local heating was assessed in 20 CKD patients [60 ± 13 yr; estimated glomerular filtration rate (eGFR) 46 ± 13 ml·kg-1·1.73 m-2] and 11 matched healthy participants (58 ± 2 yr; eGFR >90 ml·kg-1·1.73 m-2). Participants were instrumented with two microdialysis fibers for the delivery of 1) Ringer solution, and 2) the mitochondria- specific superoxide scavenger MitoTempo. Skin blood flow was measured via laser Doppler flowmetry during standardized local heating (42°C). Cutaneous vascular conductance (CVC) was calculated as a percentage of the maximum conductance achieved with sodium nitroprusside infusion at 43°C. Urinary isofuran/F2-isoprostane ratios were assessed by gas-chromatography mass spectroscopy. Isofuran-to-F2-isoprostane ratios were increased in CKD patients (3.08 ± 0.32 vs. 1.69 ± 0.12 arbitrary units; P < 0.01) indicative of mitochondria-derived oxidative stress. Cutaneous vasodilation was impaired in CKD compared with healthy controls (87 ± 1 vs. 92 ± 1%CVCmax; P < 0.01). Infusion of MitoTempo significantly increased the plateau phase CVC in CKD patients (CKD Ringer vs. CKD MitoTempo: 87 ± 1 vs. 93 ± 1%CVCmax; P < 0.01) to similar levels observed in healthy controls ( P = 0.9). These data provide in vivo evidence that mitochondria-derived reactive oxygen species contribute to microvascular dysfunction in CKD and suggest that mitochondrial dysfunction may be a potential therapeutic target to improve CKD-related vascular dysfunction.

Exposure assessment of process-related contaminants in food by biomarker monitoring.

Exposure assessment is a fundamental part of the risk assessment paradigm, but can often present a number of challenges and uncertainties. This is especially the case for process contaminants formed during the processing, e.g. heating of food, since they are in part highly reactive and/or volatile, thus making exposure assessment by analysing contents in food unreliable. New approaches are therefore required to accurately assess consumer exposure and thus better inform the risk assessment. Such novel approaches may include the use of biomarkers, physiologically based kinetic (PBK) modelling-facilitated reverse dosimetry, and/or duplicate diet studies. This review focuses on the state of the art with respect to the use of biomarkers of exposure for the process contaminants acrylamide, 3-MCPD esters, glycidyl esters, furan and acrolein. From the overview presented, it becomes clear that the field of assessing human exposure to process-related contaminants in food by biomarker monitoring is promising and strongly developing. The current state of the art as well as the existing data gaps and challenges for the future were defined. They include (1) using PBK modelling and duplicate diet studies to establish, preferably in humans, correlations between external exposure and biomarkers; (2) elucidation of the possible endogenous formation of the process-related contaminants and the resulting biomarker levels; (3) the influence of inter-individual variations and how to include that in the biomarker-based exposure predictions; (4) the correction for confounding factors; (5) the value of the different biomarkers in relation to exposure scenario's and risk assessment, and (6) the possibilities of novel methodologies. In spite of these challenges it can be concluded that biomarker-based exposure assessment provides a unique opportunity to more accurately assess consumer exposure to process-related contaminants in food and thus to better inform risk assessment.

Can creatine supplementation form carcinogenic heterocyclic amines in humans?

