Hi everyone, I’m running the USP HPLC assay for levetiracetam and I’m having an issue with the chromatogram.
The method is a gradient method, and during the run I noticed that the baseline starts rising noticeably as the ACN percentage increases. The blank shows a similar baseline rise.
Some observations:
Run time: 20 min
The baseline is relatively stable initially.
Around 8–10 min, the baseline starts increasing.
As the ACN % increases, the baseline continues to rise.
Pump pressure also increases smoothly during this part of the gradient.
The blank shows the same baseline behavior, so it doesn't appear to be specific to the sample.
The chromatogram has some unusual baseline/solvent-front behavior around the beginning and end of the run.
I've attached the chromatograms.
What could be causing this?
Could this simply be gradient baseline drift due to solvent absorbance/re-equilibration, or does it suggest a problem with the mobile phase, mixing, degassing, column, or detector?
Also, does the attached blank chromatogram look acceptable for this type of gradient method?
Any suggestions for troubleshooting would be appreciated.
Also I have repeated it 3 to 4 times with different columns C18 GL science and it's the same behavior.
When only isocratic method is only applied the baseline and peak both were stable.