Page 29 - Chiral Separation Techniques

4 1 Techniques in preparative chiral separations

1.3.1.1 High-Pressure/Medium-Pressure Liquid Chromatography
(HPLC/MPLC)

HPLC separations are one of the most important fields in the preparative resolution
of enantiomers. The instrumentation improvements and the increasing choice of
commercially available chiral stationary phases (CSPs) are some of the main reasons
for the present significance of chromatographic resolutions at large-scale by HPLC.
Proof of this interest can be seen in several reviews, and many chapters have in the
past few years dealt with preparative applications of HPLC in the resolution of chi-
ral compounds [19–23]. However, liquid chromatography has the attribute of being
a batch technique and therefore is not totally convenient for production-scale, where
continuous techniques are preferred by far.
In order to carry out a direct preparative chromatographic resolution of enan-
tiomers in batch elution mode, the same methods as are used in other nonchiral
large-scale chromatographic separations are applied [24], such as multiple close
injections, recycling or peak shaving. All of these are addressed to reduce cost and
increase yield, while saving solvent and making full use of the stationary phase.
Thus, the injections are sometimes performed repeatedly (multiple close injection),
in such a way that most of the chromatographic support is involved in the separation
at any moment. When the resolution is not sufficient, it can be improved by recycling
of the partially overlapped peaks [23]. This has almost the same effect as is obtained
when using a longer column, but clearly a broadening of peaks occurs. However,
after several cycles partially overlapped peaks can be completely resolved. This
method is often combined with the so-called peak shaving, which allows the recov-
ery of the part of the peaks corresponding to pure enantiomers while the overlapped
region is recycled. In fact, to date this method is the most often used.
The chiral environment needed for enantiomeric separations is furnished by the
chiral support into the column. Scale-up of a chiral separation can be made having
as a reference an analytical resolution, but optimization of the preparative process is
critically dependent upon the nature of the CSP. In order to increase the throughput,
the column is usually used in overloading conditions. The loading capacity of a chi-
ral stationary phase depends not only on the chiral selector density, but also on the
type of selector. Therefore, some types of CSPs are more suitable than others for
preparative purposes [21, 25]. Not all the commercially available CSPs used for ana-
lytical purposes are appropiated for large-scale resolutions (Table 1-1). CSPs with a
large application domain, such as those derived from proteins with a vast applica-
bility for analytical purposes, have a very low loadability and, therefore, they are not
well adapted to preparative separations [26, 27]. This is also the case of molecular
imprinted polymers (MIPs) [28–30]. The limited number of recognition sites
restricts their loading capacity and thus also their use in large-scale chromatography.
Some ligand-exchange CSPs have been used at preparative level [31, 32]. In this
case it must be taken into account that an extraction process, to remove the copper
salts added to the mobile phase, must be performed following the chromatographic
process [33]. Teicoplanin, in contrast, resolves all ordinary α and β-amino acids with
mobile phases consisting of alcohol/water mixtures. No buffer is needed in the