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The lack of purified standards is a bottleneck on assaying herbs  in vitro  and  in vivo . This present work proposed a strategy of relative quantification that used a herb extract as a relative standard. A rapid and selective liquid chromatography tandem mass spectrometry method was similarly developed and validated for the relative determination of dehydroevodiamine in rat plasma according to the absolute quantification. Protein precipitation was used for the pretreatment of plasma samples. Chromatographic separation was achieved on a Diamonsil C18 column with an isocratic mobile phase of a 70:30 (v/v) acetonitrile–0.3% formic acid mixture at a flow rate of 0.45 mL/min. The assay was validated in the range 100.0 ∼ 50,000.0 ngH/mL (r 2  = 0.9804), the lowest level of this range being the lower limit of quantification based on 50 µL of plasma. The precision and accuracy were within recommended limits of nominal values. The method was applied to evaluate the comparative pharmacokinetics of dehydroevodiamine in rats following oral administration of  Evodia rutaecarpa  and  Rhizoma coptidis–Evodia rutaecarpa  couple. This approach was found to be capable of providing complete pharmacokinetic parameters as well as the typical pharmacokinetic assay calibrated by authentic standards, except for the absolute plasma concentrations.

Relative Determination of Dehydroevodiamine in Rat Plasma by LC–MS and Study on its Pharmacokinetics