Theoretical and experimental studies on sequential two-diffusional sample injection for capillary electrophoresis

We report here theoretical and experimental studies on the sequential diffusion injection (SDI) for CE analysis. Based on the Fick's second law, a theoretical model for two-dimensional (2-D) diffusion has been developed for our SDI system. The 2-D diffusion model has been demonstrated via systematic experimental studies using standard nicotinamide adenine dinucleotide (NADH) as the model analyte. The results show that the dependence of the NADH peak area (corresponding to the injection amount) on the initial sample concentration, the injection time or the capillary-gap distance is consistent with the deduction of the 2-D diffusion model. It is indicated that the 2-D diffusion, both in longitudinal and transverse directions of the capillary, enhances the injection efficiency in comparison to classical concentration diffusion on the plane interface, and improves the accuracy of the sequential injection without any physical disturbance of the capillary inlet. With the insight understanding of the injection mechanism, we have successfully applied the SDI method for sequential CE analysis of amino acids mixture and online assay of the glucose-6-phosphate dehydrogenase-catalyzed reaction. The present study showed that the SDI is a versatile tool for efficient and accurate sequential CE analysis, not only for online monitoring various bioprocesses but also for continuous analyzing complex samples based on capillary electrophoresis.