peak at low wavelength

[rusatinsky]

I am using a simple isocratic elution (H2O/ACN = 50/50) at 1 mL/min on C18 column maintained at 30C. Injection volume is 50 µL and detection wavelength is 198 nm due to lack of chromophore in main analyte. There is a peak in blank diluent (mobile phase) at approximately 4 min. The peak is not present in electronic injection, significantly smaller in injection of water, doubled in injection of ACN. Interestingly, the peak from injection of MeOH is comparable to ACN. This peak interferes with elution of known impurity and therefore has to be removed. I tried a number of experiments (different lots of ACN, different HPLC vials, columns, needle wash, etc.) but the peak is still there. Of course, it’s significantly reduced at 210 nm but targeted sensitivity of related substances is lost as well. Can you advise on possible sources of that peak?

[carls]

Sounds like it could be coming from your injector. Try cycling the inject valve (load/inject) a few times during a run to see if you get the extra peak every time you cycle it. If so, you'll need to replace the rotor and perhaps the stator.

[mardexis]

It does sound like something is coming out of your injector. Do you have A/S or manual valve and which one?

Before you start throwing away rotor seals and stators try rinsing it well first. The peak is probably bigger with MeOH and ACN injections because they are better at washing the compound out of the injector than water or your mobile phase. Try rinsing with mL's of pure ACN and also try toggling the valve a few times while you rinse.

[carls]

If the rotor and/or stator are the source of contamination you'll have to at least take the valve apart and sonicate the rotor and stator. I typically use 50/50 methanol/water first followed by 100% methanol. If I go to the trouble of disassembling the valve I typically will replace at least the rotor seal.

[tom jupille]

Dissolved oxygen? Try injecting some degassed MeOH or ACN and see if that makes a difference.

[grisume]

Do you use plastic pipette tips to fill your sample vials?
We had a problem with peaks at low wave length with some sovents (ACN, MeOH).
The cause were some release agents and lubricants used in the manufacture process of our pipette tips washed out by our solvents.
Maybe try glas pipettes.