Question

in reverse phase HPLC would you expect the least polar analyte to elute first or last? Explain your answer.

Answer 1

1.Using a separation mode based on polarity, the relative chromatographic retention of different kinds of molecules is largely determined by the nature and location of functional groups

2.oil is non polar and water is polar.they will not mix. chromatographic separations based on polarity depend upon the stronger attraction between likes and the weaker attraction between opposites. Remember “Like attracts like” in polarity-based chromatography.

3.Normal Phase HPLC
a polar stationary phase (like silica) is used and mobile phase is non polar. now if we put mixture of compounds in HPLC coloumn according to like attracts like principle polar molecules will be attracted by the stationary phase and will move slowly while non polar molecules feel no attraction and will move quickly .thus in normal phase HPLC non polar molecules will be eluted first.

4. Reversed Phase HPLC

a non polar stationary phase (like C18) is used and mobile phase is polar. now if we put mixture of compounds in HPLC coloumn according to like attracts like principle non polar molecules will be attracted by the stationary phase and will move slowly while polar molecules feel no attraction and will move quickly .thus in reverse phase HPLC polar molecules will be eluted first.