What are the chromatographic terminologies used in pharmaceuticals?
Even with the numerous technical terms involved, Chromatography is challenging enough to comprehend at the best of times. Here is a guide to help clarify some of the terms frequently used to ensure a better understanding of the chromatographic technologies.
Chromatography is a procedure by which solutes are separated by a process in which a system consisting of two or more phases. Out of which one is moving continuously in a system in a given direction and in which individually substances exhibit different mobilities by reason of differences in adsorption, partition, solubility, vapour pressure, molecular size or ion charge density.
The substance that adheres near the walls of the column; in GC that will be a liquid of high viscosity; in LC that will be a packing, gel-based or solid.
The eluate is the substance being passed through the column at the end of the separation. In gas chromatography, it is a gas, but in liquid chromatography, it is a liquid.
A tube which contain the stationary phase. The stationary phase differentially interact samples constituent components as they are carried along in the mobile phase.
Tailing factor or Asymmetry:
The factor describing the shape of chromatographic peak. The peak asymmetry factor is the ratio between the peak apex and back side of the chromatographic curve to the distance peak apex and front side of the chromatographic curve.
Capacity factor (K) is a mass distribution ratio often used to describe the migration rate of the analyte in the column.
The column efficiency (N) is a measure of the band spreading of a peak. The smaller the band spreading the highest the no. of column efficiency which indicates the good column and system performance. Column efficiency also called as theoretical plates.
Retention Time (RT):
The time between injection and the appearance of the peak maximum.
Relative Retention Time (RRT):
It is a measure of the difference of affinities of two compounds for the stationary phase; used in QC, reproducibility and method validation calculations.
Normal Phase chromatography:
The stationary phase is more polar than mobile phase is a normal phase chromatography.
Reverse Phase Chromatography:
The mobile phase is more polar than stationary phase is a reverse phase chromatography.
Volume of mobile phase required to elute the unretained peak.
Dead volume is the volume of the HPLC system between the point of injection to the point of detection.
Dwell volume is a volume of gradient HPLC system between mixing chamber and column inlet.
Buffer is resist the change in pH.
The pressure above gravity at the head of the column. Expressed in Psi, Bar, kg/cm2 etc.
Baseline is a detector response to the mobile phase.
Resolution is a measure of the separation of two peaks. This resolution helps to evaluate the separation capacity of the column.
It is a series of tests, performed to check the system performance during the analysis. (From start to end).
The solvent in which sample solution is prepared.
The mobile phase composition remains same throughout the run.
The mobile phase composition is varies over the run time.
The mobile phase remains same throughout the run by using two different reservoirs at different time interval.
|RT in Mins||Mobile phase A||Mobile Phase B|
Multiple chromatographic terminologies are available in the HPLC system suitability, tried to cover some of them.
Elution of same compound/analyte in the subsequent injection.
The amount of mobile phase that is passed through the column.
Absorption occurs in chromatography when a drug is partitioned into a liquid-like stationary phase.
It is the process of accumulating chemical substances on a surface.
The process of removing dissolved gases from eluent, carried out by helium sparging, applying a vacuum, ultrasonication or heating.
A peak has a leading edge on its front that is called a leading edge/Fronting.
Componants of HPLC:
- Sample compartment
- Column compartment
Why silica gel used in HPLC column?
Silica gel is inert material and does not react with mobile phase.
These are the chromatographic terminologies used in HPLC analysis.
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