High-resolution computer simulation of the dynamics of isoelectric focusing: In quest of more realistic input parameters for carrier ampholytes

Abstract

The impact of the systematic variation of either ΔpK2 or mobility of 140 biprotic: carrier ampholytes on the conductivity proffie of a pH 3-10 gradient was studied by dynamic computer simulation. A configtiration with the greatest ΔpK2 in the pH 6-7 range and uniform mobilities produced a conductivity profile consistent with that which is experimentally observed. A similar result was observed when the neutral (pI = 7) ampholyte is assigned the lowest mobility and mobilities ofthe other carriers are systematically increased as their pIs recede from 7. When equal ΔpK2 values and mobilities are assigned to all ampholytes a conductivity plateau in the pH 5-9 region is produced which does not reflect what is seen experimentally. The variation in ΔpK2 values is considered to most accurately reflect the electrochemical parameters of commercially available mixtures of carrier ampholytes. Simulations with unequal mobilities of the cationic and anionic species of the carrier ampholytes show either cathodic (greater mobility of the cationic species) or anodic (greater mobility of the anionic species) drifts of the pH gradient. The simulated cationic drifts compare well to those observed experimentally in a capillary in which the focusing of three dyes was followed by whole column optical imaging. The cathodic drift flattens the acidic portion of the gradient and steepens the basic part. This phenomenon is an additional argument against the notion that focused zones of carrier ampholytes have no electrophoretic flux. © 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.