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Wednesday, October 2, 2019

How to Increase Retention Time in HPLC

The high-performance liquid chromatography (HPLC) is a technique of separation in chemistry which is used for quantitative and qualitative analysis of components. The retention time is the time required by a component to elute from the column and detected by the detector, generally the PDA, and UV/VIS detectors are used. The retention time of an analyte depending on the various factors such as length of the column, mobile phase composition, flow rate of analysis, the polarity of the sample and mobile phase, pH of the mobile phase and the sample, etc.

Each component has different retention times at a specific mobile phase composition, due to their affinity toward the stationary phase. In the reverse phase, HPLC the polar mobile phase and the non-polar stationary phase is used, thus the more polar molecule of a sample solution is eluted first from the HPLC column. You can increase its RT by changing the polarity of the mobile phase. The more polar mobile phase composition will increase the retention time of an analyte, and the more non-polar mobile phase will decrease the retention of the analyte.

To increase the retention reduces the flow rate of the system, it will increase the run time. Another way to increase the RT is the pH change, changing the pH of the mobile phase can cause changes in the selectivity and retention of acidic and basic compounds. As the ionization of these compounds is changed it is often the result of different interactions among the components and the column. But your ultimate goal should have a symmetrical peak shape.


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