to identify the most appropriate stationary phases. pSFC and CE are also used as a method screen due to their orthogonal separation mechanism to the reverse-phase liquid chromatography (RP-LC). Figure 14.5A illustrates different impurity profiles of the same sample generated from LC, pSFC, and CE method screening platforms. Some critical issues such as purification scale-up, buffer restriction for some bioassays, method transfer, and method validation are also taken into early consideration in this method screening stage. Based on the state-appropriate analysis concept and knowledge of the chemistry, one or two platforms can be selected instead of application of the whole system to avoid data redundancy.
There are three major challenges associated with the rapid development of chiral methods: (1) the significant number of compounds requested for chiral separation on a daily basis; (2) the wide variety of compounds that correspond to starting materials, intermediates, or drug substances; and (3) the achiral purity of the compounds can often be less than 90%, sometimes barely over 70%. Achiral impurities can severely interfere with the chiral-method development. pSFC is currently considered the primary choice over LC and CE for chiral separation. Previous approaches with SFC have not taken advantage of the MS selectivity and sensitivity to help develop chiral methods or to determine enantiomeric excess (EE%) in a complex mixture. Therefore, Zhao et al.  have developed a chiral screening strategy, which is illustrated in Figure 14.4B. The extracted ion chromatogram (EIC) function of the MS is used to distinguish the enantiomers. Sample pooling substantially increases the throughput without fear of interferences from achiral impurities, with the exception of structural isomers. This screen is performed under eight gradients on four chiral stationary phases (32 conditions). As an example, Figure 14.5B shows the chiral separation profiles of six pooled racemic samples obtained on a chiralpak-AD column at 20% B.
The downside of this multidisciplinary approach, however, is that a tremendous amount of information is produced, and consequently, data processing and interpretation are complicated. Therefore, another approach that can be used to maximize the resolving power of chromatography is to introduce a multidimensional separation with column-coupling or column-switching (CS) as an alternative procedure, especially for purity assessment.
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