I'm developing an experiment to extract some analytes for an aqueous sample and analyze them through a UPLC. My extraction solvent that I'm using is a mix of ethyl acetate/glacial acetic acid which does indeed extract my analytes as I've seen in my chromatogram. To further flesh out the method, I wanted to figure out the percent recovery by performing spikes of the various analyte standards into the sample - then extracting the sample to figure out my percent recovery.

Now, my problem is that my analyte standards are only partially soluble (about 20%) in the ethyl acetate/glacial acetic acid mixture I'm using which is terrible. My analyte standards, however, are soluble in other solvents e.g. DMF, Acetone. So my approach was to dissolve the standards in DMF for example, and spike it into my sample; then extract with ethyl acetate/glacial acetic acid. I realize by doing this that I am spiking it into a different matrix then the extraction solvent.

This leads to my questions. First off, how can I account for the fact that my standard is in a different solvent then the extraction solvent? Do I need to apply or calculate some correction factor for this? Second, is there a better way of approaching this situation where the analyte standard and analyte in the sample don't seem to be soluble in the same solvent?

In case you're wondering then how my analytes get extracted - the analytes generally tend to learn towards a more unpolar nature. Therefore they really hate water, so adding in ethyl acetate alone will cause the analytes to move into the ethyl acetate layer.