There is a long-standing concern that creatine supplementation could be associated with cancer, possibly by facilitating the formation of carcinogenic heterocyclic amines (HCAs). This study provides compelling evidence that both low and high doses of creatine supplementation, given either acutely or chronically, does not cause a significant increase in HCA formation. HCAs detection was unrelated to creatine supplementation. Diet was likely to be the main factor responsible for HCAs formation after either placebo (n = 6) or creatine supplementation (n = 3). These results directly challenge the recently suggested biological plausibility for the association between creatine use and risk of testicular germ cell cancer. Creatine supplementation has been associated with increased cancer risk. In fact, there is evidence indicating that creatine and/or creatinine are important precursors of carcinogenic heterocyclic amines (HCAs). The present study aimed to investigate the acute and chronic effects of low- and high-dose creatine supplementation on the production of HCAs in healthy humans (i.e. 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx), 2-amino-(1,6-dimethylfuro[3,2-e]imidazo[4,5-b])pyridine (IFP) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx)). This was a non-counterbalanced single-blind crossover study divided into two phases, in which low- and high-dose creatine protocols were tested. After acute (1 day) and chronic supplementation (30 days), the HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx were assessed through a newly developed HPLC-MS/MS method. Dietary HCA intake and blood and urinary creatinine were also evaluated. Out of 576 assessments performed (from 149 urine samples), only nine (3 from creatine and 6 from placebo) showed quantifiable levels of HCAs (8-MeIQx: n = 3; 4,8-DiMeIQx: n = 2; PhIP: n = 4). Individual analyses revealed that diet rather than creatine supplementation was the main responsible factor for HCA formation in these cases. This study provides compelling evidence that both low and high doses of creatine supplementation, given either acutely or chronically, did not cause increases in the carcinogenic HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx in healthy subjects. These findings challenge the long-existing notion that creatine supplementation could potentially increase the risk of cancer by stimulating the formation of these mutagens.

Abundant Rodent Furan-Derived Urinary Metabolites Are Associated with Tobacco Smoke Exposure in Humans.

Furan, a possible human carcinogen, is found in heat treated foods and tobacco smoke. Previous studies have shown that humans are capable of converting furan to its reactive metabolite, cis-2-butene-1,4-dial (BDA), and therefore may be susceptible to furan toxicity. Human risk assessment of furan exposure has been stymied because of the lack of mechanism-based exposure biomarkers. Therefore, a sensitive LC-MS/MS assay for six furan metabolites was applied to measure their levels in urine from furan-exposed rodents as well as in human urine from smokers and nonsmokers. The metabolites that result from direct reaction of BDA with lysine (BDA-N(α)-acetyllysine) and from cysteine-BDA-lysine cross-links (N-acetylcysteine-BDA-lysine, N-acetylcysteine-BDA-N(α)-acetyllysine, and their sulfoxides) were targeted in this study. Five of the six metabolites were identified in urine from rodents treated with furan by gavage. BDA-N(α)-acetyllysine, N-acetylcysteine-BDA-lysine, and its sulfoxide were detected in most human urine samples from three different groups. The levels of N-acetylcysteine-BDA-lysine sulfoxide were more than 10 times higher than that of the corresponding sulfide in many samples. The amount of this metabolite was higher in smokers relative to that in nonsmokers and was significantly reduced following smoking cessation. Our results indicate a strong relationship between BDA-derived metabolites and smoking. Future studies will determine if levels of these biomarkers are associated with adverse health effects in humans.

Acetaminophen attenuates lipid peroxidation in children undergoing cardiopulmonary bypass.

OBJECTIVE: Hemolysis, occurring during cardiopulmonary bypass, is associated with lipid peroxidation and postoperative acute kidney injury. Acetaminophen inhibits lipid peroxidation catalyzed by hemeproteins and in an animal model attenuated rhabdomyolysis-induced acute kidney injury. This pilot study tests the hypothesis that acetaminophen attenuates lipid peroxidation in children undergoing cardiopulmonary bypass. DESIGN: Single-center prospective randomized double-blinded study. SETTING: University-affiliated pediatric hospital. PATIENTS: Thirty children undergoing elective surgical correction of a congenital heart defect. INTERVENTIONS: Patients were randomized to acetaminophen (OFIRMEV [acetaminophen] injection; Cadence Pharmaceuticals, San Diego, CA) or placebo every 6 hours for four doses starting before the onset of cardiopulmonary bypass. MEASUREMENT AND MAIN RESULTS: Markers of hemolysis, lipid peroxidation (isofurans and F2-isoprostanes), and acute kidney injury were measured throughout the perioperative period. Cardiopulmonary bypass was associated with a significant increase in free hemoglobin (from a prebypass level of 9.8 ± 6.2 mg/dL to a peak of 201.5 ± 42.6 mg/dL postbypass). Plasma and urine isofuran and F2-isoprostane concentrations increased significantly during surgery. The magnitude of increase in plasma isofurans was greater than the magnitude in increase in plasma F2-isoprostanes. Acetaminophen attenuated the increase in plasma isofurans compared with placebo (p = 0.02 for effect of study drug). There was no significant effect of acetaminophen on plasma F2-isoprostanes or urinary makers of lipid peroxidation. Acetaminophen did not affect postoperative creatinine, urinary neutrophil gelatinase-associated lipocalin, or prevalence of acute kidney injury. CONCLUSION: Cardiopulmonary bypass in children is associated with hemolysis and lipid peroxidation. Acetaminophen attenuated the increase in plasma isofuran concentrations. Future studies are needed to establish whether other therapies that attenuate or prevent the effects of free hemoglobin result in more effective inhibition of lipid peroxidation in patients undergoing cardiopulmonary bypass.

