Biomedical Engineering Reference
In-Depth Information
reaction should occur quickly, and therefore the metal coordination compound must
be labile. In such kind of complexes, the bond(s) between the ligand and the metal ions
are continuously formed and broken: in this situation, the thermodynamic stability of
the complex can be conveniently expressed as the mean time spent by the ligand mol-
ecules bound to the metal ion. Consequently, we can appreciate the importance of the
stability of the complex and the degree of afi nity of the ligand toward the metal ion.
If the afi nity is too low, regardless of the lability of the complex, the ligand will spend
a negligible fraction of time bonded to the metal ion. Therefore, this process, though
occurring, will have only a slight effect on the ligand behavior. The complex stability
is conveniently expressed by the formation constant of the complex.
While the value of the complex stability depends on both the ligand and the metal
ion, the lability of the complex depends almost exclusively on the metal ion: in the
choice of the metal ion that we will use in these techniques, we will be forced to
exclude all the metal ions which give rise to inert complexes, typically chromium(III)
or cobalt(III).
The ligand exchange process was originally exploited in liquid chromatography,
starting from the pioneering work by Helfferich as far back as 1961 [1]. In this i rst
version of ligand exchange chromatography (LEC), the metal ion was bound to a cat-
ionic exchanger, and the analyte to be separated, interacting with this metal ion, could
reside on the stationary phase. By adding a ligand containing eluent, a differential
retention of the analytes occurred. By using a purpose-made chiral ion exchanger,
Davankov et al. obtained discrimination of enantiomers of
-amino acids [2]. In this
i rst developed procedure, through the ligand exchange, the afi nity of a specii c mol-
ecule for the metal ion became a sort of afi nity for the stationary phase. Thus, LEC
was a classical chromatographic separation, a two sites technique, the free ligand in
the mobile phase and the complexed ligand in the stationary phase. Later on, modii -
cations were introduced, and the methods can be conveniently classii ed as
α
•
Chiral stationary phase (CSP)
•
Chiral coated stationary phase (CCP)
•
Chiral mobile phase (CMP)
While details about all these techniques can be easily found in excellent reviews [3],
here it is interesting to give a glance at the CMP techniques, obtained by the addition
of a metal ion complex in the mobile phase. From the point of view of the procedure,
the CMP methods show some advantages, since a preliminary preparation of the sta-
tionary phase is not required: a commercially available reverse phase resin appears
more than adequate for this task. Thus, only the preparation of a suitable mobile phase
is required in this case. What soon appears new in this technique, is the possibility to
exploit the homogeneous equilibria occurring in the mobile phase. An accurate study
of such equilibria can be afforded by a plurality of techniques, e.g., spectrophotometry
and, especially, pH-metric potentiometry. Unfortunately, what goes on in the mobile
phase does not represent the totality of the processes, since, as always in chromato-
graphic techniques, we should not forget the presence of the stationary phase. All the
species in solution undergo an additional heterogeneous “quasi-equilibrium,” distribut-
ing themselves between the mobile and the stationary phase. Since what happens in
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