Capillary isoelectric focusing in presence of electroosmosis with sequential injection
of carrier ampholytes and sample was found to be suitable for mass spectrometry detection.
The separate injection of the sample and the ampholytes provides good condition to
suppress and overcome the undesirable effect of the presence of ampholytes in mass
spectrometry. By the appropriate selection of ampholyte solutions, whose pH range
not necessarily covers the pI values of the analytes, the migration of the components
can be controlled, and the impact of the ampholytes on mass spectrometric detection
is decreased. The unique applicability of this setup is shown by testing several parameters,
such as, the application of volatile electrolyte solutions, the type of the ampholytes,
the order and the number of the ampholyte and sample zones. Broad and narrow pH range
ampholytes were applied in experiments using uncoated capillaries with different length
for the analyses of substituted nitrophenol dyes to achieve optimal conditions for
the mass spectrometric detection. Although, the sample components are not leaving
the pH gradient, due to the decrease in the ampholyte concentration at the position
of the components, and because the sample components migrate in charged state, the
ionization is more effective for mass spectrometry detection. Results with model calculations
confirmed the mechanism of the process in the isoelectric focusing using one or two
ampholyte zones before and/or after the sample zone.
The work was supported by grant OTKA-NKTH-NI-68863 and by project SROP-4.2.2/08/1/2008-0011.