Separation of proteins and peptides by capillary electrophoresis in acid buffers containing high concentrations of surfactants.
Separations of proteins at acid pH in the presence of a high concentration of surfactant [sodium laurylsulfate (SDS), 50 mmol/l] was investigated. The purpose of using high concentrations of SDS as background electrolyte modifier was threefold: First, the surfactant exerts a washing effect upon the capillary wall thus preventing binding of analytes and possible clogging of the capillary. Second, it was revealed that even under very acid conditions (below pH 3) the surfactant is capable of forming associates with protein analytes which still bear considerable negative charge and can be separated on this basis. Third, the system can be applied not only for protein mixtures sufficiently soluble in neutral to alkaline media (leukocyte lysates, standard proteins), but it can be used also with proteins, that are under such conditions virtually insoluble and their solubilization is possible in acid buffers only (eggshell proteins or collagen CNBr fragments). The result was that adsorption to the capillary wall was minimized and the analytes were separated as negatively charged associates with high efficiency. With collagen fragments partition was possible on the affinity differences of the peptides to the surfactant micelles and inner wall of the capillary. Theoretical plate counts approaching 100,000 were easily achieved even with proteins which under the more conventional operation conditions exhibit considerable sticking to the capillary wall. The other feature of this system is that the associates move very rapidly to the anode. Owing to the low pH, endoosmotic flow is negligible, and therefore the system has to be operated at reversed polarity.
Miksík
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Institute of Physiology, Academy of Sciences of the Czech Republic, Prague. miksik@biomed.cas.cz
Deyl
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Z
D002021:Buffers; D019075:Electrophoresis, Capillary / Q000379:methods*; D010455:Peptides / Q000302:isolation & purification*; D011506:Proteins / Q000302:isolation & purification*; D013501:Surface-Active Agents / Q000737:chemistry*
D002021:Buffers; D010455:Peptides; D011506:Proteins; D013501:Surface-Active Agents