CHROMATOGRAPHY Flashcards
What is chromatography?
Technique for separating related chemical compounds of a mixture
Now also used to quantify compounds – single or multi-component.
What is retention in chromatography?
Non-retained compound moves with solvent front; different degrees of retention required for separation.
How does particle size affect chromatographic separation?
Smaller particle size per unit weight of stationary phase gives better and more efficient separation.
What are the two main types of chromatography used in pharmaceutical analysis?
- High performance liquid chromatography (HPLC)
- Gas chromatography (GC)
What are the main sections/topics covered in chromatography?
- Introduction
- Instrumentation
- Separation mechanisms
- Stationary phase
- Mobile phase
- Sample – choice of separation parameters
- Detectors
- The Chromatogram
- HPLC instrument care
What is chiral interaction in chromatography?
Separation of isomers based on chirality; R or S enantiomer is immobilized on silica or other polymer.
What is gel filtration in chromatography?
Separation mechanism for high molecular weight compounds (>10,000) based on ability to penetrate through pores of stationary phase.
What is partition in chromatography?
Employs the phenomenon of partition coefficient; compounds with different P/Cs will move at different rates along the column.
What materials are commonly used in ion exchange chromatography?
- Chemically modified silica
- Polymers of styrene-divinyl benzene
- Ionic groups introduced on resins
What are the key interactions involved in adsorption chromatography?
- H-bonding
- Non-bonded interactions
- Dipolar interactions
- Van der Waals attractions
What is the purpose of HPLC instrumentation?
To perform high performance liquid chromatography, involving components like pump, injector, column, and detector.
What is isocratic elution?
Same concentration of mobile phase through the run.
What is gradient elution?
Mobile phase concentration varies with time; uses binary or two pumps.
What is the purpose of quantification in chromatography?
To separate and identify components in chromatogram using standards of known purity.
What are some examples of stationary phases in normal phase chromatography?
- Silica
- Cyanopropyl silica
- Aminopropyl silica
What are some examples of stationary phases in reverse phase chromatography?
- Octadecyl silane (ODS, C-18)
- C-8
- C-4
- C-2
- Phenyl
What are the least polar mobile phase solvents?
- Fluoroalkanes
- Hexane
- Isoctane
- Carbon tetrachloride
- Toluene
What types of compounds can be used as samples in chromatography?
- Polar to non-polar
- Ionic and non-ionic
- Small molecules to macro-molecules
What is the role of the detector in HPLC?
To detect and quantify compounds based on their interaction with light or other properties.
What is the formula for efficiency (N) in chromatography?
N = 16 (tR/Wb)² or N = 5.54 (tR/Wh)²
What does the resolution (Rs) formula represent?
Rs = 2 ΔtR/(W1 + W2)
Where ΔtR is the difference in retention time and W1, W2 are the widths of the peaks.
What is the significance of the chromatogram in chromatography?
It provides a visual representation of the separation and quantification of compounds.
What is the recommended pH range for silica-based columns in chromatography?
pH 2 – 7.5
