Z. Yu, D. Westerlund and K. S. Boose, “Determination of Methotrexate and Its Metabolite 7-Hydroxymethotrexate by Direct Injection of Human Plasma into a Column-Switching Liquid Chromatographic System Using Post-Column Photochemical Reaction with Fluorimetric Detection,” Journal of Chromatography B, Vol. 689, No. 2, 1997, pp. 379-386. Hdoi:10.1016/S0378-4347(96)00357-X

has been cited by the following article:

• TITLE: Coupling of on-Line Pre-Column Oxidative Cleavage and Solid-Phase Enrichment with Liquid Chromatography Using an Eco-Friendly Analytical Procedure to Determine Low Levels of Methotrexate

  AUTHORS: Samy Emara, Walaa Zarad, Maha Kamal, Ramzia EL-Bagary

  KEYWORDS: Cerium (IV) Trihydroxyhydroperoxide; HPLC; Methotrexate; ODS Analytical Column; On-Line Pre-Column Oxidative Cleavage

  JOURNAL NAME: Journal of Analytical Sciences, Methods and Instrumentation, Vol.2 No.4, December 25, 2012

  ABSTRACT: A simple, sensitive and precise green high-performance liquid chromatographic method including on-line pre-column oxidation combined by column switching with a short Hypersil ODS analytical column (100 mm × 4.0 mm i.d.) for enrichment and separation was developed and validated to determine low levels of methotrexate (MTX). The method was based on oxidative cleavage of MTX into highly fluorescence products, 2,4-diaminopteridine-6-carboxaldehyde and the corresponding 2,4-diaminopteridine-6-carboxylic acid, during the flow of phosphate buffer (0.04 M, pH 3.4) containing the analyte through the packed reactor of cerium (IV) trihydroxyhydroperoxide (CTH) at a flow-rate of 0.2 mL/min and 40℃. The fluorescent products were enriched on the head of ODS analytical column for the final separation. The separation was performed at room temperature using an environmentally friendly mobile phase consisting of ethanol and phosphate buffer (0.04 M, pH 3.4) in the ratio of 10:90 (v/v). The eluent was monitored at emission and excitation wavelengths of 463 and 367 nm, respectively. The method was successfully applied, without any interference from the excipients, for the determination of drug in tablets and vials with a detection limit of 0.06 ng/mL from 500 ?L of sample MTX.