CAPILLARY ZONE ELECTROPHORESIS OF PEPTIDES - PREDICTION OF THE ELECTROPHORETIC MOBILITY AND RESOLUTION

The determination of the pK(a) values of some selected peptides of\r\nsimilar size was performed by microtitration, which makes possible an\r\naccurate determination of the peptide charge as a function of the solution pH. Capillary zone electrophoresis separation of these peptides\r\non modified capillaries at acidic pH showed that the electrophoretic\r\nmobility correlates with the peptide charge. This observation suggests\r\nthat when an appropriate charge value is used, the basic electrophoretic\r\nequation is respected and, at least at a peptide charge value less than\r\nI, the utilization of alternative semi-empirical predictions is not\r\nnecessary. As a general rule, a peptide separation at acidic pH values\r\nis to be preferred to that at basic pH values. In fact, at basic pH a\r\nseparation in the absence of both electroosmotic flow and of spurious\r\ninteractions between the peptides and the inner wall of the capillary is\r\ndifficult, owing to the instability of capillary modification. Further,\r\nfrom the differences in the peptide charge, a prediction of the best\r\nresolution as a function of the pH could be obtained; in fact, the\r\nresolution, for peptides of similar size and in the absence of\r\nelectroosmotic flow, is connected to a simple equation, where the\r\nprincipal term depends on the effective charge of the peptides, which is\r\na function of the pH of the solution and the pK, values of the peptides.\r\nThe predictions of resolution at acidic pH agreed well with the\r\nexperimental results; the spatial resolution measured in the separation\r\nof met- and leu-enkephalin was virtually coincident with the predicted\r\nresolution; in the case of a mixture of four model tetrapeptides of\r\nsequence GGNA, GGQA, GGDA and GGEA some anomalous results with respect\r\nto the predicted resolutions were observed. Nevertheless, an acceptable\r\nprediction can also be made in this case.