Solid-phase extraction techniques for the determination of glycopyrrolate from equine urine by liquid chromatography—tandem mass spectrometry and gas chromatography—mass spectrometry

• Published in: Journal of chromatography Vol. 573; no. 1; pp. 43 - 48
• Main Authors: Matassa, L.C., Woodard, D., Leavitt, R.K., Firby, P., Beaumier, P.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 03.01.1992; Elsevier
• Subjects: Analysis of complex biological substances; Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Blood and biological fluids; Chromatography, Ion Exchange; Chromatography, Liquid; Fundamental and applied biological sciences. Psychology; Gas Chromatography-Mass Spectrometry; Glycopyrrolate - urine; Horses - urine; Indicators and Reagents; Mass Spectrometry; Reference Standards; Quaternary ammonium compound; Vertebrata; Mammalia; Bronchodilator; Horse; Doping; Perissodactyla; Detection; Ungulata
• ISSN: 0378-4347
• DOI: 10.1016/0378-4347(92)80472-3

Glycopyrrolate (Robinul) is a quaternary ammonium salt which serves as a respiratory enhancing drug. It is reportedly used in horse racing to improve breathing. Extraction of glycopyrrolate from equine urine employing unique solid-phase extraction techniques gave a residue suitable for liquid chromatography—tandem mass spectrometry (LC—MS—MS) and gas chromatography—mass spectrometry (GC—MS). LC—MS—MS analysis employed an extract derived from 5 ml of urine subjected to cation-exchange chromatography. The daughter ion of m/z 318 monitored in the positive-ion mode was m/z 116. Recovery of glycopyrrolate was 99.5% and the within-run coefficient of variation of two quality control samples (1.0 and 10 ng/ml) was less than 5%. The between-run coefficient of variation for the same two quality control samples was less than 6.5%. The minimal detectable concentration for the assay was 250 pg/ml. Due to the extremely low concentration of glycopyrrolate in urine, qualitative detection via full-scan GC—MS required XAD-2 extraction of 50 ml of urine, cation-exchange chromatography clean-up and a tandem hydrolysis—derivatization procedure. The target analyte for GC—MS qualitative analysis was the methyl ester of hydrolyzed glycopyrrolate. Glycopyrrolate could be detected in post-administration (1 mg intravenously) urine samples for up to 9 h by both LC—MS—MS and GC—MS. The success of the method was due to a combination of the extreme sensitivity of the LC—MS—MS method and the very selective extraction process for quaternary ammonium salts.