What are the factors on which separation occurs?

[Troubleshooter]

Hi,

I have a confusion regarding basic principle of separation, whether separation is on the basis of polarity and temperature or other factors are also there, if yes then what are they and how much they contribute? I think polarity is the basic theme as we use gradient as well or in method development we use this factor foremost.

Regards,

Kashif

[AdrianF]

This is far too big a topic - you are basically asking how does HPLC work!!

Try the FAQ at the top of the page where you will find a great deal of information about HPLC. You will then be in aposition to ask more specific questions.

[DR]

Most chromatography is based on exploiting the differences between partition coefficients among a group of analytes in a given system.

Typical system critical parameters include mobile and stationary phase composition (where polarity commonly figures in), temperature, pH and (generally to a lesser extent) flow rate, pressure, column geometry. All of these can be manipulated to help maximise the separation between a pair of analytes.