A new method for the determination of l-DOPA and 3- O-methyldopa in plasma and cerebrospinal fluid using gas chromatography and electron capture negative ion mass spectrometry

Abstract

l-3-(3,4-Dihydroxyphenyl)alanine (DOPA) and its 3- O-methyl metabolite (OMD) were measured in plasma and cerebrospinal fluid by a new assay which combines N, O-acetylation of amino acids in aqueous media, preparation of pentafluorobenzyl esters under anhydrous conditions, and analysis by gas chromatography-electron capture negative ion mass spectrometry. The N, O-acetyl, carboxy-PFB derivatives gave abundant carboxylate anions ([M-CH 2C 6F 5] −) which were suitable for sensitive analysis using selected ion monitoring. Plasma and CSF samples were sufficiently purified by a simple organic solvent extraction. Analytical recovery for DOPA was 100.2 ± 3.7% at the level of 100 nmol/l. Analysis of DOPA in plasma was performed with a relative standard deviation of 5%. The limit of quantitation in plasma and CSF was at the sub-nmol/l level. In healthy adults, DOPA concentration in plasma was 9.0 ± 2 nmol/l ( n = 11) and in CSF 3.5 ± 0.9 nmol/l ( n = 9). The concentration of OMD in plasma was 99.1 nmol/l (pool of 24 samples) and 15.3 nmol/l in CSF (pool of 12 samples). Measurement of 5-[ 2H]DOPA and 5-[ 2H]OMD in plasma of a healthy individual who had been orally loaded with 3,5-[ 2H 2]tyrosine (150 mg kg body wt) was possible for several hours after the load.