Determination of methylphenidate and its main metabolite in plasma by gas chromatography-chemical ionization mass spectrometry.

A method is described for the determination of methylphenidate and its main metabolite, ritalinic acid, in plasma from various species, based on gas chromatography-chemical ionization mass spectrometry using ethylphenidate as an internal standard. Methylphenidate in plasma is extracted with cyclohexane and converted to the pentafluoropropionyl (PFP) derivative. Ritalinic acid is extracted with 2-propanol from the salt-saturated remaining aqueous phase, converted to the parent drug by treatment with a mixture of methanol and sulfuric acid (2 : 1, v/v), and then converted to the PFP derivative. The protonated molecular ions at m/z 380 for the PFP derivative of methylphenidate and m/z 394 for that of the internal standard were used for selected ion monitoring analysis. The hydrolysis of methylphenidate in plasma before the addition of the internal standard could be prevented completely by holding the plasma sample at around 0°C. The limit of quantitation was 0.5 ng for methylphenidate and 2.5 ng for ritalinic acid using 0.5 ml of plasma. The present method is highly sensitive and reliable, and should be satisfactory for application to pharmacokinetic studies of methylphenidate in laboratory animals and man.