Extraneous Peaks - Zero Volume Injection

[Firedevil]

I have a Waters 1525 pump with a manual injector. I have noticed some extraneous peaks in my analyses, and they usually don't interfere, so I have largely ignored them. Now I am looking at some very polar components and they have become a problem.

To figure out the source of my extra peaks I took the needle out of the equation, hoping I had contamination in the needle. In that case, a zero volume injection would contain no extraneous peaks and all would be right with the world when I cleaned my current needle well enough, or with a new needle.

Alas, a zero volume injection where I just flip the manual injection valve from "Load" to "Inject" (to start the date collection) shows between 3-5 peaks. I started the injection, saw the peaks, then wondered what would happen if I flipped the valve in the middle of the run. Consistently at about 45-50 seconds after I flip the valve, I see 3-5 peaks again.

I've tried rinsing my injection port with 100% water to wash off any possible salts, followed by 100% organic solvent. I still see these peaks, and am at a loss..... Anyone else have any bright ideas, or have ever experienced this problem?

Thanks for any help you can give me!

[tom jupille]

That's a *very* common problem with gradients. There is a mini-seminar on our web site that covers the causes, diagnosis, and some cures here.

[bisnettrj2]

This seems to be due to the mobile phase disruption due to the injection valve switching from the load to inject positions. Depending on your column dimensions, you are probably seeing disruption at the void volume of your column - if you are trying to quantify peaks in that region of your chromatogram, you are going to have problems with this phenomenon and other interfering peaks when you start running real samples.

[Firedevil]

Thanks for the replies. I am actually running an isocratic method 50:50 ACN:H2O with 10mM TEA.

The disruptions are occurring right around the void time of the column. Unfortunately, one of the 40 analytes in my research project is resorcinol, which is very polar and elutes very close to the void, and to top it off, I am trying to do trace analysis. These peaks are very disruptive.....

I can (will) decrease my ACN concentration and see if I can pull the very polar peaks further from the void, but I am not holding my breath that this will work. I'll let you know how it turns out.

[Bintang]

Manual injector and research, does not sound like you are doing high troughput:-). If you are not afraid to double the amount of work there is an alternative way of getting around this problem, use a different chromatographic method for your polar analytes. HILIC will give you high retention for those and the hydrophibic stuff will come in the void.
OK this approach will have you doing twice as many injections but it will work.

[Firedevil]

You wouldn't believe my saga if I told you. I will be doing extremely high throughput, but suffice to say, all my instruments with autosamplers are down until we can afford to fix them.

The HILIC advice is sound, however, I have a broad mix of ~40 compounds I will analyze ranging from resorcinol (very polar) to butylbenzene (very nonpolar). If I fix my polarity issues for one compound, I screw them up again for another.

Thanks!

[bisnettrj2]

What's the pH of your mobile phase? I took a quick Google trip through the internets and found a few papers and application notes with resorcinol retained on C8, C18, phenyl and PFP phases with an acidic mobile phase.

[Firedevil]

Neutral pH. We are not adjusting the pH of the mp for various reasons.

I'm busy with school extracurricular activities until Wednesday, can't wait to get back in the lab and try weaker MP to see if it moves my polar compounds away from the solvent front enough to not be interfered with.