Chromatography Forum

Hello, I was wondering if anyone else here has been struggling with a similar issue as us. Our Buchi C-810 flash units have been experiencing an issue where our pressure in our 5000g C-18 columns has been steadily increasing. We have attempted flushing with both pure acetonitrile and pure acetone at a rate of 15ml per minute both of which only showed a very miniscule decrease in pressure.
Thank you so much for your assistance and time!

(1) Are you making sure that 100% of your sample is 100% fully soluble in the mobile phase (The mobile phase should be used for injection)?
(2) Are you properly filtering your samples through a 0.45 or 0.22 um membrane BEFORE injecting?

You will have to divulge more about what exactly you're doing to get truly helpful answers (search "rule of Rod").

I looked - it appears that this column is packed with C-18 based media in 50µ spheres designed for use with peptides. It further appears that this is intended for process, not analytical chromatography.

Given that, I'm not sure your approach to cleaning it out is the best... Another concern is whether your peptide sample solutions, even if small and filtered, may tend to be sticky, which would account for the clogging effect. You may want to investigate whether sharp change in your aqueous buffer's pH loosens things up more than organic solvents. Addition of a zwitterion may also help.

In addition, reverse the direction of flow (insert the column backwards) to physically blow clogging material off of the frit and head of column.

Regards,
JMB