Objective Intravenous (IV) midazolam is the preferred cytochrome P450 (CYP) 3A probe for phenotyping, with systemic clearance (CL) estimating hepatic CYP3A activity. A limited sampling strategy was conducted to determine if partial area-under-the-concentration-time-curves (AUCs) could reliably estimate midazolam systemic CL during conditions of CYP3A baseline activity, inhibition and induction/activation. Methods Midazolam plasma concentrations during CYP3A baseline (n=93), inhibition (n=40), and induction/activation (n=33) were obtained from seven studies in healthy adults. Non-compartmental analysis determined observed CL (CLobs) and partial AUCs. Linear regression equations were derived from partial AUCs to estimate CL (CLpred) during CYP3A baseline, inhibition and induction/activation. Pre-established criterion for linear regression analysis was r2 ≥0.9. CLpred was compared to CLobs, and relative bias and precision were assessed using percent mean prediction error (%MPE) and percent mean absolute error (%MAE). Results During CYP3A baseline and inhibition, all evaluated partial AUCs failed to meet criterion of r2 ≥0.9 and/or %MAE <15%. During CYP3A induction/activation, equations derived from partial AUCs from 0 to 1 hour (AUC0–1), AUC0–2, and AUC0–4 were acceptable, with good precision and minimal bias. These equations provided the same conclusions regarding equivalency testing compared to intense sampling. Conclusions During CYP3A induction/activation, but not baseline or inhibition, midazolam partial AUC0–1, AUC0–2, and AUC0–4 reliably estimated systemic CL, and consequently hepatic CYP3A activity in healthy adults.