Quality by design-based capillary electrophoresis method development for the quantitative analysis of four iridoid compounds in Gentiana macrophylla Radix.

Abstract

In this study, the capillary electrophoresis-photodiode array detector was employed for the analysis of four iridoid compounds in Gentiana macrophylla Radix (RGM), and the method was optimized based on the concept of analytical quality by design (AQbD). The peak areas relative standard deviation (n=3) and resolution of the four analytes were selected as critical method attributes. Fractional factorial design test combined with Pareto analysis were employed to screen critical method parameters (buffer concentration, pH, sodium dodecyl sulfate [SDS] micelle concentration, temperature, and voltage). Subsequently, three main factors (buffer concentration, buffer pH, and SDS concentration) were selected by central composite design test for constructing the design space. The optimal separation conditions as follows: capillary column (50.2cm×50µm, detection length 40cm). Working background electrolyte consisted of 51mmol/L borax solution (pH=9.47) and 40mmol/L SDS. The samples were injected by pressure (5s at 0.5psi) and the detection was performed at 254nm. Applied voltage was 20kV and column temperature was 23°C. The developed method is rapid and reliable for the quantitative analysis of four iridoid compounds in RGM, providing a reference for the application of AQbD concept in the analysis of natural products.