The method presented describes a sensitive and specific quantitative assay for the determination of flumazenil in human plasma. Flumazenil and an internal standard, midazolam, are isolated by a basic extraction. The final extract is separated on a 25-m BP-1 capillary column and drugs are detected by selected ion monitoring at m/z 229 and m/z 310 for flumazenil and the internal standard, respectively. The minimum detectable quantity is 1.0 ng/mL, for flumazenil in plasma. Coefficients of variation for within-run data were less than 6%.