Chromatography Forum

What could the possible causes for an increase in USP tailing from 1.6 to 2.2 just by changing the mobile phases from 0.1% HCOOH in H2O (A) 0.1% HCOOH in ACN (B) to 0.1% H3PO4 in H2O (A) 0.1% H3PO4 in ACN (B)?

I can supply more information if needed, but really the question simplifies to this. Thanks in advance.

What could the possible causes for an increase in USP tailing from 1.6 to 2.2 just by changing the mobile phases from 0.1% HCOOH in H2O (A) 0.1% HCOOH in ACN (B) to 0.1% H3PO4 in H2O (A) 0.1% H3PO4 in ACN (B)?

I can supply more information if needed, but really the question simplifies to this. Thanks in advance.

Considering any liquid chromatography mode as a competition among all the mobile phase components with the analyte for the same stationary phase sites, then a change in peak shape is definitely possible. Depending on which mobile phase additive has weaker adsorption as compared to a given analyte on the stationary phase, one can expect tailed peaks for analytes which interact strongly with the stationary phase. Note that ACN or H2O will also compete, along with HCOOH or H3PO4, with the analyte on the stationary phase adsorption sites.

I'm sure that such an increase in tailing factor will be not accepted by the FDA. What was the reason to change the mobile phase?

The resolution between 2 sets of peaks in the original method were not baseline resolved. I changed the modifier to see if a change in pH would affect this and it did but at a cost of increased tailing of the main peak. The original method, <2.0 tailing method, will be used for assay and the modified method is for Area%.

The FDA question was the main reason for posting although at my last job, we had validated LC methods where the tailing for the main peak was >2.0 (related substances methods) that were approved by the FDA. I don't know why they would draw a distinction between the two method types.

If it helps to generate possible solutions, I'm using a Kinetex C18, 1.7 µm 100A, 50mm x 2.1mm column.