Enantiomeric Purification (HPLC/SFC)

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Enantiomeric Purification (HPLC/SFC)

The method for the purification of enantiomers by high performance liquid chromatography (HPLC) and supercritical fluid chromatography (SFC) is generally referred to as the continuous adsorption and desorption of compounds between the stationary phase (the column) and the mobile phase, so that different compounds can be separated. Because different compounds have different forces between the two (stationary phase and mobile phase). Some chiral substances and spatial isomers have almost the same polarity, so they are difficult to separate. Therefore, special chiral columns are needed to separate them for purification.

Enantiomeric Purification (HPLC/SFC)Fig.1 Enantiomers

BOC Sciences has state-of-the-art HPLC and SFC equipment, as well as a dedicated team of experts who can tailor-make your own chiral separation and purification services.

HPLC Chiral Separation Types

Chiral derivatization

The two enantiomers to be separated are reacted to generate a pair of diastereomers, which are then separated on a common chromatographic column (a non-chiral column). The advantage of this method is that the separation conditions are relatively simple, and only the conditions for chromatographic separation using ordinary HPLC can be used. After derivatization, the detection (UV or fluorescence) sensitivity is also improved.

The chiral additives are added to the mobile phase and a pair of diastereomeric complexes are formed with the enantiomers of the solute and separated on a common chromatographic column. Although the complexes formed by chiral additives and solutes are not as strong as those derived from the derivatization method, the diastereomeric properties of the complexes based on chiral recognition are essentially the same. Commonly used chiral additives are: cyclodextrin and ligand exchange additives. The advantage of this method is that it does not require pre-column derivatization and there are no special requirements for column packing.

The chiral stationary phase method is based on the difference in the energy or stability of the temporary diastereomeric complex formed by the chiral selector on the surface of the sample and the chiral selector, so as to achieve chiral separation. It is a method that does not undergo direct conversion into diastereomers. The advantages of the chiral stationary phase resolution method are that it is easy to prepare and can be applied to the resolution of various types of compounds. In spite of this, the chiral stationary phase method is still the most advantageous optical isomer splitting method. Up to now, the enantiomers of tens of thousands of chiral compounds have been separated by HPLC, far exceeding the sum of the enantiomers of about 7,000 chiral compounds isolated over nearly 100 years using conventional chemical methods.

SFC Chiral Separation Types

Direct method

The direct method includes the use of chiral stationary phases (CSPs) and chiral mobile phases. The specific operation is basically the same as HPLC. The chiral stationary phases of supercritical fluid chromatography are developed on the basis of the HPCL and GC chiral stationary phases. There are already a large number of commercial CSPs available. Generally, CSPs are classified into amides, cyclodextrins, and polysaccharides according to types of chiral selectors. Except for crown ethers and proteins, most CSPs can be directly used in SFC without any further improvement.

The indirect rule is based on chiral derivatization. The enantiomers are first converted to diastereomers and then separated by achiral stationary phases.

At present, the direct separation method of chiral stationary phase is the fastest growing field, and the indirect method is relatively less used.

Enantiomeric Purification (HPLC/SFC)Fig.2 Enantiomeric Purification

BOC Sciences has a large number of advanced HPLC and SFC equipment, and a team of professional technicians and experts can provide you with the chiral separation and purification services you want.

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