Update in the Technology and Applications of Chiral Stationary Phases

Apr 01, 2008

An ever-increasing need for chiral separations has led to a more generic approach for screening a variety of chiral stationary phases. These new screening methodologies have been supported by new instrument development, new chiral product performance, and a new level of user knowledge. Supercritical fluid chromatography has continued to grow, supported by published applications from the separations industry. An expanded field of polysaccharide phases has been made available from a variety of sources with some unique variants of the most common cellulose and amylose derivatives.

In 2006, at the time of the last review (1), 80% of small molecule drugs approved by the FDA were chiral and 75% were sold as single enantiomers (2). Looking forward, it has been estimated that 200 chiral compounds will enter the development process each year. A variety of chiral technologies have expanded to meet this challenge, including asymmetric synthesis involving chiral auxiliaries, salt resolutions, chemical and biocatalysis, and chromatographic separations. Throughout these processes, versatile analytical methodologies are required. Because speed is essential, due to time and cost pressures and the growing number of projects, generic screening methods have become the preferred course of action. Generic methods are typically chromatographic separation methods employing chiral stationary phases (CSPs) that have broad selectivity capabilities. The pressure of developing an increased number of analytical methods ironically has led to a reduction in the number of suitable stationary phases tested for a particular separation and, in addition, a new view that preparative separations are in reality the best course of action for obtaining pure enantiomers instead of a method of "last resort."

Also during 2006, a major shift already was taking place in the approach to chiral separations. Blind screening of a number of chiral stationary phases was seen as the best approach to getting selectivity data in the most efficient manner. High performance liquid chromatography (HPLC) systems designed to screen from six to as many as 12 columns (PDR-Chiral, Lake Park, Florida) were infiltrating the market with a configuration of eight being the most common. Sepiatec (Berlin, Germany) and Eksigent (Dublin, California) have taken this methodology a step further in offering parallel testing of eight CSPs. In the interim, many consolidations have taken place within the pharmaceutical industry and within the chiral separations businesses, creating an even stronger concentration of effort toward greater efficiency in the separation process.

As the focus on chirality has brought the chiral separation process closer to the drug discovery platform, the need for chiral analysis increased and the requirements for even shorter method development times emerged. To respond to this growing need, the focus in HPLC centered on shorter columns and smaller particles using higher pressures and temperatures in the hope of achieving faster, more efficient analyses. Many of these approaches were not directly applicable to all chiral stationary phases. Certain of the CSPs require longer columns to obtain the necessary plate height for resolution while for other CSPs, higher temperatures adversely affected column stability. Some compromises had to be made. Temperature as an operating parameter has been studied extensively (3) especially for the CHIROBIOTIC macrocyclics glycopeptides phases (4,5). An alternative approach better suited to chiral separations was demonstrated by operating a CSP in a different mode. In liquid-phase methodology, two basic modes of operation have developed; HPLC and sub- and supercritical fluid chromatography (SFC). SFC utilizing super- and subcritical carbon dioxide modified with methanol or other solvents is seen as a potential major contributor to resolving many of the method development issues, especially relating to speed but also to environmental issues, both of major concern to the pharmaceutical and chemical industries. In addition, compared with HPLC, SFC also is considered to be a more "green" technique with limited amounts of organic solvent content in a carbon dioxide environment that also helps to make solute collection a cleaner process because the majority of the solvent (that is, carbon dioxide) merely evaporates to a gaseous state.

The limiting factor for SFC methodology centers on the polarity of the molecule. As the polarity of the molecule increases additional percentages of additives are required to promote elution and the process slows back down to conventional chromatography. Fortunately, the polarity of the majority of current molecules falls within the suitable range of normal phase chromatography and, therefore, of SFC.