High-performance liquid chromatography is an analytical technique to separate, identify, and quantity components in a mixture. it is the single biggest chromatography technique essential to most laboratories worldwide.
2. What Is the principle of HPLC?
The principle of separation in normal phase mode and reverse phase mode is adsorption
When a mixture of components introduced into an HPLC column, their travel according to their relative affinities towards the stationary phase.
The component which is more affinity towards the absorbent travels slower
The component which is less affinity towards the stationary phase travels faster
3. Based on nodes and based on elution technique HPLC is how many types?
Based on modes of chromatography
Normal phase mode
Reverse phase mode
Based on elution technique
Isocratic separation
Gradient separation
4. HPLC system parts
Solvent reservoir
Degasser
Solvent delivery system
Injector
Column and oven
Detector
Recorder
5. What is the mobile phase In HPLC?
Mobile phase: the liquid that moves the solute through the column.
6. What is the stationary phase in HPLC :
The packing material of the column, which is the immobile phase involved in the chromatographic process.
the Column used as a stationary phase
7. What is retention time?
The time is taken by the analyst peak to reach the detector after sample injection
8. What are the advantages of the HPLC?
Speed, efficiency, and accuracy, compared to other chromatographic techniques, such as TLC, HPLC is extremely quick and efficient. It uses a pump, rather than gravity to force a liquid solvent through a solid absorbent material. With different chemical components separating out as they move at different speeds
9. Why do we get negative peals in HPLC?
A negative peak means that there is less absorbance while the peak is passing through the detector than When the mobile phase is passing through. It is likely due to the mobile phase has an absorbance than the analyst at the monitored wavelength
10. What are RT and RRT in HPLC?
The amount of time it took for the compounds to pass through the column is the retention time (RT)
The relative retention time (RRT) is the comparison of the RT of one compound to another
11. What is a USP tailing factor?
The tailing factor is a measure of peak tailing. It is defined as the distance from the front slope of the peak to the back slope divided by twice the distance from the centerline of the peak to the front slope, with ll measurements made at 5% of the maximum peak height.
12. What is the needle was in HPLC?
The needle in the HPLC system is used to introduce the sample into the mobile phase so that it can be separated on the HPLC column. The needle wash is used to clean the needle after an injection
13. What is the resolution in HPLC?
Resolution: the resolution of elution is a quantitative measure of how well two elution peaks can be differentiated in chromatographic separation. It is defined as the difference in retention times between the two peaks, divided by the combined widths of the elution peaks
14. What is the RRF value in HPLC?
Relative response factor(RRF) is an alternate method for the determination of the quantity of the impurities present in the pharmaceutical product and the amount of the impurity can be calculated with the help of the peak area of the components
15. What is degasses in HPLC?
Degassing is the term used to describe dissolved gasses coming out of a solution. This phenomenon can also occur in the HPLC system where rough surfaces produce nucleation sites for bubble formation. in HPLC analysis the problem produced by formation can largely be prevented, by degassing the mobile phase
16. What is SST in HPLC?
SST is commonly used to verify resolution, column efficiency, and repeatability of a chromatographic system to ensure its adequacy for a particular analysis, according to the united states pharmacopeia(USP) and the international conference on harmonization (ich) SST is n integral part of many analytical procedures
17. What is LoD and LoQ in HPLC?
Limit of detection (LOD) and limit of quantification (LOQ) are two important performance characteristics in method validation. LOD and LOQ are terms used to describe the smallest concentration of an analyte that can be reliably measured by an analytical procedure
18. What is integration In HPLC?
chromatography peak integration defines an operation on which the area under the chromatographic peak is the measured. The measurement is based on the integral technique of splitting the peak into a large number of rectangles, which are then summed to provide an estimate of the total area under the peak.
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