can be gained from an optimized solvent mixture will never be disclosed. Any evaluation of either a particular stationary phase, or solvent mixture, for the separation of closely eluting solutes must be carried out over a range of temperatures.       Optimum Operating Conditions for Chiral Separations in Liquid Chromatography Thermodynamic reasoning need not be used exclusively to examine a chromatographic problem but can also be employed together with other aspects of chromatography theory to achieve a practical goal. The following example shows how thermodynamics can be used with appropriate optimizing equations to identify the optimum conditions for particular difficult types of separation. In gas chromatography (GC), chiral selectivity is controlled by choice of stationary phase and operating temperature. From a practical point of view, chiral selectivity is achieved by introducing spatially oriented groups into the stationary phase molecules and, as a

is presently the most direct and technically viable way to resolve enantiomeric mixtures. The extremely high efficiencies available from modern chromatographic apparatus makes this approach the most effective. The use of GC for the separation of stereoisomers is not nearly so common as liquid chromatography, but nevertheless there are a number of very effective optically active stationary phases that can be used in GC for the separation of volatile enantiomers. The first effective GC chiral stationary phase adopted derivatized amino acids to provide chiral selectivity (18) in 1966. These types of stationary phase had very limited temperature stability and the optimum temperature for separation can often be greater than that at which the stationary phase was stable. The first relatively stable chiral stationary phase was introduced by Bayer (19) who combined the derivatized optically active component in a polysiloxane gum

for subsequent synthetic work (this can be particularly important in the separation of chiral mixtures). Thus, the amount of material that is separated does not necessarily determine whether the separation can be classed as preparative or not. However, all preparative separations involve the actual collection of an eluted component and does not merely comprise peak profile monitoring for quantitative estimation and elution time measurement. It is interesting to note that the technique of chromatography, originally invented by Tswett in the latter part of the nineteenth century, was not initially developed for analytical purposes, but for the isolation of some specific pigments from plant extracts. In fact, all the early applications of chromatograph were exclusively for preparative purposes and it was not until gas chromatography (GC) was introduced by Martin and Synge (1) was the technique used for analytical purposes. Even after the introduction of GC, liquid chromatography (

. The separation of the enantiomers of hexabarbital on this stationary phase by the direct injection of blood serum is shown in figure 56. Chromatogram A was obtained after 20 injections of serum and chromatogram B after 60 consecutive injections of blood serum. It is seen that here is very little column deterioration and that, although the tail of the major peak has become a little extended after 60 injections, the column could still be used very effectively for the analysis. Liquid Chromatography Applications Liquid chromatography has been used in an extremely wide range of analytical methods and it is impossible to give a comprehensive set of examples that would illustrate its wide applicability. The following are a few LC analyses that may indicate the scope of the technique and give the reader some idea of its importance and versatility. An example of the use of reversed phase chromatography (employing a C8 column) for the separation of some benzodiazepines is

. to prevent proteins from being denatured). Secondly, the stationary phase can contain molecules of a special shape that can only make close contact with molecules having a complementary shape. Other molecules can not interact so closely with the stationary phase and consequently, the stationary phase available to them will be restricted. This approach is exploited in chiral chromatography where the stationary phase is made to consist largely of a specific enantiomer that confers chiral selectivity to the distribution system Thirdly, the stationary phase can be attached to the surface of a porous support, and the pore size chosen to be commensurate with the size of the solute molecules to be separated. Under such circumstances the molecules that are smaller than the pores will enter the matrix of the material and have more stationary phase available to them. Conversely, the larger molecules will be excluded from the pores and, consequently, come in contact

Chiral Stationary Phases Modern organic chemistry and pharmaceutical research are becoming increasingly interested in methods of asymmetric syntheses. This enthusiasm has been provoked by the differing physiological activity that has been shown to exist between the geometric isomers of pharmaceutically active compounds. A tragic example being the drug Thalidomide, which was made available as a racemic mixture of N-phthalylglutamic acid imide. The important physiological activity