The elution strength increases throughout separation by gradually changing the mobile phase composition in gradient elution. Thus, the time of analysis is decreases and the quality of separation and detection limit improves. Even if the gradient method is more complex compared with isocratic elution.Advantages of Gradient Elution:
- Gradient HPLC mode has ability to separate complex samples.
- This method gives increased sensitivity.
- Gradient elution has shorter analysis times and gives much narrower chromatographic peaks.
- It gives an improved resolution.
- It can reduce the column degradation because of strongly retained analytes.
- This is the best method compared to an isocratic elution to clean the column.
Disadvantages of Gradient Elution:
- To flow the mobile phase in gradual elution, a special HPLC pump is required, which is more expensive.
- Between consecutive injections, it needs to be re-equilibration with the initial mobile phase composition.
- There is a method transfer problem since instruments vary in their dwell volume.
- Precipitation is possible when using multiple proportional valves.
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