Method of separating chemical mixtures

Details Method of separating chemical mixtures Method of separating chemical mixtures

1998-09-03

2001-05-01

Therkorn, Ernest G. (Department: 1723)

Liquid purification or separation

Processes

Liquid/liquid solvent or colloidal extraction or diffusing...

C210S656000, C210S748080, C210S198200, C204S455000

Reexamination Certificate

active

06224775

ABSTRACT:

FIELD OF THE INVENTION
The invention relates to a method of separating chemical mixtures. More particularly, the invention relates to a method of separating chiral and achiral chemical mixtures through capillary electrochromatography wherein an immobilized carbohydrate polymer is used as a chemical selector.
BACKGROUND OF THE INVENTION
Capillary electrochromatography (CEC) is a hybrid method of capillary electrophoresis (CE) and high performance liquid chromatography (HPLC). Though CEC was first demonstrated more than two decades ago, the advent of sophisticated CE instrumentation, expanded use and understanding of CE, and the continuing quest for more efficient separation methods has recently intensified interest in CEC.
CEC involves the application of an electric field between the ends of a 50-110 &mgr;m capillary containing a stationary phase. “Open tubular” CEC describes a technique where the stationary phase is bonded to the capillary wall, while “packed” CEC describes a method involving capillaries filled either with a polymer gel stationary phase or a small particle (about 1-10 &mgr;m) silica-based stationary phase.
In all CEC techniques, a liquid phase is transported through the capillary by electroosmosis or a combination of electroosmosis and pressure, and solutes are separated based on their partitioning between the stationary and mobile phases and on their charge to frictional drag. As shown in
FIG. 1
, the electroosmotic flow originates from the electrical double layer at the surface of the stationary phase as well as the capillary wall and generates a plug-like flow profile which is independent of the geometry and size of the channels between the particles. This phenomenon can provide very high efficiencies, limited primarily by the solute diffusion coefficient.
In contrast to capillary liquid chromatography, CEC can utilize long capillaries of small, very efficient particles since there is no column back pressure. It has been shown in Dittmann et al.,
LC
-
GC,
13: 800 (1995) that CEC has the potential to provide column plate numbers 5 to 10 times greater than HPLC columns. The high efficiencies attainable make CEC a very attractive technique for chiral separations since it is theoretically possible to obtain baseline resolution for solutes with very small enantioselectivities.
Polysaccharide derivatives coated onto porous derivatized silica have proven to be among the most versatile and widely used chiral stationary phases in HPLC. They have been used in both normal and reversed phase mode and have shown extremely high enantioselectivity for many solutes. Unfortunately, unlike several other chiral selectors, their use as buffer additives in CE is precluded by their poor solubility in suitable electrolytes and high UV cut-off. However, these characteristics do not preclude their use as chiral stationary phases in CEC, and open tubular electrochromatography using 50 &mgr;m I.D. fused silica capillaries coated with a cellulose derivative has been investigated by E. Francotte and M. Jung,
Chromatographia,
42: 521-527 (1996). Resolution was found to be heavily dependent on the thickness of the coating, and the highest efficiency achieved was a disappointingly low 60,000 plates/m.
OBJECTS OF THE INVENTION
Accordingly, it is an object of the invention to provide a novel capillary electrochromatographic separation method which provides improved resolutions, particularly for chiral separations.
Other objects will become apparent from the following description of the invention.
SUMMARY OF THE INVENTION
The invention is a chemical separation method involving capillary electrochromatography (CEC) or a combination of CEC and capillary liquid chromatography (CLC). The method comprises packing a capillary, preferably a fused silica capillary, with a packing material, preferably silica particles. The packing material is coated with one or more linear carbohydrate polymer chiral selectors, preferably cellulose, cellulose derivatives, amylose and/or amylose derivatives, and more preferably cellulose tris(3,5-dimethylphenylcarbamate).
The packing process first involves loading a frit material into the capillary, the frit material being distinct from the packing material and preferably being an octadecyl silica product. The frit material is then sintered to form the first of two retaining frits, with the unsintered frit material thereafter being removed from the capillary. The packing material is then loaded into the capillary to form a stationary phase, with the retaining frit defining one end of the stationary phase. The packing material is preferably pumped into the capillary as a slurry at a pressure of about 430 bar or less. More frit material is then loaded and sintered to form a second retaining frit adjacent to the stationary phase at the end opposite from the first retaining frit.
A sample of analytes (a chemical mixture) is introduced into the capillary, after which a voltage is applied across the length of the capillary to achieve bulk transport of the analyte. A pressure gradient is also preferably applied to promote bulk transport of the analyte. The components (analytes) of the sample are separated during transport across the stationary phase by the differing flow velocities of the components, the component flow velocities being determined by the differing degrees of retention by the stationary phase and mass/frictional drag.


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