Stable Isotope Dilution Analysis (SIDA) to Determine Metabolites of Furan and 2-Methylfuran in Human Urine Samples: A Pilot Study.

Furan and 2-methylfuran (2-MF) are food contaminants that are classified as potentially carcinogenic to humans. The main source of exposure for adults via food is coffee consumption. Furan and 2-MF are volatile, which complicates exposure assessment because their content measured in food prior to consumption does not afford a reliable dosimetry. Therefore, other ways of exposure assessment need to be developed, preferably by monitoring exposure biomarkers, e.g., selected metabolites excreted in urine. In this study, cis-2-buten-1,4-dial (BDA)-derived urinary furan metabolites Lys-BDA (l-2-amino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)hexanoic acid), AcLys-BDA (l-2-(acetylamino)-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)hexanoic acid) and GSH-BDA (N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-l-cysteinyl-glycine cyclic sulfide), as well as acetyl acrolein (AcA, 2-oxo-pent-2-enal)-derived metabolites Lys-AcA (l-2-(acetylamino)-6-(2,5-dihydro-5-methyl-2-oxo-1H-pyrrol-1-yl)-hexanoic acid) and AcLys-AcA (l-2-amino-6-(2,5-dihydro-5-methyl-2-oxo-1H-pyrrol-1-yl)-hexanoic acid) and their stable isotopically labeled analogs, were synthesized and characterized through NMR and MS, and a stable isotope dilution analysis (SIDA) with UPLC-ESI-MS/MS was established. As a proof of concept, urinary samples of a four-day human intervention study were used. In the frame of this study, ten subjects ingested 500 mL of coffee containing 0.648 µmol furan and 1.059 µmol 2-MF. Among the furan metabolites, AcLys-BDA was the most abundant, followed by Lys-BDA and GSH-BDA. Exposure to 2-MF via the coffee brew led to the formation of Lys-AcA and AcLys-AcA. Within 24 h, 89.1% of the ingested amount of furan and 15.4% of the ingested amount of 2-MF were detected in the urine in the form of the investigated metabolites. Therefore, GSH-BDA, Lys-BDA, AcLys-BDA, Lys-AcA and AcLys-AcA may be suitable as short-term-exposure biomarkers of furan and 2-MF exposure.

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.

Urinary Excretion of Niacin Metabolites in Humans After Coffee Consumption.

SCOPE: Coffee is a major natural source of niacin in the human diet, as it is formed during coffee roasting from the alkaloid trigonelline. The intention of our study was to monitor the urinary excretion of niacin metabolites after coffee consumption under controlled diet. METHODS AND RESULTS: We performed a 4-day human intervention study on the excretion of major niacin metabolites in the urine of volunteers after ingestion of 500 mL regular coffee containing 34.8 µmol nicotinic acid (NA) and 0.58 µmol nicotinamide (NAM). In addition to NA and NAM, the metabolites N1 -methylnicotinamide (NMNAM), N1 -methyl-2-pyridone-5-carboxamide (2-Py), and nicotinuric acid (NUA) were identified and quantified in the collected urine samples by stable isotope dilution analysis (SIVA) using HPLC-ESI-MS/MS. Rapid urinary excretion was observed for the main metabolites (NA, NAM, NMNAM, and 2-Py), with tmax values within the first hour after ingestion. NUA appeared in traces even more rapidly. In sum, 972 nmol h-1 of NA, NAM, NMNAM, and 2-Py were excreted within 12 h after coffee consumption, corresponding to 6% of the ingested NA and NAM. CONCLUSION: The results indicate regular coffee consumption to be a source of niacin in human diet.