Biomedical Engineering Reference
In-Depth Information
TABLE 14.1
Open-Tubular Columns Used in CEC Enantiomer Separations
Type of Immobilization
Selector-Type
Ref.
Adsorptive noncovalent bonding
Proteins (lysozyme, avidin)
[82,87-90]
Biospecii c binding of avidin to
biotinylated phospholipid coatings
[91]
Polyelectrolyte multilayers
Poly(
N
-undecanoyl-dipeptides)
[92,93]
Covalent linkage
Proteins (BSA, avidin)
[81,94,95]
Polymer coating (WCOT)
Cyclodextrin-bonded polysiloxane
[80]
Terguride-bonded polysiloxane
[96]
Polysaccharide
[83]
Porous-layer (PLOT type)
MIP
[85]
solution. The straightforward preparation procedure facilitates the use of 20 or even
10
m i.d. capillaries that lead to improved column efi ciencies. Liu et al. reported
on avidin-adsorbed columns (28
μ
m i.d.) and the amount of avidin adsorbed on the
capillary wall was estimated by frontal analysis to be of the order of magnitude of
10
−12
mol for a capillary of 50 cm effective length, corresponding to 10
−8
mol/m
2
[90]. A number of pharmaceuticals could be baseline resolved into enantiomers with
10 mM phosphate buffer (pH 6) containing up to 5% (v/v) methanol with reasonable
plate numbers for the i rst migrating enantiomers (10-70,000 plates/m). The second
migrating enantiomer exhibited much lower plate counts and a signii cant tailing
which is not uncommon for separations on protein CSPs. The drifting baseline in
these separations might be an indication for bleeding of the selector.
In a similar adsorptive column fabrication approach phospholipid-coated fused
silica capillaries (50
μ
m i.d.) with immobilized avidin were recently prepared by
Han et al. [91]. These OT columns were obtained in a two step procedure in which
i rst a phospholipids coating was generated by rinsing with 1 mM liposome solution
synthesized from phosphatidylserine and biotinylated phospholipids for 10-30 min.
Avidin was then adsorbed onto the biotin-residues by rinsing for about 30 min. These
OT-columns showed highly efi cient separations for PTH-amino acids (Figure 14.5).
A related procedure with great potential for OT-EC has been proposed by
Warner's group [97]. They make use of a PEM coating procedure for the fabrication
of enantioselective capillary columns [92,93]. The technique is based on an elec-
trostatic layer-by-layer deposition of positively and negatively charged polymers.
The coatings are fabricated in situ by rinsing a fused silica capillary alternating with
cationic and anionic polyelectrolytes starting with the cationic polymeric surfac-
tant. One cationic and one anionic layer constitute a so-called bilayer. Since the last
layer was usually prepared by the negatively charged polymer, the authors always
observed cathodic EOF. Several bi-layers can be applied which was shown to increase
the separation factors as illustrated for 1,1
μ
-dihydrogenphosphate in
Figure 14.6. It constitutes a straightforward approach to adjust the enantioselectiv-
ity by the demand of the separation problem. In their studies, the authors utilized
′
-binaphthyl-2,2
′
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