Retention in a chromatographic column is achieved because the retained molecule adsorbs reversibly to the stationary phase. On a molecular level this means that it spends some time in the stationary phase.
Each solute molecule can be assumed to move independently of the other solute molecules. Furthermore, the movement is a mixture of a random process, i.e. diffusion or Brownian motion, and the movement caused by the flowing mobile phase.
Animation of the principle of column chromatography
The principle of chromatography is illustrated in the following animated simulation. The molecules are represented as small circles moving in an erratic manner and randomly stick to the column walls for a certain time.
In the first animation 400 small circles( 200 blue and 200 red) are simulated as they move erratically through the column. They all adsorb randomly to the column wall and stay there for 0.5 s whereafter they move further. The velocity of the mobile phase is 1.0 units.
Click on the "Inject" button to initiate the animation. Note: the limited number of "molecules" (=400) in these animations makes the chromatograms noisy.
What happens if the time the molecules spend in the stationary phase increases?
In the second animation the same number of circles is simulated as in the first. The only difference is the time they stay on the column wall, in this case 2.0 s.
A comparison of these two chromatograms illustrates the influence of the adsorption time, i.e. the retention factor, on the peak width. What is the reason for the broader peak in the second animation?
What happens if the velocity of the mobile phase increases?
In the next animation the adsorption time is again set to 0.5 s, i.e. as in the first simulation, but the velocity of the mobile phase is 3.0. Why is the peak broader in this case compared to the first animation?
Animation of a separation
In the next animation a mixture of 200 red and 200 blue circles will be separated. The adsorption time for the blu is 0.1 s and for the red 0.5 s. The velocity of the mobile phase is 1.0.