Simultaneous HPLC Determination of 22 Components of Essential Oils; Method Robustness with Experimental Design

The aim of the present study was the development and validation of a simple, precise and specific reversed phase HPLC method for the simultaneous determination of 22 components present in different essential oils namely cinnamon bark oil, caraway oil and cardamom fruit oil. The chromatographic separation of all the components was achieved on Wakosil-II C18 column with mixture of 30 mM ammonium acetate buffer (pH 4.7), methanol and acetonitrile in different ratio as mobile phase in a ternary linear gradient mode. The calibration graphs plotted with five different concentrations of each component were linear with a regression coefficient R(2) >0.999. The limit of detection and limit of quantitation were estimated for all the components. Effect on analytical responses by small and deliberate variation of critical factors was examined by robustness testing with Design of Experiment employing Central Composite Design and established that this method was robust. The method was then validated for linearity, precision, accuracy, specificity and demonstrated to be applicable to the determination of the ingredients in commercial sample of essential oil.