Chromatography Forum

When performing filter recovery, does anyone know why peak areas might continue to increase each time another mL is passed through a filter? Selectivity seems to be fine with these filters. They are actually centrifuge tubes with small pore size filters in them. There's no plateau even after 7 mLs, for both actives. % recovery starts out right at ~ 100% for the first aliquot, and just climbs. I've heard of this happening but I don't understand why. The sample excipients are cross-linking polymers; maybe they are part of the problem.

Thanks!

There will be a plateau. Just keep filtering more and more.
And there you'll have the answer as well - saturation.

% recovery starts out right at ~ 100% for the first aliquot, and just climbs

Do I get this right: The initial aliquot is 100 % recovery, the further ones are above 100 %? Then danko's theory does not work, saturation would be possible only if the first aliquot is below 100 %. What type of membrane is this, and what solvent do you have?
I have some other ideas:
1.) A little volume of solvent is absorbed by the membrane while your target molecules can pass (the membrane kind of swells). However, I would this effect VERY little, and also I'd expect this to happen at the beginning of the filtration process.
2.) The membrane holds back some response-weakening stuff - your analyte concentration is correct, but you have higher response due to the cleaner matrix. You can check if this is true by filtering blank matrix and add your target to the aliquot steps.
3.) You have found a new way to synthesize this product out of nothing. Get patent pending on the process and make lots of money out of it

Jörg

Recovery, probably starts out at 100% because the standard for calibration is filtered as well.

Best regards