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CHROMATOGRAPHY
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Use of Achiral Columns Coupled with Chiral Columns in SFC Separations to Simplify Isolation of Chemically Pure Enantiomer Products
Manuel Ventura, Ph.D.
Principal Scientist Amgen Inc.
Introduction
Often pharmaceutical intermediates require chiral purification by SFC after a reaction step to isolate enantiomers of divergent biochemical activity[1-4]. Following a reaction, achiral impurities are always present at some level relative to the desired product, often significant even after normal phase “clean up” separation in advance of chiral purification. Frequently, conditions with one chiral stationary phase (CSP) among an SFC screening set are found that separate the enantiomers and concurrently separate interfering achiral impurities such that an efficient scale up method is possible with one chiral column. In many cases however, co-eluting or closely eluting impurities can prevent efficient purification from being achieved in a single step. In cases of insufficient resolution from achiral impurities with one CSP, one or both enantiomers collected then require a second stage of purification. A separate mobile and stationary phase process is necessary to achieve an end product of adequate chiral purity and chemical purity.
If a stationary phase exists that yields the needed achiral selectivity with the same mobile phase used for a chiral separation method, it follows that the coupling of these phases can yield selectivity for all components in one chromatographic method [5]. Achiral and chiral columns have been coupled for preparative separations of mixtures containing interfering impurities with the racemate of interest. Alexander et al. described a separation of the stereoisomers of cinnamonitrile and hydrocinnamonitrile intermediates using a silica column coupled with a Cellulose tris (3,5-dimethylphenyl carbamate) chiral column [6]. Zeng et al. designed a sophisticated 2D SFC/SFC/ MS purification system with modifications to a mass-directed prep LC and custom software for control [7]. Using mass-directed fractionation [8, 9], the racemate peak is isolated in the first dimension using a 2-ethylpyridine achiral column, and then eluted from a trapping column with the mobile phase for the second dimension onto a chiral column for enantiomer resolution.
If the necessary achiral selectivity does not occur with the single SFC achiral phase, the benefits of these processes are lost. Since no universal SFC achiral column exists similar to C18 for reversed-phase
90 | | September/October 2013 - 15TH ANNIVERSARY ISSUE
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