By Anonymous on Friday, August 30, 2002 - 01:51 am:
Here is a quick question, we are currently running a prep HPLC system using a 30mm x 100mm C18 column at 5µm packing. Our conditions start at 95%H2O/5%ACN with 0.1% Acetic Acid in both. Our samples are injected at 300mg/ml concentration (max) per injection and our sample preparation solution is 50/50 DMSO/MEOH. We inject 1ml samples. Our columns don't seem to be lasting too long (~600 injections). Does anyone know the long term effects the starting solution (50/50 DMSO/MEOH) will have on the column.
By juddc on Friday, August 30, 2002 - 07:05 am:
How exactly is the column failing? High backpressure, lousy resolution? peak tailing? All three? What's your sample and are there any other matrix components beside DMSO and MeOH (excipients, proteins, etc...)? Have you attempted a column wash? From the info that you've given, I'd suspect that something in your sample is fouling the column and building up on it. Your sample solvent is significantly less polar than your starting conditions, which can lead to such a fouling issue. Not that I've tried it, but I wouldn't expect DMSO on its own to hurt your column. Maybe a wash with 50/50 DMSO/MeOH will help. Have you tried reversing the colunm and seeing what effect that might have?
Then again, some columns seem to be less rugged than others - maybe everything's ok and you'll have to live with it or get a column that works under those conditions for a longer period of time. Your column isn't dying after 10 or 100 injections, so whatever may be wrong (if anything) isn't catastrophically wrong.
Merely my two pennies...
By Anonymous on Sunday, September 1, 2002 - 02:32 pm:
I know this isn't really related to your problem, but I was a little troubled that your sample was made up in a stronger solvent than your intial m.p.
wondered if anybody else saw this.
By david garner on Tuesday, September 24, 2002 - 02:33 am:
This failing has been attributed to the sudden pressure shock due to the high viscocity of the DMSO solution. A guard column, or lowering flow rate during injection have both been claimed to help (no direct experience). On (or At) column dilution seems effective; this reduces the problem of injecting in an overstrong solvent (direct experience). You need another pump or an injection valve to do this.
Waters are particularly strong on all this at the moment (website and reps).
It may be that 500 injections or so isn't too bad overall; it won't be a huge component of your costs.