Determination of a new non-benzodiazepine anxiolytic and its O-demethyl metabolite in plasma by high-performance liquid chromatography using automated column-switching.
An highly sensitive and fully automated high-performance liquid chromatographic assay was developed for the determination of a novel non-benzodiazepine anxiolytic (I) [(R)-2-(methoxymethyl)-1-[(7-oxo-8-phenyl-7H-thieno[2,3-a]quinolizin+ ++- 10-yl)carbonyl]pyrrolidine] and its O-demethyl metabolite (II) in plasma, using column-switching for direct injection of plasma samples. After dilution in internal standard solution, the sample was injected onto a pre-column (17 mm x 4.6 mm) dry-packed with pellicular C18 reversed-phase material. Polar plasma components were removed by flushing the pre-column with water-acetonitrile (90:10, v/v). Retained substances, including I and II, were backflushed onto an analytical column, separated by gradient elution and detected by means of fluorescence detection (excitation, 304 nm; emission, 475 nm). After washing the analytical column and re-equilibrating the pre-column, the system was ready for the next injection. The limit of quantification for I and II was 0.25 and 0.5 ng/ml, respectively, using a 350-microliter specimen of plasma. The practicability of the new method was demonstrated by analysis of more than 300 plasma samples from a tolerance study performed with human volunteers. Owing to its high sensitivity, the method can be used to calculate pharmacokinetic parameters of compounds I and II in man after a single oral dose of about 1 mg of I.
Timm
U
U
Pharmaceutical Research Department, F. Hoffmann-La Roche & Co. Ltd., Basle, Switzerland.
Hopfgartner
G
G
Erdin
R
R
D014151:Anti-Anxiety Agents / Q000097:blood*; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D011759:Pyrrolidines / Q000097:blood* / Q000378:metabolism; D011807:Quinolizines / Q000097:blood* / Q000378:metabolism
D014151:Anti-Anxiety Agents; D011759:Pyrrolidines; D011807:Quinolizines; C059292:2-(hydroxymethyl)-1-((7-oxo-8-phenyl-7H-thieno(2,3-a)quinolizin-10-yl)carbonyl)pyrrolidine; C059291:Ro 195686