Chromatography Forum

We are using waters Oasis cartrides and observed a peak split in the sample chromatogram. Sample chromatogram has more than one peak which are split. We did not see peak split in standard prep because standard was not pass through SPE.
Method is validated and we have never seen peak splitting in the sample before.

What could be the reason for peak splitting? Could it be due to error during sample prep?
FYI: sample preparation steps
1)Precondition cartridge with water and methanol
2) laod the sample
3) rinse cartridge with water and discard the water rinses
4) loaded smaple is eluted with methanol and injected to hplc
Mobile phase : acn and water

Using pure methanol as sample solvent might be the problem. Are your standards dissolved in pure methanol too?

Peak shape problems (including splitting) are not unusual when the diluent is stronger than the mobile phase. Either add some water to your sample before injecting or decrease the injection volume.