Central composite design and response surface methodology for the optimization of Ag+-HPLC/ELSD method for triglyceride profiling

Abstract

The study presents the application of central composite design (CCD) and response surface methodology (RSM) for the optimization of silver-ion normal phase HPLC/ELSD (Ag+-HPLC/ELSD) method parameters to profile the isomers of triglycerides in vegetable oils. The significance of a second-order polynomial model for predicting the optimal values of Ag+-HPLC/ELSD method parameters was evaluated by the analysis of variance, ANOVA, and 3D response surface plots for the interactions between three variables were constructed. Three experimental parameters were chosen as independent variables which are the flow rate of mobile phase, temperature of column compartment and concentration of sample. A multivariate five-level CCD and RSM were used to confirm a quadratic model as a functional relationship between the response values (Rs, N, α and k′) and variables. The optimum values of parameters were found to be a flow rate of 1.25 mL min−1, temperature of column compartment of 20 °C, and sample concentration of 5 × 10−2 mg µL−1. Regression analysis with an R2 values indicated as an adequate correlation between the experimental and predicted response values. ANOVA test results were also confirm that the models can be successfully used to predict the optimum parameters of Ag+-HPLC/ELSD method. Therefore, the proposed model provides an efficient, automated, and robust Ag+-HPLC/ELSD method for triglyceride profiling and is also suitable for a number of applications and analytical method developments for vegetable oils.