Tl;dr - As of last week, the UV-Vis instrument at my job only records negative absorbance below 230 nm for any sample when zeroing methanol to correct for solvent. Attached spectra of the same solution before/after the issue arose. Metash claims methanol is the issue.
As of last week (3/28/2024), our UV-5500PC only records negative absorbances below 230 nm for our samples (ganoderic acids extracted via MeOH) when we zero with MeOH. When zeroing this way, the gain spiked from 3-4 to 7-8 below 230 nm. This had not previously been an issue with any samples in any solvent (Dist. H2O, MeOH, EtOH, Isopropanol, hexane). Startup calibrations and lamp checks (D2, W) always cleared with no issue. Same solution, before the issue: https://imgur.com/a/2F3BpPC, and after the issue: https://imgur.com/a/fm2U8u6.
We made sure clean quartz cuvettes were used, wiped the lenses with EtOH on a Kimwipe and allowed to evaporate, and when we zeroed with air, zeroing gain remained between 3-4 and wavelength scans of each solvent remained positive across our entire 500-200 nm range. We've been in contact with Metash, and followed through with their queries. Lamp energies are above their given threshold: Tungsten at 546 nm, input amp 1 was 18,090. Deuterium at 220 nm, input amp 1 was 11,020. Metash asserts that MeOH absorbs too strongly for use in the instrument.
I have doubts that is the problem, since we've been running wavelength scans for months prior to this development without the issue. There is no difference in absorbances of our new large bottle versus the squirt bottle of MeOH which might mess with zeroing. I don't think the lamp is going bad, since it's only affecting a segment of the lamp's range.
Has anyone encountered this issue? We recently had some construction in the lab, could there be physical damage/contamination which explains this occurrence?
When I run methanol while zeroed to air: https://imgur.com/a/LB9pfpP
I've also tried diluting methanol with water as a 1:10 and a 1:3 solution, in case the issue was how strongly methanol absorbs: https://imgur.com/a/AoelYLU
I scanned the other four solvents we have:
Distilled water: https://imgur.com/rQEwJrp
Ethanol: https://imgur.com/gjTRyAU
Isopropanol: https://imgur.com/nUzYua0
Hexane: https://imgur.com/275uiTm
My main concern in finding a solution is that our procedure extracts with methanol, and we didn't have enough product to do more than one extraction. Plus, the absorbance issue is relatively new and has not been a problem with previous methanol extractions.
Not sure if you resolved the problem but based on the spectra supplied it seems like pretty clear the problem is contamination of your methanol.
Methanol spectrum should look like the one shown here which is more like ethanol spectum you posted. https://www.shimadzu.com/an/service-support/technical-support/lib/methanol-acetonitrile.html
In addition the feature near 265 is complete unexpected in pure MeOH.
Negative absorbance always means more light is getting through your sample than your blank.
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