The objective of this investigation was to determine whether the oral clearance (CL) of chlorzoxazone could be estimated accurately from a single plasma chlorzoxazone measurement. In 6 healthy male subjects the mean clearance estimate (CLest) from a single 6-hour postdose measurement of plasma chlorzoxazone was within 6% of the actual mean CL calculated conventionally from the area under the plasma concentration versus time curve (AUC). After a second dose of chlorzoxazone administered 1 week later, the 6-hour mean CLest was within approximately 10% of the value from the previous week and within 15% of the value of CL from the previous week. No significant differences occurred between these three clearances, suggesting that for purposes of phenotyping CYP2E1 activity in humans with chlorzoxazone, a single-dose, single-sample procedure may suffice.