Chromatography Forum

Hi everyone!

I have one question about the resolution...
We know, that the optimal resolution value for two separated peaks for different proteins has to be 1.5. That is well known for example HPLC columns, 15 cm long, that is filled with 5 μm size of the silica. But I also have UPLC column, 5 cm long, that is filled with 2,6 μm silica. I got perfect resolution between two narrow peaks in chromatogram, no tailing, the base line is reached between, but the resolution is only 0,96! How can I explain that? Is around 1.0 enough value for UPLC resolution?

Thanks a lot for the answer!

It does not sound like you are calculating resolution correctly. If your peaks are truly baseline resolved then the resolution will not 0.96

check your integration of peaks
one way to make look like things are good is to do a valley to valley between the peaks
make sure you do a straight line integration, then take a closer look at the baseline

Check the sampling rate and filter time constant. If you are using a TUV detector set sampling rate at 20 pts per second and use a normal filter time constant(0.200) if the resolution is still low try to use the slow filter = 0.100 lution.. Lowering the filter time constant improves resolution but increases the peak tailing.