title: Determination of methotrexate and indomethacin in urine using SPE-LC-DAD after derivatization.
abstract: A validated HPLC-DAD assay is presented for the simultaneous quantification of methotrexate and indomethacin in a drug combination which is used synergistically to intervene with tumoral or inflammatory tissue microenvironments. The analytes were isolated from urine via solid phase extraction. The method is based on derivatizing both analytes with a soluble carbodiimide coupler and 2-nitrophenylhydrazine directed to their commonly occurring carboxylate functions. The chromatographic separation was accomplished on an octylsilica column in less than 15 min using acetate buffer (pH 4; 10 mM)-methanol (60:40, v/v) as eluent at 1.5 ml/min and monitored at 400 nm. The selectivity of the method was demonstrated in a pre-dosed rheumatoid arthritis patient. Quality control samples were prepared and analyzed to reveal the validity of the method. Life samples collected from a healthy volunteer were monitored for both drugs and their urinary levels were determined.
authors.affiliation: Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Alexandria, Khartoum Square 1, Azarita 21521, Alexandria, Egypt. kmichail@pharmacy.ualberta.ca
authors.forename: Karim
authors.initials: K
authors.lastname: Michail
authors.forename: Marwa S
authors.initials: MS
authors.lastname: Moneeb
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000652:urine; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005493:Folic Acid Antagonists / Q000652:urine; D006801:Humans; D007213:Indomethacin / Q000652:urine*; D057230:Limit of Detection; D008727:Methotrexate / Q000652:urine*; D011786:Quality Control; D012015:Reference Standards; D052616:Solid Phase Extraction / Q000379:methods*
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D005493:Folic Acid Antagonists; D007213:Indomethacin; D008727:Methotrexate