Variability amongst urine toxicology amphetamine readings with concurrent administration of fenofibrate.

OBJECTIVE: The aim of this study was to highlight that concurrent administration of the common lipid-lowering agent fenofibrate may lead to false-positive amphetamine results in often-used immunoassay-based urine drug screens. It also aimed to show that there are significant moral and clinical challenges associated with the interpretation of such results amongst psychiatric inpatients. CONCLUSIONS: It is evident that different pathology laboratories may utilise different commercial urine drug-screen immunoassays in their toxicology analysis, with variability in the test specificities. Despite the relatively high prevalence of substance misuse in the population of psychiatric inpatients, there exists a need for increased vigilance towards the possibility of false-positive amphetamine results owing to likely cross-reactivity of fenofibrate with the test reagents. In cases where there is uncertainty when correlating clinically, or where false positives are suspected, gold-standard urine-sample analysis by mass spectrometry should be considered, particularly when the consequences for patients may include restrictive measures.

A Cross-Reactivity of Fenofibric Acid With MDMA DRI Assay.

BACKGROUND: Within the framework of routine fitness examinations, French Air Force military crew underwent urine testing for 3,4 methylenedioxymetamphetamine (MDMA [ecstasy]). The cross-reactivity of a dyslipidemic drug, fenofibrate, with an MDMA immunoassay was studied and confirmed on a large population sample. METHODS: A 3-year retrospective study was performed on the MDMA DRI Ecstasy Assay on the Unicel DXC 600. In the event of positive test result, a confirmatory testing was carried out by gas chromatography/mass spectrometry (GC/MS) to establish the presence of MDMA. When analysis by GC/MS did not confirm the presence of MDMA, a false-positive result was suspected and the samples were analyzed by high-performance liquid chromatography-mass spectrometry to identify a potential interfering substance. RESULTS: A total of 15,169 urine samples, from 7,803 patients, were tested for 3 years. Of the tested samples, 22 (0.15%) were positive by DRI Ecstasy Assay. None of them were positive by GC/MS. A cross-reactivity of fenofibrate's metabolite with MDMA using this assay was systematically found. CONCLUSION: Fenofibrate's interference with MDMA immunoassay was confirmed. Fenofibrate being widely prescribed, physicians had to be alerted that this treatment could lead to false-positive results.

Fenofibric Acid Can Cause False-Positive Urine Methylenedioxymethamphetamine Immunoassay Results.

We present a false-positive result of ecstasy (3,4-methylenedioxy-NN-methylamphetamine) screening due to the therapeutic use of fenofibrate, an antihyperlipidemic drug. Our hypothesis was that the main metabolite of fenofibrate, fenofibric acid, was responsible for this cross-reactivity on a DRI(®) Ecstasy Assay, using a cut-off of 500 ng/mL. We estimated that the addition of 225 µg/mL pure fenofibric acid to blank urine would be sufficient to result in a positive DRI(®) Ecstasy Assay. The results obtained on the urine samples analyses of the patient show that the DRI(®) Ecstasy Assay resulted negative 2 days after discontinuing fenofibrate treatment, when the urine fenofibric acid concentration corrected by creatinine and determinated by gas chromatography-mass spectrometry was 20.3 µg/mg creatinine. The cross-reactivity data for fenofibric acid would seem to indicate that there was insufficient concentration of measured compound to account for the positive immunochemical results for ecstasy. This apparent discrepancy can be explained in several ways, one of them is that the ß-glucuronidase-resistent fenofibric acid isomers are responsible. This process could explain the low recovery of free fenofibric acid when we use the developed method for its quantification in urine samples. Positive results on immunoassay screening must be considered presumptive until confirmation with another method based on a different principle, preferably gas chromatography-mass spectrometry or liquid chromatography-mass spectrometry.

Development of a LC-MS/MS method for the estimation of clinofibrate in human urine.

A highly sensitive and rapid liquid chromatography-tandem mass spectrometry method was developed for the determination of clinofibrate in human urine. The analyte and IS were extracted through a simple protein precipitation by the mixture of acetonitrile and 1 mol/L hydrochloric acid (95:5, v/v) and separated on an Inspire C18 (150 mm x 4.6 mm I.D., 5 µm particle size) column using isocratic elution with methanol and water containing 0.1% formic acid and 10 mM ammonium acetate (90:10, v/v). Mass spectrometric detection was performed in electrospray positive ionization MRM mode. The mass transition was m/z 486.3-->175.0 for clinofibrate and m/z 361.1-->233.1 for IS, respectively. The flow rate was 0.6 mL/min and the column oven temperature was set at 35 °C. The total run time was 6.5 min. Good linear relationships were obtained for all analytes over the concentrations ranging of 0.1002-10.02 µg/mL (r2 = 0.9991) and the limit of quantification was 0.1002 µg/mL. The extraction recovery was larger than 87.4% and intra- and inter-batch precision and accuracy with RSD were all less than 6.5%. The total amount of unchanged clinofibrate excreted in urine was less than 0.34%. This method was successfully applied to the pharmacokinetic study of clinofibrate in human urine.

