Development and validation of a liquid chromatography-tandem mass spectrometry method for the determination of Febuxostat in human plasma

Zhejiang University of Technology, Chaowang Road, Hangzhou, 310014, People's Republic of China
Biomedical Chromatography (Impact Factor: 1.72). 01/2013; 27(1). DOI: 10.1002/bmc.2744
Source: PubMed


A rapid and sensitive analytical method based on liquid chromatography coupled to tandem mass spectrometry detection with positive ion electrospray ionization was developed for the determination of febuxostat in human plasma using d(7) -febuxostat as the internal standard (IS). A simple protein precipitation was performed using acetonitrile. The analyte and IS were subjected to chromatographic analysis on a Capcell PAK C(18) column (4.6 × 100 mm, 5 µm) using acetonitrile-5 mm ammonium acetate-formic acid (85:15:0.015, v/v/v) as the mobile phase at a flow rate of 0.6 mL/min. An Agilent 6460 electrospray tandem mass spectrometer was operated in the multiple reaction monitoring mode. The precursor-to-product ion transitions m/z 317 → m/z 261 (febuxsotat) and m/z 324 → m/z (261 + 262) (d(7) -febuxostat, IS) were used for quantitation. The results were linear over the studied range (10.0-5000 ng/mL), and the total analysis time for each chromatograph was 3 min. The intra- and inter-day precisions were less than 7.9 and 7.2%, respectively, and the accuracy was within ±4.2%. No evidence of analyte instability in human plasma was observed storage at -20°C for 31 days. This method was successfully applied in the determination of febuxostat concentrations in plasma samples from healthy Chinese volunteers. Copyright © 2012 John Wiley & Sons, Ltd.

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    • "67M-1, 67M-2 and 67M-4 have been shown to inhibit XO with similar potency to febuxostat and are also important indexes for characterizing the pharmacokinetics of febuxostat in humans [6]. HPLC with fluorescence detection (HPLC-FD) [7] [8] and LC–MS/MS [9] [10] [11] [12] methods have been previously developed to determine febuxostat in human plasma for pharmacokinetic study of febuxostat. The quantification of metabolites 67M-1, 67M-2 and 67M-4 in human plasma has been accomplished by two groups of researchers using LC–MS/MS methods [6] [8]. "
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    ABSTRACT: A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of febuxostat and its three active metabolites in human plasma was developed using a ZORBAX SB-C18 column (50mm×4.6mm, 5μm) and an optimized gradient mobile phase consisting of acetonitrile, water and formic acid. Plasma samples were spiked with the internal standard losartan and then pre-treated using one-step protein precipitation with methanol. Mass spectrometric detection was performed by selective reaction monitoring mode via electrospray ionization source operating in positive ionization mode. The method exhibited good linearity over the concentration range of 10-20,000ng/mL for febuxostat, 1.0-270ng/mL for 67M-1 and 67M-2, and 0.8-250ng/mL for 67M-4, respectively. The intra- and inter-day precisions were less than 14.7% and the accuracy ranged from -4.3% to 5.1%. The method was successfully applied to a clinical pharmacokinetic study of febuxostat in humans after oral administration of a single dose of febuxostat at 40, 80 and 120mg. Copyright © 2015 Elsevier B.V. All rights reserved.
    Full-text · Article · Oct 2015 · Journal of pharmaceutical and biomedical analysis
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    ABSTRACT: Febuxostat is a selective inhibitor of xanthine oxidase that is used for the treatment of hyperuricaemia in patients with gout. An isocratic liquid chromatographic method was developed and validated for the determination of febuxostat. Chromatographic separation was achieved on a C18 column using sodium acetate buffer (pH 4.0)–acetonitrile (40:60, v/v), with a flow rate 1.2 mL/min (ultraviolet detection at 254 nm). Linearity was observed in the concentration range of 0.1–200 µg/mL (R2 = 0.9999) with a linear regression equation of y = 21148x – 2025.1. The limit of quantification was found to be 0.0783 µg/mL and the limit of detection was found to be 0.0257 µg/mL. Febuxostat was subjected to stress conditions of degradation in aqueous solutions, including acidic, alkaline, oxidation, photolysis and thermal degradation. The forced degradation studies were performed by using sodium hydroxide, hydrochloric acid, hydrogen peroxide and thermal and ultraviolet radiation. Febuxostat is more sensitive toward acidic conditions than oxidation and very resistant toward alkaline, thermal and photolytic degradations. The method was validated as per the guidelines of the International Conference on Harmonization. The intra-day and inter-day precision (relative standard deviation) was found to be 0.29–0.41 and 0.63–0.76, respectively. The method is simple, specific, precise, robust and accurate for the determination of febuxostat in pharmaceutical dosage forms (tablets).
    Full-text · Article · Dec 2012 · Journal of chromatographic science
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    ABSTRACT: An improved, simple and highly sensitive LC-MS/MS method has been developed and validated for quantification of febuxostat with 100 μL human plasma using febuxostat-d7 as an internal standard (IS) according to regulatory guidelines. The analyte and IS were extracted from human plasma via liquid-liquid extraction using diethyl ether. The chromatographic separation was achieved on a Zorbax C18 column using a mixture of acetonitrile and 5 mm ammonium formate (60:40, v/v) as the mobile phase at a flow rate of 0.5 mL/min. The total run time was 5.0 min and the elution of febuxostat and IS occurred at 1.0 and 1.5 min, respectively. A linear response function was established for the range of concentrations 1-6000 ng/mL (r > 0.99). The precursor to product ion transitions monitored for febuxostat and IS were m/z 317.1 → 261.1 and 324.2 → 262.1, respectively. The intra- and inter-day precisions (%RSD) were within 1.29-9.19 and 2.85-7.69%, respectively. The proposed method was successfully applied to pharmacokinetic studies in humans. Copyright © 2013 John Wiley & Sons, Ltd.
    No preview · Article · Nov 2013 · Biomedical Chromatography
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