A2 Analytical Chem Chromatography
A2 Analytical Chem Chromatography
A2 Analytical Chem Chromatography
KCH20501P
ANALYTICAL CHEMISTRY
CLASSIFICATION OF CHROMATOGRAPHY
TECHNIQUES BY CHROMATOGRAPHIC BED SHAPE
Column chromatography
Planar chromatography
Paper chromatography
Thin layer chromatography
TECHNIQUES BY PHYSICAL STATE OF MOBILE PHASE
Gas chromatography
Liquid chromatography
AFFINITY CHROMATOGRAPHY
Supercritical fluid chromatography
COLUMN CHROMATOGRAPHY
PRINCIPLES
Solid materials (Adsorbants) – Ability to hold the molecules at their surface
Attractive forces (Vanderwalls & Hydrogen )
Functional groups (Hydroxyl/ Aromatic)
Silica
B. PLANAR CHROMATOGRAPHY
Separation technique - Stationary phase is present as or on a plane. Paper – Paper
Chromatography
C. PAPER CHROMATOGRAPHY
Paper chromatography is a technique that involves placing a small dot or line of
sample solution onto a strip of chromatography paper. The paper is placed in a container with a shallow layer of
solvent and sealed. As the solvent rises through the paper, it meets the sample mixture, which starts to travel up
the paper with the solvent. This paper is made of cellulose, a polar substance, and the compounds within the
mixture travel further if they are less polar. More polar substances bond with the cellulose paper more quickly, and
therefore do not travel as far.
D. THIN-LAYER CHROMATOGRAPHY (TLC)
Thin-layer chromatography (TLC) is a widely employed laboratory technique used to
separate different biochemicals on the basis of their relative attractions to the stationary
and mobile phases. It is similar to paper chromatography.
Used to identify the unknown compounds and to determine the purity of mixture.
Advantages :
Simple, Rapid and Cheap
Faster runs
Better separations
Choice between different adsorbents.
Better resolution
Allow for quantification
TLC is very versatile; multiple samples can be separated simultaneously on the same layer, making it very
useful for screening applications such as testing drug levels and water purity.
GAS CHROMATOGRAPHY
Gas-Liquid chromatography, (GLC)
Mobile phase – Gas (Helium) Carrier Gas Pressure = 4 kg/cm2 Stationary phase -
Column, which is typically "packed" or "capillary".
The stationary phase is adhered to the inside of a small-diameter glass tube (a capillary column) or a solid
matrix inside a larger metal tube (a packed column).
Partition Coefficient of Volatile analyte between a solid stationary phase (Silicone) and a mobile gas (Helium).
AFFINITY CHROMATOGRAPHY
Based on specific & non-covalent binding of the proteins to other molecules – Ligands (His-tags, biotin or antigens)
Physical properties of the analyte.
Biochemistry in the purification of proteins (Enzymes) bound to tags.
After purification, some of these tags are usually removed and the pure protein is obtained.
APPLICATION
Use in industry primarily for separation of Chiral (Asymmetric Carbon atoms) molecules.
• Soman
• Glyceraldehyde
• Phosphours (Phosphine)
• Sulfar metal
• Cobalt
• Enkephalins
DETECTOR
Gas Chromatography or liquid Chromatography
To visualize components of the mixture being eluted off the chromatography column.
CONCLUSION
Chromatography is a process for separating components of a mixture. To get the process started, the mixture is dissolved in a
substance called the mobile phase, which carries it through a second substance called the stationary phase.
Keller, Roy A. and Giddings, J. Calvin. "chromatography". Encyclopedia
Britannica, 10 Nov. 2020,
https://www.britannica.com/science/chromatography. Accessed 1 June
2022.
C.F. Poole,
CHROMATOGRAPHY,
Editor(s): Ian D. Wilson,
Encyclopedia of Separation Science,
Academic Press,
2000,
Pages 40-64,
ISBN 9780122267703,
https://doi.org/10.1016/B0-12-226770-2/00021-1.
(https://www.sciencedirect.com/science/article/pii/B012226770200021
1)
INSTRUCTIONS FOR CARD GAME
- True / False
- True / False
- True / False
- True / False
- True / False
- True / False
7) Chromatography separate the mixture of dyes on the basis of their gravity
- True/False
- True/False
9) In paper chromatography , the LEAST soluble solute stays closest to the start point
- True/ False
- True/False
11) TLC is used to identify the known compounds and to determine the purity of mixture.
- True/False
- True/False
- True/False
- True/False