Chromatography Forum

We are experiencing an issue with our Agilent 1100 HPLC.
When similar samples were run in 2009, the peaks were large (the peaks of interest were 200-1600 mAu high). On the sample I ran yesterday, the highest amplitude peak was around 600. Most were much lower. The peak we are interested in was about 650 units high in 2009 but only 50 yesterday. Another biology research group used the HPLC in January and noticed that the peaks were much lower than when they used it several years ago. Is this an indicator that we need to change the bulb or would you recommend other options/tests? Our protocol runs at 205 nm.
I checked the lamp hours in the diagnostic screen in chemstation and this is what was reported back:

Accum UV on time 18387.20 h, 584 ignitions
Accum Vis on time 17884.75 h, vis lamp switch on 467 times.

The main user of this instrument told me he has never changed the bulbs.

Also we are noticing a lot of noise on the baseline compared to the 2009 experiments.

Is it possible for the lamp to have this many hours on it and still function?

Thanks for any advice.

Chris

Those are veritable Methuselahs among lamps! They are waaaay overdue for replacement.

That said, an old lamp is more likely to cause baseline noise problems than a loss in absorbance. If the detector hasn't been serviced in 4+ years, it's possible that the mirrors in the monochromator are showing some corrosion and/or the flowcell windows have solarized, both of which can decrease the absorbance.

It's time that system went in for it's 100,000 mile tune-up.

Strewth, I'm amazed the deuterium lamp still lights. Wherever that lamp came from, get the same again, it was clearly a very high quality product!

Yes, noisy base-line is highly likely to be due to dreadfully low light intensity. Do keep in mind that this may not be the only problem though; in theory absorbance is just what proportion of light is taken out by your sample, so it shouldn't vary with lamp intensity. In your case it may be so extreme that it does!