An LC-MS/MS assay for the quantitative determination of 2-pyridyl acetic acid, a major metabolite and key surrogate for betahistine, using low-volume human K₂EDTA plasma

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• 作者：Krunal Soni ; Chandrakant Bhatt ; Kanchan Singh ; P.C. Bhuvaneshwari ; Anil Jha ; Palak Patel ; Harilal Patel and Nuggehally R. Srinivas
• 刊名：Biomedical Chromatography
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：31
• 期：2
• 全文大小：425K
• ISSN：1099-0801

文摘

Betahistine is widely used for the treatment of vertigo. Owing to first-pass metabolism, 2-pyridyl acetic acid (2PAA, major metabolite of betahistine) was considered as surrogate for quantitation. A specific and sensitive LC–MS/MS method was developed and validated for quantitation of 2PAA using turbo-ion spray in a positive ion mode. A solid-phase extraction was employed for the extraction of 2PAA and 2PAA d₆ (IS) from human plasma. Chromatographic separation of analytes was achieved using an ACE CN, 5&thinsp;μm (50&thinsp;×&thinsp;4.6&thinsp;mm) column with a gradient mobile phase comprising acetonitrile–methanol (90:10% v/v) and 0.7% v/v formic acid in 0.5&thinsp;mpan class="smallCaps">mpan> ammonium trifluoroacetate in purified water (100% v/v). The retention times of 1.15 and 1.17&thinsp;min for 2PAA and internal standard, respectively, were achieved. Quantitation of 2PAA and internal standard was achieved by monitoring multiple reaction monitoring transition pairs (m/z 138.1 to m/z 92.0 and m/z 142.1 to m/z 96.1, respectively). The developed method was validated for various parameters. The calibration curves of 2PAA showed linearity from 5.0 to 1500&thinsp;ng/mL, with a lower limit of quantitation of 5.0&thinsp;ng/mL. The bias and precision for inter- and intra-batch assays were <10%. The developed method was used to support clinical sample analysis.