Frans_M_Everaerts_Isotachophoresis_378342.pdf

1.00 CHOICE OF ELECTROLYTE SYSTEMS
differences between the effective mobilities of the ionic species to be separated are too
small for them to be separated, the addition of a spacer will further reduce the differences
in mobilities so that a separation cannot be achieved. Sometimes, however, it can be
advantageous to use a spacer, for instance if ionic species can be separated easily, but if
their zones are small and close together so that there is a risk that the detector cannot
distinguish the separated zones. In such a case we can use, for example, a non-UV-
absorbing spacer (if a UV absorption detector is being used) that separates these zones so
that they can be detected separately. Similarly, we can utilize a UV-absorbing spacer in
order to make the detection of consecutive zones of non-UV-absorbing ionic species
possible.
Another application (see Chapter 13) of spacers is in the separation of molecules
with a high molecular weight, e.g., proteins. For the separation and detection, a series of
compounds can be used with a large range of effective mobilities in the operational
system chosen and, almost always related to this, a large pH gradient. Of course, these
additives are a combination of spacers and carriers (see Chapter 13), but proteins normally
require stabilization with electrolytes. If a protein is introduced in such a gradient, it will
migrate together with the carrier, by which it is diluted, at such a position that its effec-
tive mobility corresponds to the related pH. This subject is dealt with in greater detail in
the Section Applications, where practical information on protein separations is presented.
5.7. DISCUSSION
Prescriptions for the selection of electrolyte systems that are always valid cannot be
given, because the choice of an electrolyte system depends to a great extent on the sample
being analyzed. In order to demonstrate the method of choosing a suitable electrolyte
system, some examples of electrolyte systems are considered in the remainder of this
chapter. Further information and experimental data are given in the Section Applications.
A scheme that can be of help in choosing an electrolyte system is shown in Fig.5.4.
5.8. EXAMPLES
Example A. Suppose we wish to separate two anionic species A and B with ionic
mobilities* of 30 and 50 and with pK values of 1 and 3, respectively. We must decide the
preferred type of separation (according to pK values or according to mobilities), the
pH, and the limits for the effective mobilities of the leading and terminating ionic species.
The effective mobility of the leading ionic species must be greater than 50 and that
of the terminating ionic species must be less than 30. (One must consider the possibility
that at certain pH values the effective mobilities of the sample ionic species can be
much lower than 30 or 50). In this instance we would prefer a separation according to
mobilities. Here the differences between the ionic species are sufficiently large and the
pH, should be about 2 (because the pK values are 1 and 3) for a separation to be carried
out according to pK values. At this pH, the concentration of the hydrogen ions is so high
that the current carried by the counter ions and hydrogen ions is the largest part of the
*.10-5 - cm*/V sec.