Capital HPLC

Types of Chiral Phases for HPLC

Types of Chiral Phases for HPLC

Since the dependence of the chemical or biological
activity of a substance often depends upon it’s
stereochemistry, a number of stereoselective
separation methods have been developed over
the past few years. Currently three HPLC
separation strategies are in common use for the
separation of enantiomers, these are (1)
diastereomer formation, (2) chiral mobile phase
additives, and (3) chiral stationary phases (CSPs).

(1) Diastereomer formation is an indirect method
in which the sample mixture of enantiomers is
reacted with a chiral reagent, to form a pair of
diastereomers. The diastereomers possess
different physicochemical properties and can
be separated on conventional columns, which
is the major advantage to this approach.
However, it also has a number of
disadvantages, the derivatisation stage can
be laborious, display differential reaction rates,
requires a pure chiral reagent to avoid the
misleading formation of interfering
diastereomeric pairs, also further chemical
treatment is necessary if the starting
enantiomers are to be recovered.

(2) The use of chiral mobile phase additives eg.
camphorsulphonic acid, and cyclodextrins,
are also popular as they offer direct separation
of enantiomers, via the formation of stable
diastereomeric complexes with the
enantiomeric solutes, again on conventional
columns. Its disadvantages include that the
additive may interfere with detection, might be
difficult to remove in the case of preparative
LC, and can be expensive.

(3) CSP involves the binding of a chiral selector to
a support material, usually silica. Transient
diastereomeric complexes are formed (with
different stabilities) between the solute
enantiomers and the bound chiral selector.

Chiral column technology is a rapidly expanding
field in which well over fifty phases are commerically
available, with new ones appearing all the time. In
1987 Dr Irving W. Wainer, (1) classified the
available phases into the following five categories
according to their chiral recognition mechanisms:

Type I - in which the solute-CSP complexes
are formed by mechanisms such
as attractive interactions, hydrogen
bonding, dipole stacking, and pi-pi

Type II - in which the primary mechanism
for the formation of solute-CSP
complexes is through attractive
interactions, but where inclusion
complexes also play an important

Type III - in which the solute enters into chiral
cavities within the CSP to form
inclusion complexes.

Type IV - in which the solute forms part of a
metal diastereomeric complex, also
known as chiral ligand exchange

Type V - in which the CSP is a protein and
the solute-CSP complexes are
based on combinations of polar
and hydrophobic interactions.



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