Method development for the certification of a ginsenoside calibration solution via liquid chromatography with absorbance and mass spectrometric detection
Walter B. Wilson, Lane C. Sander
The research presented here describes the development of two analytical methods for use in the certification of a ginsenoside calibration solution Standard Reference Material (SRM) 3389 consisting of seven ginsenosides: Rg1, Re, Rf, Rb1, Rc, Rb2, and Rd. The new methods utilized the liquid chromatographic (LC) separation of ginsenoside mixtures with absorbance detection (UV) and mass spectrometry (MS). Ginsenosides Rb3, Rg2, Rg3, Rh1, and Rh2 were evaluated for use as internal standards for LC/MS measurements. Two new analytical methods were developed for the separation and detection of 12 ginsenosides by combing liquid chromatography with ultraviolet-visible absorbance detection (LC/UV) and mass spectrometry (LC/MS). The 12 ginsenosides were baseline resolved via LC/UV with a 0.7 mL/min flow rate, and by gradient elution LC/UV, with an initial mobile phase composition of 22 % acetonitrile and 78 % water, flow rate of 0.7 mL/min, and column temperature of 25 oC. The work presented here includes a detailed investigation into the optimization of the chromatographic conditions to minimize measurement biases that result from unresolved constituents. Column tTemperature and mobile phase composition are known to play a significant role in columnthe selectivity of an LC column; however, flow rate is expected to influence primarily the separation efficiency and detection sensitivity. In the current study, column selectivity changed with changes in flow rate and the relative retention of ginsenoside Rg2 and Rh1 decreased changed as the flow rate increased from 0.6 mL/min to 1.0 mL/min. The LC/UV and LC/MS methods were used for the certification of a new ginsenoside calibration solution (Standard Reference Material 3389).