High performance liquid chromatography method development to determine inhibition of CYP2D6 by andrographis paniculata / Norlaila Kartina Malini Mamat

This paper describes the development of an HPLC method for the determination of Bufuralol and 1-Hydroxybufuralol. Separations were achieved by using a Luna CN Cl8 column with Fluorescent detection (excitation 252nm and emission 302nm) for quantification. The mobile phase consists of 70% 10mM Ammonium Acetate (pH3) and 30% Methanol. The method was validated for linearity in three days assay. The peak of Bufuralol was observed at 7.1 minutes and 1-Hydroxybufuralol at 3.2 minutes. This method produced narrower and taller peaks for both samples. Besides, this method was used to detect andrographolide, isoandrographolide, neoandrographolide and l 4-deoxy- 11, 12-didehydroandrographolide in commercial product and plant materials of Andrographis paniculata (Hempedu bumi). Andrographolide is believed to prevent free radical production and the peak was observed at 2.4 minutes, while Isoandrographolide was reported to possess cell differentiating inducing activity and for this compound the peak was observed at 2.8 minutes. Neoandrographolide was reported to possess anti­ hepatotoxicity and the peak was observed at 2.1 minutes. On the other hand, l 4-deoxy- 11, 12-didehydroandrographolide' speak was observed at 4.0 minutes. This compound shows marked hypotensive effects. In this method, the Andrographis paniculata components were eluted faster than the previous researches. However, the effect of these components on Bufuralol metabolism by CYP2D6 enzyme cannot be concluded as the enzyme did not metabolized the substrate under in vitro condition.