Gradient problems....

<p><span>Hi all,</span></p><p></p><p><span>We are having few gradients problems where we can see small "steps" during the gradient part of the run. We are using a PDA detector and our mobile phase is a) 38:62:0.1 b) 78:22:0.1 ACN:H2O:TFA .</span></p><p></p><p><span>Any ideas where this problem came from?</span></p><p></p><p><span>See attached file for Chromatogram..</span></p><p></p><p><span>Thanks</span></p>

Answers

  • Hello:

    This feels like a pump issiue or possiblly a mixing issue. The first thing would be to look at the console traces of pump pressure - are the "jumps" corresponding to changes in the pump perfromance, look at the system overlays in the console.

    You may not have the console data available, as it saves 96 hours and then over writes, so alternatively run just your gradient with a blank injection - no sample and ensure that you save the pump data (channels tab in the pump instrument method) and see what this reveals.

    regards,

    Liz

  • Hi Liz,

    There are no "jumps" on the pump pressure corresponding to the plateaus. The system overlays in the console looks fine too. Any other ideas. Thanks

  • Hello:

    Actually that is good, we have ruled out the pump, now, since the chromatogram has not changed, just the baseline it could be the detector. Can you send me your method and can you tell me what your reference energy/autoexpose and the type of detector.you are using.

    Many thanks

    Liz

  • Hi Liz,

    We have a PDA detector at 217nm. We had been running this method for very long and we never have that problem. What other information you will need about the detector?

    Thanks again

  • That's why we are a bit confused here, we suspect the detector as the baseline is skipping, but I am not sure why so can you send us the full instrument method details and the autoexposure time for the injection before and after the one you showed. PLease send the specifics of the wavelength channel that you pulled out to show us.

    Many TX

    Liz

  • Hi Liz,

    This is all the information that I get from the instrument methods:

    We are running PDA @ 217 (not 3D)

    Sampling Rate 20 points

    Filter time constant: normal

    Exposure time: auto msec

    absorbance @ 217 4.8 nm resolution

    Let me know if this is what you were looking for?

    Thanks again,

    Maria

  • Almost, what might be better, in the Console at the systems view under troubleshooting menu - you can create a service profile, can you do this and post (or email privately to me) what ever suits. I was looking for the SM, BSM method data too and the chemistry details. Additionally the value chose by autoexposure is very helpful for diagnostics - it should be the same value run to run. However, the profile will collect much of that data for me automatically.

    Many TX

    Liz

  • Hi Liz,

    Guess what?...we change and put back the old lamp and all the plateau dissapeared.... test injections look great. Now any ideas of why the new lamp (6 months) was doing that?

    Thanks so much for all your help.

    Maria

  • Hello:

    Once we had rules out the pump, that's why I was asking for further data on the exposure time - that is the reference/raw lamp energy. It looks like the lamp energy was changing. The console data would tell me how many hours and what the energy levels were on the lamp

    I was speculating bad lamp, or a power supply issue. I have sent the email off to a specialist and will get you his thoughts.

    Many Thanks

    Liz

  • Hi Liz,

    Th eplateau are back again. Someone from Waters expend the whole day trying to figure out what going on, however seems like nobody run into this problem before. They change the check valves, and all the lines in the pump....at the end of the day the plateau were missing however the baseline is very noisy. I leave the system overnight running....

    Tomorrow I will see what we have again.

    So, please keep me posted of what the specialist have to said about this problem.

    Thanks so much

    Maria

  • Hello

    I did talk to them and it's possible, though very rare, that the lamp is defective and is emitting an unsteady flow of light energy. But I can't think of the last time that happened. Now the phenomenon is back, again, can we get that service profile and exposure time values.

    Liz

  • One final point, RDE got back to me and based upon the reoccurance and they suspect the electronics, so I sent the information on where to look, (probably the personality card) to your FSE who should be in touch.

    Have a great weekend,

    Liz

  • Hi Liz,

    Thanks so much! I didn't have time today to copy that file to send it to you. If you think it can help I can send it on Monday. The system is running with a lot of noise also.

    What "RDE" and "FSE" stand for?

    Thanks again and have a great weekend too!

    Maria

  • I am sorry, no I will not need, I think we are all set, once the FSE is back in, best regards,

    Liz

  • Hi,

    RDE: Research, Development & Engineering

    FSE: Field Service Engineer

    --Doug

  • Hello:

    I wonder if there is a lamp issue here, regular steps like this seems we should look at the pump as well as the detector.

    First thing is to look at the pressure trace, does this correlate to a pump pressure channel anomally?

    I would then check the lamp energy, what is the value for exposre time?

    Best reagrds,

    Liz

  • Hi all,

    We send the detector and the pump back to waters...it will take couple weeks. I will post what they found.

    Thanks for all the help.

    Maria

  • Hi

    We had a training by Waters techincal personnel and we rise this issue. As per his thought if you are using TFA in the mobile phase. You need different mixture. Bydefault aquity has 100uL size, but if you use TFA in the mobile phase you might need bigger size mixture. Ask about this to Watersyou will not get this problem again. After this we kept one system dedicated for the use of TFA.

    this might help you.

    J

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