Bioequivalence and Bioavailability Forum

Dear Group Members,

Let's assume that we are performing the Bioequivalence study of the formulation of a highly protein bound drug which is 98% bound to plasma proteins. After direct Liquid-Liquid extraction or Solid Phase Extraction of the plasma samples followed by LC-MS/MS analysis, do we need to apply a correction factor for the protein binding? Eg: If a plasma concentration of a time point comes to 50ng/mL, do we need to calculate the bound form as 50*100/(100-98) = 2500ng/mL & use 50 (Free Form) + 2500 (Bound Form) = 2550 ng/mL, for Winnonlin data analysis?

In case we initially precipitate the plasma proteins by addition of Acetonitrile / Methanol before Liquid-Liquid or Solid Phase Extraction, do we need to consider the results as sum of the free & bound concentrations?

Can anyone provide any reference in this regard?

Thanks & regards,
Dr. Harish L. Rao

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Edit: capitals changed to mixed case in title [Ohlbe]