Chromatography Forum

Solved.

Welcome to the forum.

As a chemist, I think it could be confusing to think of this synthetic pesticide mixture as being 'natural' and 'in equilibrium'. The common formulation seems to be indeed 3:1, for whatever reason, but CMIT and MIT are 2 different molecules that do not convert in one another when they are mixed.

I personally wouldn't call your original standard material a standard material, and would highly prefer to develop the method starting from 2 separate, pure standards for CMIT and MIT before injecting mixtures.

What else is in the original mixture, and what solvent do you use to prepare a solution of that and the standard from Sigma?

First you need to make sure that whatever you see is your target compound which elutes from the column. You need to inject it without the column and measure peak area. Peak area without the column and sum of all peaks with the column should be the same if you are eluting your compound. Second, I would check proportional valve and equilibration time in your gradient method.