Frans_M_Everaerts_Isotachophoresis_378342.pdf

ISOTACHOPHORESIS 13
2.4. PRINCIPLE OF iSOTACHOPHORESIS
2.4.1. Introduction
We shall consider here the separation of anionic species in narrow-bore tubes. For
the separation of anionic species, the narrow-bore tube and anode compartment are
filled with the so-called leading electrolyte, the anions of which must have a mobility
that is higher than that of any of the sample anionic species. The cations of the leading
electrolyte must have a buffering capacity at the pH at which the analyses will be
performed. The cathode compartment is filled with the terminating electrolyte, the
anions of whch must have a mobility that is lower than that of any of the sample
anionic species. The sample is introduced between the leading and terminating
electrolyte, e.g., by means of a sample tap or a micro-syringe.
#en an electric current is passed through such a system (see Fig.2.5a), a uniform
electric field strength over the sample zone occurs and hence each sample anionic species
will have a different migration velocity according to eqn. 2.1. The sample anionic species
with the highest effective mobility will run forwards and those with lower mobilities will
remain behind. Hence, both in front of and behind the original sample zone, the moving-
boundary procedure results in two series of mixed zones (comparable with the Tiselius
method). In the series of mixed zones, the sample anionic species are arranged in order
of their decreasing effective mobilities (see Fig.2.5b),
The anionic species of the leading electrolyte can never be passed by sample anions,
because its effective mobility is chosen so as to be higher. Similarly, the terminating
anions can never pass the anionic species of the sample. In this way, the sample zones
are sandwiched between the leading and terminating electrolyte. In the mixed zones of
the sample (see Fig.2.5b), the separation continues and, after some time, when the
separation is complete, a series of zones is obtained in which each zone contains only one
anionic species of the sample if no anionic species with identical effective mobilities are
present in the sample. Of course, this series of zones is still sandwiched between leading
and terminating electrolyte (see Fig.2.5~).
The first sample zone contains the anionic species of the sample with the highest
effective mobility, the last zone that with the lowest effective mobility. After this stage,
no further changes to the system occur and a steady state has been reached. In such a
case, we can speak of an isotachophoretic separated system. (Of course, one or more
unmixable ‘mixed zones’, i.e., zones that contain one or more anionic species with
identical effective mobilities, may still be present.) In this state, all of the zones must run
connected together, in contrast to zone electrophoresis, where all zones release. Here the
zones cannot release as there is no background electrolyte that can support the electric
current (a requirement for the solvent is that its self-conductance must be negligible; see
section 5.2 .)* .
*If it is assumed that the zones release, then the concentration of the ionic species at that position will
decrease, the electric field strength will increase (working at a constant current density) and hence the
migration velocity of the ionic species involved will be higher. Therefore, finally these ionic species will
reach the preceding zone.