Chromatography Forum

Hi everybody.

I use Waters 2690 with RI detector. My mobile phase is ACN/H20 80/20 and I premix it. I use Nh2 column from Kromasil.
Recently I changed my column and I see that I have to purge the detector for over 2 hours in order to have a stable baseline. Previously I needed only 5-10 minutes. Do you have any suggestions?

Hi Kalaikon,

Your query is generally vague, to receive some more help I would recommend providing more information about your analysis, the new column you installed, if you have any purge-steps following installation, etc. This issue can manifest for a variety of reasons so being specific will help us help you better. Check out this post in blogspot about common baseline disruptions and see if any look like your issue: https://hplctips.blogspot.com/2014/09/c ... noise.html

It's hard to usher a reason without knowing more, but I wonder is this "other" column also an NH2? Is this a new column (straight from the manufacturer) or another column that you have? What does your analysis look like/what are you actually running on these columns? Are there two different methods being used on the two columns?

Please get back to us with more information so that we may be a better help to you!

This is what I'd do when I had to run an assay using RI detector:
We had quaternary pumps (Agilent 1050 and 1100). I'd pre-mix the mobile phase.

I'd bypass the mixing valve with the little adapter.

I'd run the system overnight in the recycle mode, routing the flow from the detector back into the mobile phase reservoir.

I found this to produce stable baseline and good results.

I run sugar analysis. The new column is the same with the old now, from the same manufacturer too.

RIs take a long time to equilibrate with respect to temperature stability. Ideally, you just don't shut them off unless you know you won't be using them for days on end.

That said, I briefly flush the flow cell w/ new MP, then switch to purge and don't switch back until I'm about ready to start my run.

New Amino columns are shipped in hexane which is incompatible with your mobile phase composition. The fix is to flush the new column with IPA first to remove hexane and then with your mobile phase until the baseline is stable.

I flushed the new column with isopropanol, before the use of the original mobile phase. I will try it again and see if something changes.

Purge time of the RI detector used to be around 5-10 minutes only. Now I need 4-5 hours minimum.

My concern is that the column stationary phase is NH2. So it is hydrolised with the aqueous mobile phase and I believe that it will reduce the column lifetime pretty fast

The detector should be disconnected from the column when flushing. You can flush the column with IPA first followed by ACN and then 80-20 ACN-water.

100% water is not good to the amino column, but it should be safe to use 80-20 ACN-water on it.