Simultaneous quantitation of levodopa and 3-O-methyldopa in human plasma by HPLC-ESI-MS/MS: application for a pharmacokinetic study with a levodopa/benserazide formulation.

Abstract A sensitive and simple method was developed for the quantitation of levodopa and its metabolite 3- O -methyldopa, in human plasma, after oral administration of tablet formulations containing levodopa (200 mg) and benserazide (50 mg). The analytes were extracted by a protein precipitation procedure, using carbidopa as an internal standard. A mobile phase consisting of 0.2% formic acid and acetonitrile (94:6, v/v) was used and chromatographic separation was achieved using ACE C 18 column (50 mm × 4.6 mm i.d.; 5 μm particle size). Selected reaction monitoring was performed using the fragmentation transitions m / z 198 →  m / z 107, m / z 212 →  m / z 166 and m / z 227 →  m / z 181 for levodopa, 3- O -methyldopa and carbidopa, respectively. Calibration curves were constructed over the range 50.0–6000.0 ng/mL for levodopa and 25.0–4000.0 ng/mL for 3- O -methyldopa. The method shown to be specific, precise, accurate and provided recovery rates higher than 85% for all analytes. No matrix effect was detected in the samples. The validated method was applied in a pharmacokinetic study with a levodopa/benserazide tablet formulation in healthy volunteers.