Dosimetry of human exposure to furan and 2-methylfuran by monitoring urinary biomarkers.

Furan and 2-methylfuran (2-MF) can form during food processing and accumulate in foods at various concentrations depending on processing technology and beverage/meal preparation methods applied prior to consumption. Here, we report a controlled dosimetry study with 20 volunteers (10 male, 10 female) to monitor dietary furan/2-MF exposure. The volunteers followed an eleven-day furan/2-MF-restricted diet in which they consumed freshly prepared coffee brew containing known amounts of furan and 2-MF on two separate occasions (250 mL and 500 mL on days 4 and 8, respectively). Urine was collected over the whole study period and analyzed for key metabolites derived from the primary oxidative furan metabolite cis-2-butene-1,4-dial (BDA) (i.e., Lys-BDA, AcLys-BDA and cyclic GSH-BDA) and the primary 2-MF metabolite acetylacrolein (AcA, 4-oxo-pent-2-enal) (i.e., Lys-AcA and AcLys-AcA). A previously established stable isotope dilution analysis (SIDA) method was utilized. Excretion kinetics revealed two peaks (at 0-2 and 24-36 h) for AcLys-BDA, Lys-BDA, AcLysAcA and LysAcA, whereas GSH-BDA showed a single peak. Notably, women on average excreted the metabolite GSH-BDA slightly faster than men, indicating gender differences. Overall, the study provided further insights into the spectrum of possible biomarkers of furan and 2-methyfuran metabolites occurring in the urine of volunteers after coffee consumption.

Mass balance study of [¹4C]eribulin in patients with advanced solid tumors.

This mass balance study investigated the metabolism and excretion of eribulin, a nontaxane microtubule dynamics inhibitor with a novel mechanism of action, in patients with advanced solid tumors. A single approximately 2 mg (approximately 80 µCi) dose of [¹4C]eribulin acetate was administered as a 2 to 5 min bolus injection to six patients on day 1. Blood, urine, and fecal samples were collected at specified time points on days 1 to 8 or until sample radioactivity was ≤1% of the administered dose. Mean plasma eribulin exposure (627 ng · h/ml) was comparable with that of total radioactivity (568 ng Eq · h/ml). Time-matched concentration ratios of eribulin to total radioactivity approached unity in blood and plasma, indicating that unchanged parent compound constituted almost all of the eribulin-derived radioactivity. Only minor metabolites were detected in plasma samples up to 60 min postdose, pooled across patients, each metabolite representing ≤0.6% of eribulin. Elimination half-lives for eribulin (45.6 h) and total radioactivity (42.3 h) were comparable. Eribulin-derived radioactivity excreted in feces was 81.5%, and that of unchanged eribulin was 61.9%. Renal clearance (0.301 l/h) was a minor component of total eribulin clearance (3.93 l/h). Eribulin-derived radioactivity excreted in urine (8.9%) was comparable with that of unchanged eribulin (8.1%), indicating minimal excretion of metabolite(s) in urine. Total recovery of the radioactive dose was 90.4% in urine and feces. Overall, no major metabolites of eribulin were detected in plasma. Eribulin is eliminated primarily unchanged in feces, whereas urine constitutes a minor route of elimination.

