Mobility‐based correction for accurate determination of binding constants by capillary electrophoresis‐frontal analysis

Abstract

Capillary electrophoresis frontal analysis (CE‐FA) can be used to determine binding affinity of molecular interactions. However, its current data processing method mandate specific requirement on the mobilities of the binding pair in order to obtain accurate binding constants. This work shows that significant errors are resulted when the mobilities of the interacting species do not meet these requirements. Therefore, the applicability of CE‐FA in many real word applications becomes questionable. An electrophoretic mobility‐based correction method is developed in this work based on the flux of each species. A simulation program and a pair of model compounds are used to verify the new equations and evaluate the effectiveness of this method. Ibuprofen and hydroxypropyl‐β‐cyclodextrinare used to demonstrate the differences in the obtained binding constant by CE‐FA when different calculation methods are used, and the results are compared with those obtained by affinity capillary electrophoresis (ACE). The results suggest that CE‐FA, with the mobility‐based correction method, can be a generally applicable method for a much wider range of applications.

Capillary Electrophoresis/Dynamic Frontal Analysis for the Enzyme Assay of 4-Nitrophenyl Phosphate with Alkaline Phosphatase

“…[9][10][11] Simulation of the plateau signal was also studied with binding of ibuprofen to hydroxypropyl--cyclodextrin. 12 A simulation program of Simul 5 Complex is applicable to CE/FA. 13,14 As for the enzyme assays, CE is widely used in the dynamic or kinetic reactions of enzymes.…”

Pressure‐assisted capillary electrophoresis frontal analysis for faster binding constant determination

“…Some previous works have discussed this issue . {Winzor, 2006 #406}In our recent work, a mobility‐correcting algorithm was established to expand the application range of CE‐FA . The relationship of the true unbound ligand concentration at equilibria with measured concentration is described by: where [ L ], [ L ] t , [ L ] m are the actual concentrations of unbound ligand at equilibria, total concentration of the ligand, and measured ligand concentration, respectively; μ HL , μ H and μ L are the mobilities of the complex, the host and the ligand molecules, respectively.…”

“…Frontal analysis, before being used in CE, had its genesis in the field of liquid chromatography. Since reported by Kraak et.al in 1992, CE‐FA has been used as a technique for the characterization of non‐covalent interactions between molecules . Compared with other CE‐based techniques, a much larger amount of sample ( ca 100–150 nL) is used in CE‐FA.…”

Determining binding constants for 1:1 and 1:2 inclusion complexes of ester betulin derivatives with (2‐hydroxypropyl)‐β‐cyclodextrin by affinity capillary electrophoresis

“…In 2017, Qian et al. [19] suggested a mobility‐based correction to use FA ACE for the case of different mobilities of the analyte and 1:1 complex. But this approach was tested on one system, and the simulated electropherograms differed from those obtained experimentally.…”

“…But this approach was tested on one system, and the simulated electropherograms differed from those obtained experimentally. In addition, the authors [19] compared the results of ms ACE and FA ACE and found that these results were similar, but the authors did not fit the triangular peaks from ms ACE with the Haarhoff–Van der Linde (HVL) function. Thus, ms ACE provides more reliable and precise values of binding constants as compared to other CE‐based techniques and is the most suitable ACE mode for studying most CD complexes.…”