Extraction Technologies for Medicinal and Aromatic ... - Capacity4Dev

12 FLASH CHROMATOGRAPHY AND LOW PRESSURE CHROMATOGRAPHIC TECHNIQUES FOR SEPARATION OF PHYTOMOLECULES
• Collect the aliquot in the vial in position 2 of the collection
rack.
• Continue until all the components of interest have been collected.
Alternatively, multiple columns can be processed by replacing
collection vials at each elution step. A typical volume for each fraction is
two CV.
12.7.2 On-line Flash Chromatography
Using an automated system equipped with a fraction collector,
fractions can be collected in a variety of ways, for example:
• Fixed volume fraction collection.
• Individual peak fraction collection under microprocessor
control with UV detector input.
12.8 Low Pressure Liquid Chromatography
In low pressure column chromatography, a column of particulate
material such as silica or alumina has a solvent passed through it at
atmospheric or low pressure. There are different kinds of low pressure chromatographic
techniques:
i) Gel fi ltration chromatography (separation on the basis of
size)
ii) Ion exchange chromatography (separation of the basis of
charge)
iii) Affi nity chromatography (separation on the basis of specifi c
binding sites on the proteins)
12.8.1 Gel Filtration Chromatography
Proteins of different sizes are separated on a column in which
the stationary phase consists of polymerized agarose or acrylamide beads
with pores of particular sizes. A small protein in the mobile phase (aqueous
buffered solution) can enter the pores in the beads while a large protein
cannot due to size restriction. The result is that a smaller fraction of the
overall volume of the column is available to the large protein than to the
small protein, which thus spends a longer time on the column and is eluted
by the mobile solvent after the large protein.
12.8.2 Ion Exchange Chromatography
The material used for this type of chromatography consists of
an agarose, acryl amide or cellulose resin or bead which is derivatized to
contain covalently linked positively or negatively charged groups. Proteins
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