New metabolites of fenofibrate in Sprague-Dawley rats by UPLC-ESI-QTOF-MS-based metabolomics coupled with LC-MS/MS.

Fenofibrate has been widely used for the treatment of dyslipidaemia with a long history. Species differences of its metabolism were reported, but its metabolites in rodent have not been fully investigated. Urine and plasma samples were collected before and after oral dosages of fenofibrate in Sprague-Dawley rats. Urine samples were subjected to ultra-performance liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOF-MS) analysis, and projection to latent structures discriminant analysis was used for the identification of metabolites. New metabolites in urine and plasma were also studied by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The metabolism pathway was studied in rat hepatocytes. Synthesized and purchased authentic compounds were used for metabolite identification by LC-MS/MS. Five ever-reported metabolites were identified and another four new ones were found. Among these new metabolites, fenofibric acid taurine and reduced fenofibric acid taurine indicate new phase II conjugation pathway of fenofibrate.

[High-performance liquid chromatographic method for the determination of fenofibric acid and reduced fenofibric acid in human blood, plasma and urine].

In this study, a very reliable HPLC method was developed for the determination of fenofibric acid and reduced fenofibric acid in the biological samples described as follows. After addition of the internal standard solution and 0.5 M HCl to the biological sample, fenofibric acid, reduced fenofibric acid and the internal standard were extracted with a mixed solvent of n-hexane and ethyl acetate (90:10) from the mixture. The acids were back-extracted from the organic phase with 0.1 M Na2HPO4 and then re-extracted from the aqueous phase with a mixed solution of n-hexane and ethyl acetate (95:5) after addition of 0.5 M HCl. The organic phase was evaporated to dryness under the vacuum. The residue was dissolved in MeOH and diluted with distilled water. An aliquot of the resulting solution was injected on the HPLC. High reproducibility was observed in this HPLC method (C.V.% less than 4%). Moreover it was confirmed that the conjugates in the urine could be hydrolyzed by incubation at 37 degrees C for 18 h after addition of 400 IU of beta-glucuronidase.

Urinary glucuronide excretion of fenofibric and clofibric acid glucuronides in man. Is it polymorphic?

The possible polymorphism of the glucuronidation reaction in man has been investigated using two hypolipidaemic compounds, fenofibrate and clofibrate, as the test probes. The formation of fenofibryl and clofibryl glucuronides was identified by their susceptibility to hydrolyses by beta-glucuronidase. The urinary excretion of the glucuronides was measured in 72 healthy volunteers after a single dose of fenofibrate, and in 104 subjects given a single dose of clofibric acid. Fenofibrate was excreted at a lower rate than clofibrate, since 13.94% and 26.55% of the doses of fenofibrate and clofibrate respectively, were recovered in urine in 8 h. Correlation analysis indicated that sex and body mass index significantly influenced the formation of fenofibryl glucuronide, whereas age and oral contraceptives affected the excretion of clofibryl acid glucuronide. The 8-hour urinary excretion patterns of clofibryl glucuronide and of clofibric acid presented a Gaussian distribution, whereas those of fenofibryl glucuronide and fenofibric acid showed 2 populations. When the metabolic ratio free fenofibric acid/glucuronide was considered, 84.7% of subjects presented the ratio 0.147, and 15.3% had the 3-fold higher ratio of 0.421. The study has shown, in the human population studied, that the glucuronidation of fenofibric acid but not that of clofibric acid may present a polymorphism.

The metabolism and disposition of 14C-fenofibrate in human volunteers.

The metabolic fate of a single dose of 14C-fenofibrate has been studied in a panel of eight healthy volunteers (four males and four females). In 7 days, a total of 84% of the administered dose was recovered, with 59% in the urine and 25% in the feces. The majority of the urinary 14C was excreted within 24 hr, whereas the bulk of the fecal 14C was recovered over the first 3 days after dosing. The major urinary metabolite was the ester glucuronide of fenofibric acid, accompanied by much smaller amounts of fenofibric acid and the benzhydrol and its glucuronide. The principal compound in feces was unchanged fenofibrate, together with smaller quantities of fenofibric acid and polar unknown metabolite(s). Experiments on the stability of fenofibryl glucuronide showed it to be less reactive than most ester glucuronides.