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Abstract: Carvedilol is widely prescribed for the treatment of hypertension, heart failure and left ventricular dysfunction following myocardial infarction. A sensitive and reliable liquid chromatography–tandem mass spectrometry (LC–MS/MS) assay was developed and validated to enable reliable and efficient bioanalysis of the (R)- and (S)-enantiomers of carvedilol and its pharmacologically active 4′-hydroxyphenyl metabolite in human plasma. Following plasma extraction using supported liquid extraction (SLE) in a 96-well plate format, extracted samples were derivatized with 2,3,4,6-tetra-O-acetyl-β-d-glucopyranosyl isothiocyanate (GITC). Chromatographic separation was achieved by gradient elution on an ACQUITY UPLC HSS T3 analytical column. The impact of several potentially interfering isobaric metabolites on the quantification of the 4′-hydroxyphenyl metabolite (R)- and (S)-enantiomers was minimized by implementation of a combination of chromatographic and mass spectrometric techniques. Derivatized analytes and stable-labeled internal standards were detected by positive ion electrospray tandem mass spectrometry. The assay was validated over concentration ranges of 0.200–100ng/mL for (R)- and (S)-carvedilol and 0.0200–10.0ng/mL for (R)- and (S)-4′-hydroxyphenyl carvedilol. Intra- and inter-assay precision values for replicate quality control samples were within 11.9% for all analytes during the assay validation. Mean quality control accuracy values were within ±9.4% of nominal values for all analytes. Assay recoveries were high (>76%) and internal standard normalized matrix effects were minimal. The four analytes were stable in human plasma for at least 24h at room temperature, 89 days at −20°C and −70°C, and following at least five freeze–thaw cycles. The validated assay was successfully applied to the quantification of the (R)- and (S)-enantiomers of both carvedilol and its pharmacologically active 4′-hydroxyphenyl metabolite in human plasma in support of a human pharmacokinetic study. [Copyright &y& Elsevier]

Abstract: A sensitive method was developed and validated for the measurement of ixabepilone (BMS-247550, Ixempra™) using a demethylated analogue of ixabepilone (BMS-212188) as an internal standard. A 0.050mL portion of each plasma sample was extracted with 0.450mL of acetonitrile containing the internal standard via protein precipitation. The supernatant was analyzed on a LC–MS/MS system. Chromatography was carried out on a 2.0mm×100mm YMC ODS-AQ 3μm column using an isocractic mobile phase consisting of acetonitrile:10mM ammonium acetate, pH 5.0 (70:30, v/v) at a flow rate of 0.30mL/min. The mass spectrometer was fitted with a TurboIonSpray® source and operated in negative ionization mode. Detection of ixabepilone and BMS-212188 were accomplished using multiple reaction monitoring (MRM) of precursor>product ion pairs of m/z 505.2>405.2, and 492.1>392.1, respectively. The assay range was 2.00–500ng/mL and was fitted to a 1/x 2 weighted quadratic regression model. Replicate sample analysis indicated that intra- and inter-day accuracy and precision are within ±15.0%. The recovery of ixabepilone from 0.050mL of plasma containing 5.00 and 400ng/mL was greater than 94%. The method was demonstrated to be sensitive, selective and robust, and was successfully used to support clinical studies. This paper also discussed approaches used for resolving a curve splitting issue observed during quantitative analysis of ixabepilone in biological matrices. Finally, to adapt the methodology to pharmacokinetics of ixabepilone after oral administration, the potential interference of chemical degradants on the determination of ixabepilone was evaluated. [Copyright &y& Elsevier]

Clopidogrel is prescribed for the treatment of Acute Coronary Syndrome and recent myocardial infarction, recent stroke, or established peripheral arterial disease. A sensitive and reliable high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) assay was developed and validated to enable reliable quantification of four diastereomeric and chemically reactive thiol metabolites, two of which are pharmacologically active, in human plasma. The metabolites were stabilized by alkylation of their reactive thiol moieties with 2-bromo-3′-methoxyacetophenone (MPB). Following organic solvent mediated-protein precipitation in a 96-well plate format, chromatographic separation was achieved by gradient elution on an Ascentis Express RP-amide column. Chromatographic conditions were optimized to ensure separation of the four derivatized active metabolites. Derivatized metabolites and stable isotope-labeled internal standards were detected by positive ion electrospray tandem mass spectrometry. The HPLC–MS/MS assay was validated over concentration ranges of 0.125–125ng/mL for metabolites H1–H3 and 0.101–101ng/mL for H4. Intra- and inter-assay precision values for replicate quality control samples were within 14.3% for all analytes during the assay validation. Mean quality control accuracy values were within ±6.3% of nominal values for all analytes. Assay recoveries were high (>79%). The four derivatized analytes were stable in human blood for at least 2h at room temperature and on ice. The analytes were also stable in human plasma for at least 25h at room temperature, 372 days at −20°C and −70°C, and following at least five freeze–thaw cycles. The validated assay was successfully applied to the quantification of all four thiol metabolites in human plasma in support of a human pharmacokinetic study. [ABSTRACT FROM AUTHOR]