Method development to assay the inhibition of CYP2D6 by morinda citrifolia L. using high performance liquid chromatography / Wan Shamsun Bahari Wan Mohamad

Recently, there are many studies that show the interaction between CYP206 enzyme and herbs as the use of herbs as alternative and/or complementary therapy is on the rise. In this research, the interaction of CYP206 and Morinda citrifolia L. was being investigated. But first of all the the sensitive and selective method to analyze bufuralol and its main metabolite, 1'-hidroxybufuralol which were used as a probe had been developed using high-performance liquid chromatographic (HPLC). The chromatogram of both compounds were achieved using Agilent 1200 Series Automatic Injector system, Luna Su CN I OOA (5 µm, 150 x 4.6 mm i.d) column with fluorescence detection set at excitation wavelength 252 nm and emission wavelength 302 nm. The mobile phase consisting of methanol HPLC grade and ammonium acetate (CH3COONH4) buffer, pH 3 (30:70, v/v) have been used at a flow rate of 1.0 ml/min. The method was highly specific where other coformulated compounds did not interfere. The results showed good separation of the peaks which were at 7.11 minutes for bufuralol and 3.37 minutes for 1'-hydroxybufuralol. The method was validated for its linearity, accuracy, precision, sensitivity and selectivity. An experimental design was used during validation to evaluate method robustness. The calibration curves in plasma were linear over the range of 2.5-100 ng/ml for 1'­ hydroxybufuralol with detection limit of 2.5 ng/ml. The coefficient variation (CV) of the results of within-day precision and accuracy of the drug were <7%. There was no significant difference between inter-day and intra-day studies for 1'­ hydroxybufuralol which confirmed the reproducibility of the assay method. The validated method then had been applied for the incubation of bufuralol and CYP206