shifting retention times
Posted: Thu Jan 13, 2011 9:48 am
I am working with an dionex RSLC-MicrotofII, using a Agilent HPLC column Zorbax Eclipse XDB-C18, 2.1*100mm, 18µm.
Mobile phase A: 94 water, 5 MeOH, 1 acetic acid
Mobile phase B: 97 MeOH, 2 water, 1 acetic acid
I have put a standard on the column (alternariol monomethyl ether) and let it run 4 times. So it takes the sample 4 times from the same vial, with the same conditions.
Normally the retention time should be the same all the runs, but this is not the case... It even shiftes sometimes more then 1.5min.
The area for the 4 runs is the same, as also the intensity.
BUT: when I do another run, between the 4 runs and the 5th there is another sample (with other methode: positive modus instead of negative), the area is dubble the amount of the others, and also intensity is much bigger (10^5 instead of 10^3).
I have used 2 different columns to make sure the original column was ok, but it gives the same effect: different retention time, and when an other sample is inbetween the area is double the amount and intensity much higher.
Does anyone have some kind of experience with this or solutions...?
Mobile phase A: 94 water, 5 MeOH, 1 acetic acid
Mobile phase B: 97 MeOH, 2 water, 1 acetic acid
I have put a standard on the column (alternariol monomethyl ether) and let it run 4 times. So it takes the sample 4 times from the same vial, with the same conditions.
Normally the retention time should be the same all the runs, but this is not the case... It even shiftes sometimes more then 1.5min.
The area for the 4 runs is the same, as also the intensity.
BUT: when I do another run, between the 4 runs and the 5th there is another sample (with other methode: positive modus instead of negative), the area is dubble the amount of the others, and also intensity is much bigger (10^5 instead of 10^3).
I have used 2 different columns to make sure the original column was ok, but it gives the same effect: different retention time, and when an other sample is inbetween the area is double the amount and intensity much higher.
Does anyone have some kind of experience with this or solutions...?