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What is the cause of a shift in the FLD signal during a run?

mvsteenl

Hi 

 

We often run amino acid analysis with a HPLC FLD (G1321A). But recently we have some problems with our chromatograms.

 

For the detection of primary and secondary amino acids, a different excitation and emission wavelengt is needed. So for the analysis a timetable for the FLD signal is set up:

 

 

Recently we have noticed that, at 17,40 minutes, when the FLD switches its signal, it also causes a shift of the baseline in our chromatogram. Which means we are not able anymore to detect our secondary amino acids.

 

 

Does anybody has expercience with such a problem?

 

Thank you!

 

MV

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  • 0

    That looks unusual, especially the peak with the flat top. In the Chemstation logbook there would be an ADC overflow error, I reckon? Not taking the baseline drop into consideration, the peak at 22min seems to saturate the detector. Remember, the highest peak has to be between 0 and 100 LU.

     

    I would run an intensity test, the signal/noise test and the dark current test. A wavelength accuracy test should be done after  thorough flushing with water at 2 ml/min for about 15 mins. Pulling the diagnostic buffers and a system report might be required later. The tests itself are run with a restriction capillary installed and a flow of 0.25ml/min of fresh water.

     

    Note that to be on the safe side we request to keep a distance between EX and EM of 40nm, so your parameters are borderline.

Reply
  • 0

    That looks unusual, especially the peak with the flat top. In the Chemstation logbook there would be an ADC overflow error, I reckon? Not taking the baseline drop into consideration, the peak at 22min seems to saturate the detector. Remember, the highest peak has to be between 0 and 100 LU.

     

    I would run an intensity test, the signal/noise test and the dark current test. A wavelength accuracy test should be done after  thorough flushing with water at 2 ml/min for about 15 mins. Pulling the diagnostic buffers and a system report might be required later. The tests itself are run with a restriction capillary installed and a flow of 0.25ml/min of fresh water.

     

    Note that to be on the safe side we request to keep a distance between EX and EM of 40nm, so your parameters are borderline.

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