A reliable, selective and sensitive LC-MS/MS assay has been proposed for the determination of nitrofurantoin in human plasma. The analyte and nitrofurazone were extracted from 100 μL of human plasma via SPE on Strata-X 33 μm extraction cartridges. Chromatography was done on a BDS Hypersil C18 (100 mm × 4.6 mm, 5 μm) column under isocratic conditions. Quantitation was done using the multiple reaction monitoring (MRM) mode for deprotonated precursor to product ion transitions of nitrofurantoin (m/z 237.0 → 151.8) and nitrofurazone (m/z 197.0 → 123.9). The limit of detection and the lowest limit of quantitation of the method were 0.25 ng mL-1 and 5.00 ng mL-1, respectively, with a linear dynamic range of 5.00-1500 ng mL-1 for nitrofurantoin. The intra- -batch and inter-batch precision (RSD, %) was ≤ 5.8 %, while the mean extraction recovery was > 92 %. The method was successfully applied to a bioequivalence study of a 100 mg nitrofurantoin capsule formulation in 36 healthy subjects.