Simultaneous Determination of Naphthalene and Anthraquinone Derivatives in Rumex nepalensis Spreng. Roots by HPLC: Comparison of Different Extraction Methods and Validation

Introduction - Rumex nepalensis contains mainly anthraquinone and naphthalene derivatives. Although HPLC methods have been reported for the analysis of anthraquinones, neither a phytochemical analysis of Rumex species nor the simultaneous determination of anthraquinone and naphthalene derivatives in other samples has been reported so far. Objective - To develop and validate a HPLC method for the simultaneous determination of anthraquinone and naphthalene derivatives in R. nepalensis roots. Methodology - Anthraquinones and naphthalenes were extracted from R. nepalensis roots by three methods (reflux, ultrasonication and pressurized liquid extraction) using methanol. Separation was achieved on an RP C-18 column with a gradient mobile phase consisting of 0.05% orthophosphoric acid in water (solvent A) and methanol (solvent B) using a UV detector (254 nm). Results - Small differences were observed in the contents of anthraquinone and naphthalene derivatives extracted by the three methods. Chrysophanol-8-O-beta-D-glucopyranoside and nepodin were detected as major constituents. The method showed a good linearity (r(2) > 0.9992), high precision (RSD < 5%) and a good recovery (97-105%) of the compounds. The lowest detection limit was found to be 0.97 ng and the method was found to be robust. Conclusion - Reflux and ultrasonication were found to be the best suited methods for the extraction of glycosides and aglycones, respectively. The developed and validated HPLC method is simple, precise and accurate; and can hence be recommended as the method of choice for the analysis of anthraquinones and naphthalenes in R. nepalensis and other Rumex species for both quality control as well as routine analytical purposes. Copyright (c) 2010 John Wiley & Sons, Ltd.