Surveillance program for former PCB-exposed workers of a transformer and capacitor recycling company, family members, employees of surrounding companies, and area residents--executive summary.

In a German company polychlorinated biphenyls (PCB)-containing transformers and capacitors were recycled on a large scale. Human biomonitoring revealed a high PCB body burden in workers of the recycling company, in surrounding locations of this plant, in companies in the neighborhood of this plant, and in family members of these employees. In order to clarify whether possible adverse health effects occurred or may occur in the future, a prospective surveillance program was initiated. After an extensive literature search, an interdisciplinary group of experts developed a surveillance program based on current knowledge with respect to possible adverse health effects that might occur in the recycling process of transformers and capacitors. Exposure to various hazardous substances (PCB, polychlorinated dibenzo-p-dioxins and dibenzo-furans [PCDD/F], metals, solvents) was considered. Criteria derived from human biomonitoring results of PCB were used for admission to the program. Participants in the surveillance program are first informed about risks and aims of the program. Subsequently, physicians started a detailed documentation of participants' general and occupational history, with their complaints, diseases, and nutritional habits, as well as information regarding their living areas, by means of a standardized questionnaire. In addition, separate examinations were performed to detect possible neurological, immunological, (neuro)psychological, hormonal, and skin effects. Moreover, DNA exposure as assessed by the comet assay and antioxidative status were determined. The program will be offered at yearly intervals for 3 years, and then at 5 and 10 years after program onset. Until now the program has proved to be feasible, and acceptance among workers and their families has been high. Based on the results, criteria will be developed to define adverse health effects that might be attributable to a hazardous substance exposure.

[Determination of tetrahydrofuran in urine by headspace solid-phase microextraction and gas chromatography].

OBJECTIVE: Headspace solid-phase microextraction (HS-SPME) was used pre-concentration procedure for the determination of tetrahydrofuran in urine by gas chromatography with hydrogen flame detector. METHODS: Several parameters controlling SPME was studied and optimised: SPME fiber, extraction time and extraction temperature, desorption time and desorption temperature. RESULTS: Under optimal conditions, the correlation coefficient was 0.9998 and good recoveries (range from 93.0% ∼ 100.8%) were achieved, the detection limit was 0.5 µg/L. CONCLUSION: The method can be applied to the determination of trace amount of tetrahydrofuran in urine.

Identification of furan metabolites derived from cysteine-cis-2-butene-1,4-dial-lysine cross-links.

Furan is a rodent hepatotoxicant and carcinogen. Because this compound is an important industrial intermediate and has been detected in heat-processed foods and smoke, humans are likely exposed to this toxic compound. Characterization of urinary metabolites of furan will lead to the development of biomarkers to assess human health risks associated with furan exposure. Previous studies indicate that furan is oxidized to a reactive alpha,beta-unsaturated dialdehyde, cis-2-butene-1,4-dial (BDA), in a reaction catalyzed by cytochrome P450. Five previously characterized metabolites are derived from the reaction of BDA with cellular nucleophiles such as glutathione and protein. They include the monoglutathione reaction product, N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-l-cysteinylglycine cyclic sulfide, and its downstream metabolite, S-[1-(1,3-dicarboxypropyl)-1H-pyrrol-3-yl]methylthiol, as well as (R)-2-acetylamino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid and N-acetyl-S-[1-(5-acetylamino-5-carboxypentyl)-1H-pyrrol-3-yl]-l-cysteine and its sulfoxide. The last two compounds are downstream metabolites of a BDA-derived cysteine-lysine cross-link, S-[1-(5-amino-5-carboxypentyl)-1H-pyrrol-3-yl]-l-cysteine. In this report, we present the characterization of seven additional urinary furan metabolites, all of which are derived from this cross-link. The cysteinyl residue is subject to several biotransformation reactions, including N-acetylation and S-oxidation. Alternatively, it can undergo beta-elimination followed by S-methylation to a methylthiol intermediate that is further oxidized to a sulfoxide. The lysine portion of the cross-link either is N-acetylated or undergoes a transamination reaction to generate an alpha-ketoacid metabolite that undergoes oxidative decarboxylation. Some of these metabolites are among the most abundant furan metabolites present in urine as judged by LC-MS/MS analysis, indicating that the oxidation of furan to BDA and BDA's subsequent reaction with cellular cysteine and lysine residues may represent a significant in vivo pathway of furan biotransformation. Because they are derived from cellular BDA reaction products, these metabolites are markers of furan exposure and bioactivation and could be explored as potential biomarkers in human studies.

