Quantitative determination of glycopyrrolate in human plasma by liquid chromatography–electrospray ionization mass spectrometry: The use of a volatile ion-pairing agent during both liquid–liquid extraction and liquid chromatography

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Vol. 876; no. 1; pp. 24 - 30
• Main Authors: Storme, M.L., t’Kindt, R.S., Goeteyn, W., Reyntjens, K., Van Bocxlaer, J.F.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.12.2008; Elsevier
• Subjects: Analysis; Analytical, structural and metabolic biochemistry; Biological and medical sciences; Chemical Fractionation - methods; Chromatography, High Pressure Liquid - methods; Fundamental and applied biological sciences. Psychology; General pharmacology; Glycopyrrolate; Glycopyrrolate - blood; Humans; LC-MS; LC–MS/MS; Medical sciences; Pharmacokinetics; Pharmacology. Drug treatments; Reproducibility of Results; Spectrometry, Mass, Electrospray Ionization - methods; Tandem Mass Spectrometry - methods; Pharmacokinetics; LC–MS/MS; Glycopyrrolate; Volatile compound; Human; Biological fluid; Organic cation; Liquid liquid extraction; Ion pair; HPLC chromatography; Electrospray; Blood plasma; Quaternary ammonium compound; LC-MS/MS; Cholinergic receptor; Glycopyrronium bromide; Mass spectrometry MS/MS; Muscarinic receptor; Antagonist; Mass spectrometry; Quantitative analysis
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2008.10.013

The work presented here deals with the development of a quantitative tool for the determination of the quaternary ammonium anticholinergic glycopyrrolate in human plasma samples. Mepenzolate was used as an internal standard. The plasma samples were subjected to a suitable sample clean-up consisting of a simple and relatively fast, two step liquid–liquid ion-pair extraction procedure. The chromatography, using the same volatile ion-pair reagent heptafluorobutyric acid (HFBA), takes only 10 min. Relative standard deviation of retention times was never above 2.26% ( n = 36). The method was fully validated based on the US FDA Bioanalytical Method Validation Guidance for Industry. As such, a quantitative ESI-LC–MS(/MS) (TOF mass spectrometry) method was optimized for the absolute quantification of glycopyrrolate in human plasma in a concentration range from 0.101 to 101 ng/mL using a quadratic calibration function ( R 2 = 0.9995), y = −2.21 × 10 −4 (±3.93 × 10 −5) × x 2 + 5.85 × 10 −2 (±5.27 × 10 −3) × x + 4.08 × 10 −3 (±4.82 × 10 −4). For the three QC concentrations (QC 1 0.252, QC 2 2.52, and QC 3 25.2 ng/mL) and the LLOQ (0.101 ng/mL), total precision was under 20% (18.0% ( n = 6) at the LLOQ) and maximum accuracy was 112% (88.9% for the LLOQ, n = 6). Absolute matrix effect (maximum 133% ± 9.59, n = 3), absolute recovery (better than 41.8% ± 2.22, n = 3), relative (inter-subject) matrix effect (maximum 10.9% ± 1.45, n = 4) and process efficiency (better than 45.2% ± 5.74, n = 3) too were assessed at the 3 QC concentrations.