Determination of Myrtucommulone from Myrtus communis in Human and Rat Plasma by Liquid Chromatography/Tandem Mass Spectrometry

Recent studies revealed that the non-prenylated acylphloroglucinol myrtucommulone (MC) from myrtle (Myrtus communis) potently suppresses the biosynthesis of eicosanoids by direct inhibition of cyclooxygenase-1, microsomal prostaglandin E2 synthase (mPGES)-1, and 5-lipoxygenase at IC(50) values in the range of 1 to 29 mu M. Moreover, MC showed potent efficacy in animal models of inflammation after intraperitoneal administration. Since the main prerequisite for therapeutic efficacy is sufficient bioavailability, it is important to evaluate whether the concentrations of MC achieved in plasma coincide with the pharmacological active concentrations determined in vitro. For that reason, a sensitive LC/MS/MS-method has been developed and validated for the determination of MC in human plasma. This method is based on liquid-liquid extraction of plasma samples with 20% ethyl acetate in tert-butyl methyl ether using the structurally related acyl-phloroglucinol hyperforin as the internal standard. Chromatographic separation was achieved on a Gemini C6 Phenyl column using a mixture of acetonitrile/water (85 : 15 v/v) containing 6 mM ammonium formate in a run time of 15 min at a flow rate of 1 mL/min, a column temperature of 40 degrees C, and an autosampler temperature of 5 degrees C. Mass spectrometric quantification was carried out in the negative ion mode using electrospray ionization (ESI) and multiple-reaction monitoring (MRM). The most intense [M - H](-) MRM transition at m/z 667.4 -> m/z 194.9 was used for quantification of MC and the transition at m/z 535.4 to m/z 383.2 was used to monitor hyperforin. The method was linear in the range of 1-100 ng/mL with r > 0.998, an intra- and inter-day RSD of 1.1-8.4 and 7.1-11.8%, respectively, and a maximum R. E. of 13.8% at the lowest concentration level. Moreover, cross validation revealed the suitability of the developed LC/MS method for application in rat studies.