Enantioseparation of d-, l-threo-ritalinic acids, the metabolites of d-, l-threo-methylphenidates, by high performance liquid chromatography

A novel high performance liquid chromatographic method was developed and validated for the quantitative analysis of enantiomeric ritalinic acid (RA), the metabolites of d- , l-threo-methylphenidates (MPH), in human plasma and enzyme buffer solution. An α₁-acid glycoprotein column was used with mobile phase of 0.4 % (v/v) acetic acid solution containing 0.1 % (v/v) dimethyloctylamine (pH 3.4). The assay was successfully applied for in vitro analysis of enantiomeric ritalinic acids produced in human and dog plasma and dog liver. The results demonstrate that the esterase in human plasma metabolize d-MPH faster than l-MPH.