By Anonymous on Saturday, May 22, 2004 - 06:12 pm:

I'm working with:

MPA: 0.05% TFA in H2O
MPB: 0.05% TFA in ACN

TFA - Fluka
ACN - HPLC grade
H2O - UHP water

Column: 5-cm C18
Run: Gradient
System: Agilent 1100 HPLC
Wavelength: 272nm
Lamps: UV and Vis lamp are both switched on
Signal: DAD, ref: "off"
Pump: Quartenary

I've encountered a series of "V~V~V~V big waves" baseline now and then. It often happens initially when I switched on my system. When I switched off and on the lamps again. The baseline went back to normal but after a few runs (8 mins injection), the problem comes back.

I've also tried switching off the Vis lamp and leaving only the UV lamp on and the baseline looks okay. The detector lamps have just been changed. Is it a lamp problem or does it come from the pump?

Help would be much appreciated! - Fran

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By Anonymous on Sunday, May 23, 2004 - 10:59 am:

Several things come to mind.

1- An electronics problem with the detector (faulty pre-amp board or something like that)

2- An intermitent check valve failure

3- Outgassing in the detector flow cell

Call Agilent service and have them help you diagnose this.

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By Anonymous on Sunday, May 23, 2004 - 05:21 pm:

We've already called them but they can't seem to be able to find out why. It's been a second check from two engineers. No conclusion. - Fran

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By Anonymous on Monday, May 24, 2004 - 02:23 am:

How many hours had the UV lamp been working ??
I think that is a problem of de UV lamp ??

Change for a new one or form other HPLC 1100 and check it !!

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By Henrik Vogelius on Monday, May 24, 2004 - 05:26 am:

To find out if it is the pump or detector switch of the flow when you see the problem. Another thing switch the ref wavelenght on. Set the ref wavelenght close to the sample wavelenght with 50 nm bandwith without overlapping.

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By Anonymous on Monday, May 24, 2004 - 06:18 am:

Check that the pump pressure is stable and watch the waste line when priming for intermittant flow this can give you an idea that it is a pump problem.

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By Anonymous on Monday, May 24, 2004 - 06:35 am:

Hi Henry,

How do I determine "sample wavelength"? I'm having a stated 272nm I had to follow and I've also read your statement on similar topics.

Is your sample wavelength referring to 272nm in my case? And "with 50nm bandwidth" is referring to + and - 50nm from my stated 272nm, am I right?
- Fran

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By Henrik Vogelius on Monday, May 24, 2004 - 07:32 am:

Yes, but with the 50nm I mean +25 nm -25nm.
Quantification wavelength might be a better word for sample wavelength.
Please let me know if it helped.
- Henrik

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By Henrik Vogelius on Monday, May 24, 2004 - 07:44 am:

Hi Fran
If you still have problems, you can just switch the vis-lamp off, because if you analyse your sample at 272 nm you don't need it. You only need it for wavelength higher than 350nm. And if I understood you right you don't have this problem when the vis lamp was off

Best Regards
Henrik

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By Anonymous on Monday, May 24, 2004 - 09:07 am:

Step 1. Remove the flowcell from the light path. By changing only one thing at a time, you have a better chance of nailing down the culprit.

step 2. If the problem persists, replace the lamp(s). If it goes away, it's a flow issue.

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By Anonymous on Wednesday, June 2, 2004 - 12:05 am:

Hi Henrik,

I've tried switching off the Vis-lamp and it does helped. But I haven't had a chance to run analysis with the lamp switched off for my samples. I'll try it when time permits.

I've also checked the methods and unfortunately, it has stated that I must use a wavelength of 272nm with "ref=off". So, basically, I couldn't play around with that anymore. I'll try my own experiments and feedback to my seniors if it does consistently helped.

- Fran

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By Anonymous on Wednesday, June 2, 2004 - 12:07 am:

Let you guys know the results again sometime later?? And thanks very much for all the contributions. I've really had a headache thinking of where to search for help and lucky, I found this forum!

- Fran