Determination of memantine in plasma and vitreous humour by HPLC with precolumn derivatization and fluorescence detection.

Belen Puente, Esther Hernandez, Susana Perez, Luis Pablo, Esther Prieto, Maria Angeles Garcia, Miguel Angel Bregante

Index: J. Chromatogr. Sci. 49(10) , 745-52, (2011)

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Abstract

A new HPLC procedure with precolumn derivatization and rimantadine as the internal standard for determining memantine, a candidate agent for the treatment of glaucoma in plasma and vitreous humour, has been developed and validated. Precolumn derivatization was performed with 9-fluorenylmethyl-chloroformate-chloride (FMOC-Cl) as the derivatization reagent and followed by a liquid-liquid extraction with n-hexane. Optimal conditions for derivatization were an FMOC-Cl concentration of 1.5 mM, a reaction time of 20 min, the temperature at 30°C, the borate buffer pH 8.5, and a borate buffer-acetonitrile ratio of 1:1. The derivatives were analyzed by isocratic HPLC with the fluorescence detector λex 260 nm λem 315 nm on a Novapack C(18) reversed-phase column with a mobile phase of acetonitrile-water (73:27, v/v), 40°C, and a flow rate of 1.2 mL/min. The linear range was 10-1000 ng/mL with a quantification limit of ∼ 10 ng/mL for both types of samples. This analytical method may be suitable for using in ocular availability studies.