Reverse Phase High Performance Liquid Chromatography for the Quantification of Eurycomanone in <i>Eurycoma longifolia</i> Jack (Simaroubaceae) Extracts and their Commercial Products

Purpose: To develop a reverse phase high performance liquid chromatography (HPLC) method for the determination of eurycomanone in E. longifolia Jack (Simaroubaceae) aqueous root extract and their commercial products.Methods: Analysis was carried out using reverse phase HPLC at 254 nm and a gradient mobile phase that comprised of acetonitrile and 0.1 % formic acid. Flow rate was 1 ml/min and separation was done using Phenomenex, Luna C18 column (150 mm x 4.6 mm, 5 μm). Validation tests were performed in order to demonstrate the linearity, precision, accuracy, limit of detection (LOD) and limit of quantification (LOQ) of the method.Results: Linearity was in the range 0.1 – 50.0 μg/ml (R2 = 0.9999). Precision, as relative standard deviation of retention time and peak area of reference compound was < 0.14, and < 2.75 %, respectively. Accuracy, as percent recovery of eurycomanone, was in the range 94.2 – 99.8 % while LOD and LOQ were 0.293 ± 0.100 and 0.887 ± 0.300 μg/ml, respectively. Eurycomanone concentration in E. longifolia extracts and commercial products was 0.89 – 3.28, and 0.07 – 0.16 %, respectively. Analysis of the ethanol extract and chloroform sub-extract showed high resolution of > 10 peaks, which indicate the suitability of the method for the analysis of extracts prepared in organic solvents as well.Conclusion: The proposed method shows good linearity, precision, accuracy and high sensitivity. The method can be applied in the routine quantification of eurycomanone for quality control of E. longifolia extracts and commercial products.Keywords: Eurycoma longifolia Jack, Tongkat Ali, Eurycomanone, Validation