Simultaneous quantification of methylene blue and its major metabolite, azure B, in plasma by LC-MS/MS and its application for a pharmacokinetic study

• Published in: Biomedical chromatography Vol. 28; no. 4; pp. 518 - 524
• Main Authors: Kim, Soo-Jin, Ha, Dong-Jin, Koo, Tae-Sung
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01.04.2014
• Subjects: Administration, Oral; Animals; azure B; Azure Stains - analysis; Azure Stains - chemistry; Azure Stains - pharmacokinetics; Blood Chemical Analysis; Chromatography, Liquid - methods; LC/MS/MS; Linear Models; Male; methylene blue; Methylene Blue - analysis; Methylene Blue - chemistry; Methylene Blue - pharmacokinetics; pharmacokinetics; rat plasma; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Sensitivity and Specificity; Tandem Mass Spectrometry - methods; azure B; pharmacokinetics; methylene blue; LC/MS/MS; rat plasma
• ISSN: 0269-3879; 1099-0801
• DOI: 10.1002/bmc.3063

ABSTRACT A simple and sensitive liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) method was developed for the quantification of methylene blue (MB) and its major metabolite, azure B (AZB), in rat plasma. A simple protein precipitation using acetonitrile was followed by injection of the supernatant on to a Zorbax HILIC Plus column (3.5 µm, 2.1 × 100 mm) with isocratic mobile phase consisting of 5 mM ammonium acetate in 10:90 (v/v) water:methanol at a flow rate of 0.3 mL/min and detection in positive ionization mode. The standard curve was linear over the concentration range from 1 to 1000 ng/mL for MB and AZB with coefficient of determination above 0.9930. The lower limit of quantification was 1 ng/mL using 20 μL of rat plasma sample. The intra‐ and inter‐assay precision and accuracy were <12%. The developed analytical method was successfully applied to the pharmacokinetic study of MB and AZB in rats. Copyright © 2013 John Wiley & Sons, Ltd.