Technologies for Chiral Analysis and Separation

As one of the key elements for any pharmaceutical development program, analytic method development, validation...

Technologies for Chiral Analysis and Separation

BOC Sciences utilizes accumulated rich experience and the most advanced equipment to provide customers with rapid, economical and high-quality technical services for chiral problems.

The important molecules that make up organisms are mostly chiral substances, such as amino acids, carbohydrates, proteins, and nucleic acids. In fact, they mostly exist as single enantiomers. Since the different enantiomers of chiral substances have different physiological activities to organisms, it is very important to separate and synthesize pure single enantiomers.

Numerous studies have been focused on chiral analysis and the separation of chiral compounds resulting in various approaches to chiral recognition, analysis and separation, including crystallization, chemical resolution, kinetic resolution, capillary electrophoresis(CE), high performance liquid chromatography(HPLC), gas chromatography(GC), thin-layer chromatography(TLC), supercritical fluid chromatography(SFC), nuclear magnetic resonance(NMR), chiral ligand-exchange chromatography(CLEC), circular dichroism(CD), and molecular imprinting techniques(MIT). Chromatographic separation and analysis technology has shown great advantages in the separation and determination of chirality.

Our Advantages

  • Multiple separation and analysis techniques
  • Professional team and standardized laboratory
  • Efficient, accurate and low cost

The common chiral separation technologies could be classified into several subtypes as follow:

High Performance Liquid Chromatography(HPLC)

HPLC is ideally suited to analyze and separate chiral drugs. These drugs are transported by the body fluids after entering the organism to further produce action. HPLC can easily achieve online pretreatment of biological samples and can automate to a high degree.

HPLC is suitable for chiral analysis and separation with strong polarity and poor thermal stability. Direct and indirect methods are used for enantiomer resolution by HPLC. The former is divided into chiral mobile phase method (GMP) and chiral stationary phase method (GSP).The latter is the chiral derivatization reagent method(CDR).

Compared with other analytical methods, GC has the advantages of accuracy, speed, and strong recognition ability. It is suitable for the resolution of some enantiomeric compounds without aromatic rings that are usually difficult to separate and detect under HPLC conditions.

GC can also be divided into two methods: direct method and indirect method. The former is a chiral stationary phase method, which uses the chirality of the stationary phase to provide the required environment for separation. The latter chiral derivatization reagent method uses a chiral resolution reagent to convert enantiomers into diastereomers for analysis.

SFC is similar to HPLC in both hardware and software systems, and its mobile phase is a supercritical fluid, such as a ternary or quaternary mixed component based on CO2.

Compared with HPLC, SFC has the advantages of fast equilibrium speed, short analysis cycle, high throughput, low pollution, economical and practical, etc., and is suitable for the analysis of poor thermal stability and low volatile substances. Unlike in HPLC, temperature and pressure are important parameters in SFC and, hence, they have been used for the optimization of chiral separation in the modality of liquid chromatography.

There are two types of chromatographic columns, packed columns and capillary columns. Commonly used chiral stationary phases are cyclodextrin, metal complexes, polysaccharides, amides and amino acids.

CE has the advantages of simple operation, high separation efficiency and low cost, so it has become one of the important methods of chiral separation and analysis. At present, CE is a major trend in analytical science, and numerous publications have been appeared on the chiral resolution of a wide variety of racemates.

When separating enantiomers by CE, a chiral selective agent is usually added to the buffer first. The enantiomeric molecules can then form complexes with different stability with the chiral selector. The migration speed of these complexes is different, so the separation of enantiomers can be achieved.

Commonly used methods are capillary zone electrophoresis, micellar electrokinetic chromatography and capillary electrochromatography. Commonly used chiral selectors are cyclodextrin, crown ether, mixed micelles, cellulose, protein, polysaccharides, macrocyclic antibiotics and so on.

CEC is considered to be a good technique for chiral separations as it works on the principles of both liquid chromatography and capillary electrophoresis. CEC mainly performs chiral separation in three ways. The first method uses an achiral stationary phase combined with a chiral additive mobile phase, and its chiral selection depends on the chiral additive in the mobile phase. The second method uses a chiral stationary phase. The third way is to imprint the stationary phase with chiral molecules.

TLC is a chromatographic technique for the separation of non-volatile mixtures. Thin layer chromatography is carried out on a piece of glass, plastic or aluminum foil coated with a thin layer of adsorbent material, usually silica gel, alumina or cellulose. This is one of the easiest chromatographic techniques for the analysis of organic mixtures. It has the characteristics of easy operation, simple equipment, fast analysis speed, intuitive results and quick flow change.

CLEC is currently one of the most effective methods for separating chiral compounds with diligands such as amino acids, hydroxy acids, amino alcohols, dipeptides, sugars, nucleosides, nucleotides and derivatives. CLEC can be divided into two categories: chiral mobile phase additive method and chiral stationary phase method. The substance to be separated, the chiral stationary phase (or chiral mobile phase additive) and the central metal ion form a coordination complex (diastereomer complex). The difference in the relative stability of the diastereomeric complexes is used to realize the separation of enantiomers.

The principle of NMR for separating enantiomeric molecules is to convert enantiomers into diastereomers, or to provide a chiral environment to make the nuclear magnetic resonance signal difference between the enantiomers.

MIT is a technology for preparing recognition materials with selectivity and memory effect. Imprinting molecules are the target molecules we operate, such as drug molecules, vitamins, enzymes, antigens, etc. The identification material is a molecular imprinted polymer (MIP) with selective and memory effects prepared using molecular imprinting technology and imprinted molecules as a template. Because of its unique "predetermined" high chiral selectivity, MIP has a bright future in chiral separation, especially preparation separation.

CD only responds to optically active substances and has advantages in selectivity. In addition, CD has high sensitivity and good reproducibility, and is widely used in the analysis of chiral compounds. The separation and analysis of enantiomers often use chiral chromatography or capillary electrophoresis, but these methods require chiral columns or chiral additives, and circular dichroism can achieve the detection of enantiomeric purity under achiral chromatographic conditions.

Our Services

There are different variations of the technology for chiral analysis and separation. BOC Sciences has a large number of testing instruments and technologies. Our experienced scientists will select the most appropriate methods and provide 100% guaranteed service for customers. Typical services include:

  • Chiral separation service
  • Chiral Analysis report
  • Technical consultation


  1. Kim, J. M., Chang, S. M., He, X. K., & Kim, W. S. (2012). Development of real-time sensitive chiral analysis technique using quartz crystal analyzer. Sensors and Actuators B: Chemical, 171, 478-485.
  2. Guo, H. S., Kim, J. M., Kim, S. J., Chang, S. M., & Kim, W. S. (2008). Versatile method for chiral recognition by the quartz crystal microbalance: chiral mandelic acid as the detection model. Langmuir, 25(2), 648-652.
  3. Subramanian, G. (Ed.). (2008). Chiral separation techniques: a practical approach. John Wiley & Sons.

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