I analyse SSRIs (pKa values of them between 9-10) in human plasma by HPLC following liquide-liquide phase extraction. The residue was resuspended with mobile phase (0.25 ml), water: ACN (42:58) (pH:3.5, adjusted by TFA, isocratic), vortex 15 sec., ultrasonic bed 2 min. then vortex 15 sec. The recoveries were lower than 40%. Same tubes were resuspended with 0.25 ml DMSO and re-analysed again and the additional recoveries were more than %40. So, the mobile phase does not solve all residue of analytes from the bottom of the glass tube. I think mobile phase is not always an ideal solvent for resuspension!
Is there any relation between pKa values of molecules analysed and the pH of solvent used for resuspension of residue after extraction?
Thank you
Its compound should be soluble in mobile phase. That is an inconvenience. The extraction solvent could be another problem of low recovery. Another possibility would be to try to increase the percentage of ACN in the resuspension
Have you thought about Kow oil:water partition coefficient or tried variant son mobile phase with more hydrophobic vs. aqueous fractions
Mobile phase is always recommendable to resuspend your residue...even when you inject system suitability sample diluted with other solvent than mobile phase, the system sometimes is not suitable even..sample made in diluent (mobile phase) rectifies it..
In agricultural chemistry, we often face much more difficult matrices than plasma samples. The matrix we encounter frequently prevents complete dissolution of the residues of interest. To circumvent this issue, we typically dissolve samples with vortex mixing in a small amount of organic solvent (50-100 µL of MeOH or ACN) followed by aqueous mobile phase to a final volume of 1 to 2 mL. This usually allows for analysis of compounds with extremely different polarities in the presence of extracted plant and animal tissues. By keeping the percent organic at or below 10%, there is usually minimal peak shifting (any peak shifting can be readily accounted for by dissolving standards in the same solvent). We frequently use this technique to dissolve samples even when the analyses require large injection volumes (900 µL).
As I understand your method, you are trying to dissolve your preparation in 0.25 mL of the eluent. Perhaps you are exceeding the solubility of your preparation in eluent. If you were to increase the amount of solvent and then increase the injection volume you might reach a happy medium.
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