The State of the Art in Chiral Capillary Gas Chromatography

May 01, 2012
Volume 25, Issue 5

The present state-of-the-art in chiral capillary gas chromatography (GC) is reviewed in this article. The article provide a short historical overview of phase development followed by a comparison of GC and high performance liquid chromatography (HPLC) chiral separations. After a short discussion of mechanisms of the most popular cyclodextrin phases, practical recommendations on how to select the best stationary phase and then how to use it to optimize chiral GC separations are described. Examples of elution order reversal of racemates, injection of large volumes of diluted sample using retention gaps and other applications are also provided.

The origins of chiral gas chromatography (GC) technology can be found in an article by Volker Schurig and Leslie S. Ettre entitled "Emanuel Gil-Av and the Separation of Enantiomers on Chiral Stationary Phases by Chromatography" (1). For GC, there were two methods described, the direct approach utilizing chiral stationary phases with or without achiral derivatization and the indirect approach requiring derivatization of the racemate with a chiral reagent and separating the diastereoisomers on conventional achiral GC stationary phases. In today's world, the former is the preferred method given the extensive validation problems of chiral derivatization and the increased stability, reliability and diversity of available chiral stationary phases for capillary GC. In 1966, Gil-Av, Feibush and Sigler (2) achieved the first ever GC chiral separation using GC with N-trifluoroacetylLisoleucine lauryl ester chiral stationary phase (CSP) and separated N-trifluororacetyl alkyl esters of (±)-natural amino acids. Other stationary phases based on other amino acid or diamide derivatives have been developed and there are examples of chiral metal complexes that can interact with analytes through coordination complexation but none have been as successful or expansive as the cyclodextrin technology (3).

Chiral GC has seen a dramatic explosion of applications in many fields of study including natural products, asymmetric synthesis, biological studies, environmental contaminants, agriculture, food, flavour and fragrance with the evaluation of essential oils. See reference 3, in which 185 papers on chiral applications were reviewed using CSPs in capillary GC.

In this article, I attempt to review the types of columns that are currently available for chiral separations by GC, citing the advantages and disadvantages of each type. I also review the options available to solve a particular separation problem and the approaches to developing a separation. Optimization and troubleshooting tips are also included.

Advantages of GC Over HPLC

GC offers a number of advantages over high performance liquid chromatography (HPLC) for the separation of enantiomers including the ease of generating theoretical plate numbers ranging from 12000 to 60000, depending on column length, resulting in the separation of more pairs of enantiomers for each column; high speed of analysis meaning higher sample throughput; and with flame ionization detection (FID) as close to a universal detection method as you can get, so a lack of aromaticity or functionality of the target molecules are not negatives. The downside is that injected samples need to be quite clean of extraneous material (precolumns, so called retention gaps, can help here), have a vapour pressure below 250 °C (achiral derivatization can help here), and be thermally stable and not racemize (there are techniques that can help here such as lowering elution temperature with proper achiral derivatives). Please note that elution temperatures are related to analyte vapour pressure, not boiling point. There are also other advantages, such as short column equilibration time, ease of connection to mass spectrometry (MS) and ease of trace impurity quantitation.

Figure 1
Selectivity of other detection methods such as electron capture, the techniques of headspace extraction and the more routine use of two dimensional GC (GC×GC) have made the technique ideal for the analysis of enantiomers in complicated matrices like environmental, biological and agriculture samples. Development time and optimization are also faster than for HPLC. Increased thermal stability, high resolution, and increased peak capacity of current GC capillary columns make these tools ideal for the analysis of complex mixtures commonly encountered in samples from biological or natural sources. With current technological advances in CSPs there are often multiple choices for a particular racemate to maximize speed and increase efficiency leading to lower limits of detection. It is also easier to reverse elution order — that will be explained in more detail later. Also, despite the great success by the use of chiral LC, there are no chiral LC columns that can routinely separate a variety of volatile, nonaromatic enantiomers. And, of course, the biggest advantage is no waste solvent. The biggest drawbacks are the lack of preparative capabilities, the fact that the final analyte must be volatile and the inability to analyze thermally labile compounds.

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