Urinary metabolites of furan in waterpipe tobacco smokers compared to non-smokers in home settings in the US.

OBJECTIVES: Determine uptake of furan, a potential human carcinogen, in waterpipe tobacco (WPT) smokers in home settings. METHODS: We analysed data from a US convenience sample of 50 exclusive WPT smokers, mean age 25.3 years, and 25 non-smokers, mean age 25.5 years. For WPT smokers, data were collected at a home visit by research assistants during which participants smoked one WPT head of one brand for a mean of 33.1 min in their homes. Research assistants provided and prepared a WP for participants by weighing and loading 10 g of WPT in the WP head. At the completion of the smoking session, research assistants measured the remaining WPT. Cotinine and six furan metabolites were quantified in first morning urine samples provided on 2 consecutive days for non-smokers, and on the morning of a WPT smoking session and on the following morning for smokers. RESULTS: WPT smokers consumed a mean of 2.99 g WPT. In WPT smokers, urinary cotinine levels increased significantly 26.1 times the following morning; however, urinary metabolites of furan did not increase significantly. Compared to non-smokers, 2 furan metabolites, N-acetyl-S-[1-(5-acetylamino-5-carboxylpentyl)-1H-pyrrol-3-yl]-L-cysteine and N-acetyl-S-[1-(5-amino-5-carboxypentyl)-1H-pyrrol-3-yl]-L-cysteine sulfoxide, were significantly higher in WPT smokers in pre and in post WPT smoking levels. CONCLUSIONS: To enable a more rigorous assessment of furan exposure from WPT smoking, future research should determine furan concentrations in WPT smoke, quantify furan metabolites from users of various WPT brands; and extend the investigation to social settings where WPT smoking is habitually practiced.

Multiplex detection of 14 fentanyl analogues and U-47700 in biological samples: Application to a panel of French hospitalized patients.

Synthetic opioids (SO) associated with the recent alarming increase of deaths and intoxications in United States of America and Europe are not detected by the usual first-line opiates drug screening assays. We developed a liquid chromatography tandem mass spectrometry analytical method for the multiplex detection of 14 fentanyl analogues (2-furanylfentanyl, 4-ANPP, 4-methoxybutyrylfentanyl, acrylfentanyl, alfentanil, carfentanil, despropionyl-2-fluorofentanyl, fentanyl, methoxyacetylfentanyl, norfentanyl, ocfentanil, remifentanil, sufentanil and valerylfentanyl) and U-47700 in whole blood and urine samples. The method was validated according to the requirements of ISO 15189. A simple and fast liquid-liquid extraction (LLE) with De-Tox Tube-A was performed leading to better recovery of molecules in urine than in blood samples. Depending on the compound, the limits of detection (LODs) ranged from 0.01 to 0.10 ng/mL and from 0.02 to 0.05 ng/mL in whole blood and urine, respectively. Calibration curves were linear in the range 0.5-50.0 ng/mL and the limit of quantification (LOQ) ranged from 0.10 to 0.40 ng/mL in blood. Internal quality controls at 1 and 40 ng/mL showed intra-day and between-day precision and accuracy bias below 10% in urine and 15% in blood. The method was applied to the screening of 211 urine samples from patients admitted in emergency or addiction departments. The presence of legal fentanyl analogues in 5 urine samples was justified by their therapeutic use as analgesics. Only one patient was concerned by fentanyl misuse and addiction whereas no illegal SO was detected. This study is not in favor of a huge misuse of SO in the Lorraine region.

