CHIRAL SEPARATION USING THIN LAYER CHROMATOGRAPHY

CHIRAL SEPARATION USING THIN LAYER CHROMATOGRAPHY

Chiral Chromatography Chiral - adjective: not superimposable on its mirror image: used to describe a molecule whose arrangement of atoms is such that it cannot be superimposed on its mirror image. Enantiomers - noun: mirror-image molecule: either of a pair of molecules that are a mirror image of each other in structure but cannot be superimposed.

Chiral Separations An enantiomer is one of two stereoisomers that are mirror images of each other that are non-superposable (not identical), much as one's left and right hands are the same except for opposite orientation. A racemic mixture, or racemate, is one that has equal amounts of left- and right-handed enantiomers of a chiral molecule. It important to resolve racemates as pharmaceutical activity is principally if not exclusively depend upon the one enantiomer or the other.

Diastereomer D-Threose D-Erythrose

As in HPLC β-cyclodextrins have approved particularly useful in the separation of mixtures of chiral compounds. These chiral-cavity type media act by allowing selective occlusion or intercalation of one enantiomer into chiral cavities in the matrix of the phase.

Chiral Chromatography Because of their chemical and physical similarity, enantiomers can be difficult to separate. However, they can be separated by chromatography, provided the system is also chiral. This can be achieved by the use of a chiral mobile phase, by a chiral liquid stationary phase or a chiral solid stationary phase.

Chiral Chromatography Mobile Phase The addition of chiral atoms to the mobile phase is one way to perform separations. If the chiral reagent form a complex of some type with one type of molecule of a pair of enantiomers, chiral separation may result.

Chiral Chromatography There are many solid stationary phases available. Brush type phases, cavity phases (such as cyclodextrins, crown ethers and macrocyclic glycopeptide antibiotics), protein phases, and ligand-exchange phases are common examples. Brush Type Stationary Phases.

Dual Phase TLC

Rod Thin-Layer Chromatography Instead of thin plates this method uses a thin layer of stationary phase coated onto small diameter quartz rods. The rods are spotted and developed in a similar manner to regular TLC plates After development and drying, the rods are automatically passed through a specially constructed flame ionization detector (FID) at a constant speed. The result is a chromatogram similar to a normal HPLC chromatogram.

Affinity Chromatography

Affinity Chromatography. Is based on the interactions between two components that are ideally suited to each other both electrostatically and spatially. One component is bonded to a solid support. The bonded component interacts with the analyte and the analytes is adsorbed from the solution. Molecules such as these do not match the ligand and are not adsorbed

Affinity Chromatography Affinity chromatography is the most specific chromatographic method and the separation in based on biochemical interactions such as: antigen antibody enzyme inhibitor hormone carrier

Affinity Chromatography Elution

Preparation of plates Glass plates are original support however, flexible plates have become increasingly popular. Most of commercial apparatus is designed for 20 5 or 20 20 cm, and those are now regarded as standards. Plates are cleaned before preparation and it is important not to touch plates with the fingers.

Usually xg of adsorbent and 2x cm 3 of water is used to make slurry. If a binder is used, the time available from mixing the slurry to completion of spreading is about 4min (after which setting will have begun). Buffer solution can be used instead of water to modify the properties of the layer. Similarly fluorescent indicators or other agents can by mixed with slurry. The standard thickness is 250 μm.

Spreading An absolutely even layer without any lumps or gaps is required. Commercial spreaders are of two types: moving spreader and moving plate type

Pouring If the adsorbent is very finely divided and of homogeneous particle size, and if no binder is used, a slurry can be poured on a plate and allowed to flow over it so that it is evenly covered. Certain type of alumina can be used for pouring, but water alone is not usually suitable for making the slurry; a volatile liquid ethanol or ethyl acetate is preferable.

Spraying They do not seem to have nay particular advantage over spreading. Difficult to produce even layer using this method.

Dipping Small plates, such as microscope slide, can be spread by dipping in a slurry of the adsorbent in chloroform, or other volatile liquid. Thickness and evenness of plate not known.