Chromatography Forum

Hi all,

I am having a problem with one of the HPLCs I am working with. For no obvious to me reason i am having a number of base line "jumps" every now and again. It might happen every 5 minutes or once a day without a pattern. It happens with different mobile phases, different wavelenghts and different temperatures. Here is a chromatogram to give you a better idea



This particular chromatogram has been done on:
Agilent 1050 machine
Thermoscientific Hypersil Gold 150x4.6 column
Mobile Phase: 50/50 Methanol/Water
Wavelenght: 254nm
Temperature: 35oC

I have tried figuring out a problem myself, like phone calls, air conditioning, dirty mobile phase, pump fluctuations but nothing seems to help.
Did anyone else had a similar problem and knows how can i eliminate those jumps.
Thanks in advance

2 ideas:
- worn detector lamp. Do you see those jumps when you run without flow?
- leakage somewhere in the system. Can you record the system pressure? If there's a pressure jump at the same time as the baseline jumps than it looks like a leakage.

2 ideas:
- worn detector lamp. Do you see those jumps when you run without flow?
- leakage somewhere in the system. Can you record the system pressure? If there's a pressure jump at the same time as the baseline jumps than it looks like a leakage.

The pressure is constant, i have monitored it, it will rise or drop a few units from time to time but it does not affect the base line in this way. I do have a little issue with an AIV though, it is leaking (one drop in a few hours) from time to time, but those base line "jumps" appeared before an AIV problem. Lamp sounds as a good idea as its response is lower compared to the same runs/systems on different machine (same model, column, conditions).

Per HPLCaddict's suggestion to see what happens with no-flow. If it *is* flow related, you might be looking at small air bubbles gradually accumulating in the detector cell and then suddenly breaking loose. In that case, check your degassing or look for a leaky fitting on the intake side of the pump that might be aspirating some air.

Thanks for your replies,

I have testet wether it is flow related or not and it seems like it is not flow related. I am still getting those "jumps" even though the pump is off. On the other hand my AIV is dead now, so I will try replacing that first and see if it helps.

Not sure if you have a VWD or DAD detector, I'm not to familiar with the Agilent 1050.
I have seen a similar disturbance when one of our detectors had a failing Vis lamp.
Our test was run a 254 & 415 nm so we needed both D2 and Vis lamp on.
As you're only aquiring 254 nm if you've got the Vis lamp on turn it off and see if the problem goes away.

Not sure if you have a VWD or DAD detector, I'm not to familiar with the Agilent 1050.
I have seen a similar disturbance when one of our detectors had a failing Vis lamp.
Our test was run a 254 & 415 nm so we needed both D2 and Vis lamp on.
As you're only aquiring 254 nm if you've got the Vis lamp on turn it off and see if the problem goes away.

We are using VWD only. If I turn the lamp off I can not see anything

Not sure if you have a VWD or DAD detector, I'm not to familiar with the Agilent 1050.
I have seen a similar disturbance when one of our detectors had a failing Vis lamp.
Our test was run a 254 & 415 nm so we needed both D2 and Vis lamp on.
As you're only aquiring 254 nm if you've got the Vis lamp on turn it off and see if the problem goes away.

We are using VWD only. If I turn the lamp off I can not see anything

What ExLet was aiming at: If your detector has both D2- and Vis-Lamps, you might try to turn the Vis-Lamp off.

Thanks for your replies.

Problem solved by changing the lamp