Disappearing Peak

RHuber

<p>Hello,</p><p>I have a method for tocopherol acetate. At the beginning of a run everything looks fine, all of the standards look good. But at the end of the run, the standards are injected again (same vials) and no peak. If I start the run again, the peak is there, but gone again at the end of the run. The retention time does not drift and the peak area is consistent for multiple injections. The method is a gradient method starting with 10:90 0.1% Acetic:MECN to 90:10 0.1% Acetic:MeCN.</p><p></p><p>This issue started after the installation of a sample organizer and an instrument PM. It is also a new method, but I have been having some odd results with other methods. So I am not sure if this is an instrument or method issue.</p><p></p><p>Any ideas why the peak disappears?</p><p></p><p>Thanks,</p><p>Rob</p>

lizh

These types of problems are always tricky. First of all you have lost confidence in the instrument, so a good first test would be go back to a known good method and see if all is well. Then can you elaborate on the "peak is gone" is it mising. Does this mean when you re-inject from the same vials the peak is not there, sorry to be  apain. Can you forward the instrument method and details. This is an interesting one.

Many TX

Liz

RHuber

I am running another method today, so tomorrow I will have a better idea whether it is instrument or method. I will let you know what I find out.

Correct...when I inject say a 10ppm standard at the beginning I have a good peak, but an injection from the same vial at the end of the run results in no peak. If I reinject the sample the peak is there again, but if I run the entire sample set, it is gone again by the end. The majority of my unknown samples have no detection of the compound, but since the end standard also has no injection, I can not trust the results of the unknowns.

The sample diluent is 100mM tris buffer pH 7 (samples are from a degradation study of some material). The analyte has low solubility in the solution ~40ppm, so this may also contribute to the issue.

The gradient method is as follows:

Initial    10     90          0.1% Acetic : 0.1% Acetic in MeCN

0.5       10     90

3          90     10

6          90     10

7          10     90

Injection - partial loop w/ needle overfill (5uL)

Run time - 8 min

RT - 4.2 min

The reason for starting with such a low MeCN concentration is that I am worried that I may precipitate the sample buffer. It is intended to flush the buffer through prior to introducing too much organic.

Thanks,

Rob

lizh

Hello

I ahve asked for some experienced help here and their thought is that the sample might be attaching itself to the walls of the vial or precipitating out of solution. You seem to be seeig solubility issues. What is not clear is whether you are ever able to get a peak out of a particular vial once it has gone missing. If the answer is no, we suggest adding a little acetonitrile (up to, maybe, 10%) to any of the standard vials and re-injecting. If the peak returns,you will  need to take steps to keep this sample in solution while it sits in the vial. What do you think?

BR

Liz

RHuber

Well...it was not an issue with the instrument as several runs using another method were fine. I is something with this method. I agree that it is likely a solubility issue. I will work with the person submitting these samples to see if they can be put in a more suitable solvent.

Thanks,

Rob