difference between low pressure gradient and high pressure gradient hplc

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lakshminarayana
lakshminarayana's picture
difference between low pressure gradient and high pressure gradient hplc

hai i am lakshminarayana is there any change in retention time by using low pressure gradient and hight pressure gradient hplc
regards

Dr. Analytical
Dr. Analytical's picture
Yes, there can be some

Yes, there can be some differences in retention times when using gradients with high pressure and low pressure mixing systems.  The difference is caused by the "dwell volume" in the two systems.  Dwell volume is the physical volume of liquid between the point of mixing and the column.  On a high pressure mixing system, the mixing occurs after the pumps, and usually there is only a small dwell volume.  On a low pressure mixing system, the mixing occurs before the pump, and the dwell volume includes the volume in the mixing chamber and the low pressure tubing leading to the pump, in addition to the tubing after the pump.  This volume is usually much larger than with high pressure mixing, and can be 1 mL or more.

When you program a gradient, the actual composition that you program is delayed in reaching the column, becauase it has to pass through the dwell volume.  On a low pressure system, this take longer, so the column see the specified composition at a later time.  So, usually the retention times will be later on a low pressure system.  However, the early eluting peaks are usually affected more than the later eluting peaks.

Remember that these difference are not important in isocratic separations, and retention times should be equal.

Thanks for posting on scientistsolutions.com.

sachintailor80
sachintailor80's picture
 what id gradient

 what id gradient