Degraded protein adducts of cis-2-butene-1,4-dial are urinary and hepatocyte metabolites of furan.

Furan is a liver toxicant and carcinogen in rodents. On the basis of these observations and the large potential for human exposure, furan has been classified as a possible human carcinogen. The mechanism of tumor induction by furan is unknown. However, the toxicity requires cytochrome P450-catalyzed oxidation of furan. The product of this oxidation, cis-2-butene-1,4-dial (BDA), reacts readily with glutathione, amino acids, and DNA and is a bacterial mutagen in Ames assay strain TA104. Characterization of the urinary metabolites of furan is expected to provide information regarding the structure(s) of the reactive metabolite(s). Recently, several urinary metabolites have been identified. We reported the presence of a monoglutathione-BDA reaction product, N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-l-cysteinylglycine cyclic sulfide. Three additional urinary metabolites of furan were also characterized as follows: R-2-acetylamino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid, N-acetyl-S-[1-(5-acetylamino-5-carboxypentyl)-1H-pyrrol-3-yl]-l-cysteine, and its sulfoxide. It was postulated that these three metabolites are derived from degraded protein adducts. However, the possibility that these metabolites result from the reaction of BDA with free lysine and/or cysteine was not ruled out. In this latter case, one might predict that the reaction of thiol-BDA with free lysine would not occur exclusively on the epsilon-amino group. Reaction of BDA with N-acetylcysteine or GSH in the presence of lysine indicated that both the alpha- and the epsilon-amino groups of lysine can be modified by thiol-BDA. The N-acetylcysteine-BDA-N-acetyllysine urinary metabolites were solely linked through the epsilon-amino group of lysine. A GSH-BDA-lysine cross-link was a significant hepatocyte metabolite of furan. In this case, the major product resulted from reaction with the epsilon-amino group of lysine; however, small amounts of the alpha-amino reaction product were also observed. Western analysis of liver and hepatocyte protein extracts using anti-GSH antibody indicated that GSH was covalently linked to proteins in tissues or cells exposed to furan. Our data support the hypothesis that GSH-BDA can react with either free lysine or protein lysine groups. These data suggest that there are multiple pathways by which furan can modify cellular nucleophiles. In one pathway, BDA reacts directly with proteins to form cysteine-lysine reaction products. In another, BDA reacts with GSH to form GSH-BDA conjugates, which then react with cellular nucleophiles like free lysine or lysine moieties in proteins. Both pathways will give rise to N-acetyl-S-[1-(5-acetylamino-5-carboxypentyl)-1H-pyrrol-3-yl]-l-cysteine. Given the abundance of these metabolites in urine of furan-treated rats, these pathways appear to be major pathways of furan biotransformation in vivo.

Highly potent fentanyl analogs: apnea from exposure to small quantities of ß-hydroxyfentanyl and furanylfentanyl.

Context: Fentanyl analogs are synthetic opioids that bind to mu receptors with high potency. We report two cases where users developed apnea and required naloxone reversal after exposure to small quantities of the fentanyl analogs, furanylfentanyl and ß-hydroxyfentanyl. Case details: A 19 year-old woman reported insufflating a small number of granules of "fentanyl" powder and developed apnea. A 22 year-old woman touched her tongue to powdered "fentanyl" to treat a migraine headache and became apneic. Results: Furanylfentanyl was identified in the first patient's serum (3.6 ng/mL furanylfentanyl), urine (17.6 ng/mL furanylfentanyl) , and in the product (109mg/g powder). ß-hydroxyfentanyl was detected in the second patient's serum (6.5 ng/mL ß-hydroxyfentanyl) and in the product (120 mg/g powder). Discussion: In both cases, furanylfentanyl and ß-hydroxyfentanyl were identified in the serum/plasma and urine. Fentanyl analogs were detected in the product at 109-120mg per gram of powder, which suggests that a 500mcg dose of fentanyl analog is the equivalent to 4.17-4.59mg of powder, or about the mass in 1/500th of a packet of sugar. Unique aspects of these cases are the rarely reported use of furanylfentanyl and ß-hydroxyfentanyl, intentional insufflation of "fentanyl" for therapeutic and recreational use, the small quantities of powder ingested, and the use of non-targeted analysis to identify the fentanyl analogs.

