Determination of L-dopa, carbidopa, 3-O-methyldopa and entacapone in human plasma by HPLC-ED

The aim of the study was the development of analytical methods suitable for the quantification of L-dopa, carbidopa and entacapone in plasma of Parkinsonian patients treated with Stalevo (R). The metabolite 3-O-methyldopa was also determined to obtain some indications on the pharmacokinetics of L-dopa. For the simultaneous analysis of L-dopa, 3-O-methyldopa and carbidopa, a RP 08 column as the stationary phase and a mixture of methanol and a pH 2.88 phosphate buffer (8:92, v/v) as the mobile phase were used. A feasible plasma pre-treatment based on protein precipitation was implemented, obtaining extraction yield higher than 94% for all the analytes. For the analysis of entacapone a RP C8 column and a mixture of methanol, acetonitrile and a pH 1.90 phosphate buffer as the mobile phase (17.5:22.5:60, v/v/v) were used. A plasma pre-treatment procedure was developed, based on solid phase extraction of entacapone using Oasis HLB cartridges. Extraction yields were good, being always higher than 96%. Both methods, based on HPLC-ED (V = +0.8 V), have been fully validated. Good linearity was obtained over the following concentration ranges: 100-4000 ng mL(-1) for L-dopa, 200-10,000 ng mL(-1) for 3-O-methyldopa, 25-4000 ng mL(-1) for carbidopa and 20-4000 ng mL(-1) for entacapone. Precision data were satisfactory, being R.S.D.% values lower than 5.64%; accuracy also resulted very good with recovery data higher than 90%. The proposed methods have been successfully applied to the analysis of patient plasma samples and seem to be suitable for therapeutic drug monitoring purposes. (C) 2010 Elsevier B.V. All rights reserved.