Bioavailability (BA) and bioequivalence (BE) studies

Published

March 18, 2026

Formulations change across development:

  1. Solution/capsule-in-envelope (Phase 1)
  2. Tablet/capsule (Phase 2)
  3. Commercial formulation (Phase 3/launch)
  4. Post-approval changes.

Each switch may require a bridging study, with increasing confidence required as development progresses. Clinical studies are the primary tool for managing formulation changes, but other methods exist (e.g. in vitro dissolution-based biowaivers).

Absolute bioavailability (Fabs)

Compares oral (or other extravascular) administration to an IV reference. Gives the true fraction of dose reaching systemic circulation. Should be determined for new active substances intended for systemic action. Important for interpreting mass balance data (distinguishing incomplete absorption from first-pass metabolism), evaluating elimination route contributions, and supporting BCS-based biowaivers. When no IV formulation is available, an IV microdose can sometimes be used.

Relative bioavailability (Frel)

Compares two extravascular formulations. Used in early development (Phase 1–2b) when formulations change.

BA vs. BE: the key distinction

Both BE and relative BA studies share the same objective, design, and statistical analysis (comparing AUC and Cmax via the geometric mean ratio (GMR) and its 90% CI). The key distinction:

  • Relative BA: GMR and 90% CI reported without pre-defined boundaries (effect estimation).
  • Bioequivalence: GMR and 90% CI must fall within pre-defined boundaries (generally 0.80–1.25).

BE demonstrates that two formulations deliver the drug at the same rate and extent of absorption:

  • Rate of absorption is estimated by Cmax (and tmax).
  • Extent of absorption is estimated by AUC.

When to conduct BE

  • New formulation vs. marketed product (generics, line extensions, fixed-dose combinations).
  • Post-approval changes to a marketed product (new excipient composition/supplier, new equipment, new manufacturing site).
  • Comparators reformulated for blinding in pivotal clinical studies.
  • Marketed formulation vs. formulation used in pivotal clinical studies (“BE-like”).

BE study design

  • 90% CI of the GMR (test/reference) for AUC and Cmax must fall within 0.80–1.25.

The acceptance range appears asymmetric on the original scale but is symmetric on the log scale: ln(0.80) ≈ −0.223 and ln(1.25) ≈ +0.223. PK parameters are log-normally distributed, so statistical analysis is performed on log-transformed data. The ±20% threshold is based on the assumption that differences in systemic exposure smaller than 20% are not clinically significant for most drugs.

  • Randomized, two-way crossover, single dose. The number of subjects with evaluable data should not be less than 12 for a crossover design or less than 12 per treatment group for a parallel design (ICH M13A). Actual sample size based on a power calculation.
  • Replicate design for highly variable drugs (intra-subject CV >30%).
  • BCS class 1 and class 3 drugs may qualify for biowaivers (in vitro dissolution-based BE). See BCS (Biopharmaceutics Classification System) for the solubility/permeability classification that underpins biowaivers.
  • A biowaiver for lower strengths may be applied if formulations are proportionally similar (immediate-release and multiple-unit modified-release formulations only).
  • Sampling: ≥12 blood samples per subject; <20% of AUC extrapolated; adequate density around tmax. Analyte: parent drug (even for prodrugs). PK analysis: non-compartmental analysis (NCA) of AUC and Cmax.

Food effect studies

Characterizes the effect of food on the PK of an oral drug to inform dosing recommendations (e.g. “take with or without food”, dose adjustments). Structurally a type of relative BA study – same formulation under two conditions (fed vs. fasted) with no predefined acceptance criteria.

  • Randomized, single-dose, two-period crossover (fed vs. fasted), in healthy participants.
  • Fed condition: high-fat (~50% of calories), high-calorie (~800–1000 kcal) standardized meal; dose 30 minutes after start of meal.
  • Fasted condition: overnight fast ≥10 hours; water allowed until 1 hour before dosing.
  • The result informs labeling (e.g. no food restriction, take with food, avoid with high-fat meals).
  • Can be included as a cohort within the FIH study or run as a standalone study.
  • Minimum 12 evaluable subjects (FDA).

Fasted vs. fed BE studies (ICH M13A)

  • A fasting study provides the greatest discrimination between formulations and is sufficient for most immediate-release (IR) products.
  • Both fasted and fed studies are required for:
    • High-risk IR products (solid dispersions, lipid-based, nanotech, or other specialized formulations).
    • Modified-release (MR) formulations (e.g. extended-release, delayed-release).
  • A fed-only study is acceptable when the product is labelled “take with food only” (e.g. for tolerability reasons).
  • Fed condition: high-calorie (~800–1000 kcal), high-fat (~50% of calories) standardized meal; dose 30 minutes after start of meal.