Adrenic acid non-enzymatic peroxidation products in biofluids of moderate preterm infants.

Oxidative stress plays an essential role in processes of signaling and damage to biomolecules during early perinatal life. Isoprostanoids and isofuranoids from the free radical-catalyzed peroxidation of polyunsaturated fatty acids (PUFAs) are widely recognized as reliable biomarkers of oxidative stress. However, their quantification is not straightforward due to high structural similarity of the compounds formed. In this work, a semiquantitative method for the analysis of adrenic acid (AdA, C22:4 n-6) non-enzymatic peroxidation products (i.e. dihomo-isoprostanes and dihomo-isofurans) was developed. The proposed ultra-performance liquid chromatography - tandem mass spectrometry (UPLC-MS/MS) method was applied to the analysis of blood plasma and urine from preterm infants providing information about AdA peroxidation.

A Fatality Involving Furanylfentanyl and MMMP, with Presumptive Identification of Three MMMP Metabolites in Urine.

The prevalence of new psychoactive substances (NPS) on the illicit drug market continues to grow, with new analogs being routinely synthesized. Routes of administration for these compounds are also diversifying, and recent research has shown an increase in the incorporation of NPS into vaping liquids. Among the most commonly encountered NPS are the cathinone and fentanyl analogs. Fentanyl analogs in particular have been implicated in a significant number of deaths, usually in combination with other prescription and illicit drugs. We report the case of a 44-year-old male with a history of polysubstance abuse found deceased at his home address. Items located within the vicinity of the deceased were found to contain furanylfentanyl and 2-methyl-4'-(methylthio)-2-morpholinopropiophenone (MMMP also known as MTMP, MMTMP, Irgacure 907 and Caccure 907). Both of these compounds were detected in the post-mortem peripheral blood of the deceased: furanylfentanyl at 1.6 ng/mL and MMMP at 6.7 ng/mL. MMMP is an unrestricted, commercially available photo-initiator used in the printing and polymer industry, which structurally can be classed as a highly modified cathinone. Although MMMP has been found previously in drug seizures, this is the first fatality in which MMMP has been detected. A number of other prescription and illicit drugs were also detected in the blood. MMMP was not detected in the post-mortem urine; however three metabolites, beta-hydroxy-MMMP, beta-hydroxy-MMMP-sulfoxide and beta-hydroxy-MMMP-sulfone, were presumptively identified. The significance of MMMP to the cause of death is uncertain as its pharmacological and toxicological profile is unclear.

A high-performance liquid chromatography-tandem mass spectrometry method for the clinical combination study of carboplatin and anti-tumor agent eribulin mesylate (E7389) in human plasma.

An LC/MS/MS method was developed to quantify carboplatin and eribulin mesylate (E7389) in human plasma and urine. For carboplatin, sample clean-up by protein precipitation and supernatant injection into a Waters Spherisorb((R)) S5 SCX column was used. Liquid-phase extraction and reverse-phase chromatography on a Polaris C18 column were used for eribulin. Quantitation involved LC/MS/MS with positive electrospray ionization. Accuracy, precision, linearity, range, specificity, recovery and stability were also evaluated. Both compounds were stable in human plasma (>or=80 days at -80 degrees C), at room temperature (>or=4h), following three freeze-thaw cycles and in 50/50 methanol/H(2)O (<4 degrees C for >or=252 days).