title: Rapid quantitative determination of a collagenase inhibitor and its major metabolite by on-line liquid chromatography with ionspray tandem mass spectrometric detection.
abstract: A rapid, sensitive and specific analytical method has been developed and validated to quantify the collagenase inhibitor N2-(2(5)-[(hydroxycarbamoyl)methyl)-4-methylvaleryl]-N1, 3-dimethyl-L-valin-amide (I) and its major metabolite (II) from plasma and urine. This method employs an automated solid-phase extraction procedure to isolate the analytes and the internal standard from the biological matrix. Reconstituted extracts were analyzed by HPLC-ionspray MS-MS. Chromatography was performed on a 4.6 mm I.D. reversed-phase guard column. The retention times of the analytes and the internal standard were approximately 1.3 min. The assay has a limit of quantification of 5 ng/ml plasma and a limit of detection of 1 ng/ml, based on 100-micro l plasma aliquots. No sample-drying step was required. The standard curves were linear form 5 to 5000 ng/ml using weighted linear regression analysis (1/y2). The intra- and inter-assay precision were better than + or - 10% with intra- and inter-assay accuracies between 95 and 105%. this new HPLC-MS-MS assay procedure for I and II in plasma and urine has proven to be specific, sensitive, accurate and robust, and is being used routinely for the analysis of I in pre-clinical and clinical trial samples. Up to 200 unknowns may be analyzed each 24 h per analyst.
authors.affiliation: Department of Pharmacokinetics and Metabolism, Roche Products Ltd., Hertfordshire, UK.
authors.forename: N G
authors.initials: NG
authors.lastname: Knebel
authors.forename: S R
authors.initials: SR
authors.lastname: Sharp
authors.forename: M J
authors.initials: MJ
authors.lastname: Madigan
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000706:statistics & numerical data; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000097:blood / Q000652:urine; D006801:Humans; D006877:Hydroxamic Acids / Q000032:analysis* / Q000097:blood / Q000652:urine; D013058:Mass Spectrometry / Q000379:methods* / Q000706:statistics & numerical data; D061965:Matrix Metalloproteinase Inhibitors; D011786:Quality Control; D012680:Sensitivity and Specificity; D014633:Valine / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood / Q000652:urine
chemical_list: D004791:Enzyme Inhibitors; D006877:Hydroxamic Acids; D061965:Matrix Metalloproteinase Inhibitors; C099123:N2-(2-((hydroxycarbamoyl)methyl)-4-methylvaleryl)-N1,3-dimethylvalinamide; C099124:N2-(2-(carbamoylmethyl)-4-methylvaleryl)-N1,3-dimethylvalinamide; D014633:Valine