Chromatography Forum

I am running samples at very short wavelength, around 210 nm. I got a whole bunch of peaks from 1.5 min to 6 min in blank injection (20 ul 50/50 H2O/ACN). Is it likely that those are just solvent peaks considering the short wavelength? Could I possibly have solvent peaks at almost 6 min?

Do you use mixer? I mean 2 pumps? This may be a mixing problem. Thought I am not the expert yet but I work only with that wavelength with my peptide and 20:80 ACN:H2O or 48:52 MeOH:H2O. I even had good looking peaks of my peptide at 205nm. If I were you I would clean injector by injection pure ACN several times without column and observe the spectrum.

? There's mo files or pictures there.

1. Try injecting mobile phase.

2. Try injecting nothing ie just activating the injector.

These experiments should indicate if the system is the problem.

If these are OK it must be something in the blank or a disturbance in the column caused by the blank being different to the mobile phase.

Can you give more details ie column/mob phase/blank.

the above chromatogram is an injection of mobile phase (50/50 ACN/H2O)

our system doesn't allow zero volume injection

what do you think?

Try injecting the lowest possible volume ie 1ul. It would also be interesting to see if varying volumes give different result.

What is your to? Looks like you are injecting air. Strange apparatus that doesn´t allow zero injection. Possibly you can trick it to do so?

I think that you have dirty injector. Yesterday I used sample dissolved in buffer (acetic acid/TEA) and ACN. Today I had to inject 200 ul pure ACN several times before I didn't get any peaks. Mobile phase was also pure ACN. I also discovered that in my HP 1090 in injector, which is glass, there is sometimes a bubble of gas. Maybe this is the problem.