The choice of an appropriate background electrolyte (BGE) and its components for capillary electrophoresis analysis is the main step in capillary electrophoresis method development. The use of an inadequate co-ion component could lead to asymmetrical peaks and selecting an inappropriate counter-ion could affect the buffer capacity and the pH of the BGE, leading to unreliable analysis. In this paper, we describe a systematic procedure for the development of a capillary electrophoresis method, based on the effect of varying pH on the ion effective mobility, to optimize the BGE composition. The method was applied to the separation of L-ascorbic acid in different samples. The optimized background electrolyte composition was 40 mmol L-1 tris(hydroxymethyl)aminomethane and 20 mmol L-1 2-morpholinoethanesulfonic acid, at pH 8.1. Sorbic acid was used as the internal standard and separation was carried out in a fused-silica capillary (32 cm total length and 8.5 cm effective length, 50 µm inner diameter), with a short-end-injection configuration and direct ultraviolet (UV) detection at 266 nm. The separation was performed in 26 s. The method shows good linearity (R2 > 0.999), excellent values for inter-day and intra-day precision and good recovery (in the range of 94-107%). The values obtained for limit of detection (LOD) and limit of quantification (LOQ) were 0.14 and 41 mg L-1, respectively. The systematic procedure applied shows to be a very useful tool for the first step method development for capillary electrophoresis.
Keywords:
capillary electrophoresis; effective mobility curves; optimization; L-ascorbic acid