Bioequivalence and Bioavailability Forum

Hi giang nidqc,

❝ Could it be a possibility that the absence of one APIs may affect the final concentration of other APIs (regarding sample preparation and peak response)?

Sample preparation

Impossible. Regardless whether you use LLE or SPE each analyte follows the law of mass action (driven by pH, lipophilicity) independent from other analytes. The amount of the “target phase” (organic solvent, RP- or exchange resin) is 6 to 12 orders of magnitude higher than the one of the analyte. Hence, the presence / absence of other analyte(s) in a similar – low – amount is simply irrelevant. It can never change the equilibrium constant.

Chromatography

Unless you have an insufficient separation, no. Only if peaks overlap, in MS ion-suppression/enhancement by one analyte could influence the response of the other.

Ligand-binding assays

Can be problematic if you have a substantial degree of cross-reactivity.

Dear giang nidqc

❝ Could it be a possibility that the absence of one APIs may affect the final concentration of other APIs (regarding sample preparation and peak response)?

I agree with Helmut's post above. There is just one thing I'm thinking of: how was the spiking of calibration and QC samples done when you used 5 analytes ? Did you use a single spiking solution containing all analytes (usual practice), or separate working solutions ? If you used a single, combined solution at each level concentration, no problem. If you used separate spiking solutions, you will have a different proportion of solvent added to the plasma if you have less analytes, which may influence your recovery and matrix effects.