RID wavy baseline

[FBProc]

I've got an 1100 HPLC and I just installed a brand new Zorbax Carbohydrate column a few days ago. I ran through some methanol by accident (this shouldn't affect it?) but then I changed to 75% ACN and let it run through for a couple hours at 1mL/min. the baseline seemed good and stable. I ran it again today and got a really wavy baseline. Hasn't stablised even after letting it run for hours and purging the reference cell a couple times. WHy is this happening to a new column, it should work perfectly?

[kubowicz.tomasz]

Hello

RI signal is very dependent on temperature so make sure there is no air circulation around detector (draught or missing front cover)

First I'd check:
1.Pump - make sure that all components are working fine (valves), you can perform leak test to check it. If you run Water/ACN using MCGV (multi channel gradient valve) you can try to prepare mobile phase as one solvent (to check if it is not mixing problem)
Make sure that degasser is working fine.
2.Disconnect column and put zero dead volume union instead. Than flush both cells and monitor baseline to see if RI signal is stable (use pure water for RI flushing). If baseline is stable you can suspect your column.
3.If column is suspected you can try to clean it (read specification or use your own "cleaning" method)
Good luck

Regards

Tomasz Kubowicz

[Gerhard Kratz]

Which Zorbax Carbohydrate column do you run with which counter ion?

[Gerhard Kratz]

3-Aminopropylsilan is used. Flush with 20 column volumes of 75%ACN/25%Water. Keep the temperature under control.

[FBProc]

it's a C18 Zorbax Carbohydrate 250mm x 4.6mm x 0.5um. Not sure what you mean which counter ion?
pump is working fine and RI is working fine, it gives a stable baseline when I switch to the other column.
my other Zorbax Carb C18 had this problem so I switched it for a BRAND NEW one, I broke the seal on the packet. it was good for a short while but then the same problem occured. is it possible there is something contaminating in the flow path for the 2nd column? I flush through so much solvent 75/25 ACN for 8 hours, baseline is still wavy. how would you recommend to clean or recondition the column? it doesn't say anything about it in the instruction/specifications sheet

[FBProc]

heres some images to help explain the problem


here is what the baseline looks like realtime after flushing for about 6 hours with 75% ACN at 1mL/min


here is a blank sample which was ran on the brand new C18 column


here is what sugar standards look like being run on my brand new column

so bizarre what is happening?
Also I noticed the software straightens out the baseline post analysis, in realtime it's very wavy

[Hollow]

What you're seeing is the injection solvent peak.quite normal.
Did you dissolve your sample in mobile phase or is it different? if possible, use mobile phase as diluent. Make a blank inj with mobile phase and see if its gone.
if you don't have any peaks of interest at this retention time, nevermind about it and go ahead (if baseline is fine afterwards and integration is good)
I guess the carbohydrate column is a kind of amine/amide phase, so it works in HILIC mode (?).
therefore, if you need to move your peaks away from this, try to increase the % of ACN to get more retention of your analytes.