HPLC signal at -500 mAU

[AriadneY]

What does it mean? I just read ACN using it also as mobile phase with H2O. I have already checked the flowcell and every part of the instrument. Moreover, when I changed solvent composition, the noise started again, after the first run in which the signal was perfectly flat. (first run: 36%ACN, 2nd run: 100%, 3rd again 36%)

(Instrument Agilent compact LC 1120, detector Uv-Vis)

Thanksss!

[tom jupille]

A "perfectly flat" baseline is *not* a good thing. There should be "some" noise (check the Agilent specifications; probably something like 5 X 10^-6 AU).

First off, are you running at a reasonable wavelength? If assume you have already attempted to zero the signal. If -0.5 AU is as close as you can get, the chances are you probably have one of the following problems:
- a severely mis-aligned flow cell
- severely solarized flow cell windows
- an occluded optical path in the flow cell (precipitated junk or material photolyzed and adsorbed onto the cell windows
- a dead lamp
- a bad electronic connection

You should be able to check the voltages from the reference and sample side photodetectors. If it's a dead lamp, then both will be zero. If it's a sample-side electrical fault, then that side will be zero but the reference side will be within spec. If it's a misaligned or occluded flow cell, then the sample side will be low (but non-zero) and the reference side will be within spec.