A simple, precise, accurate and stability-indicating reverse phase high performance liquid chromatography (RP-HPLC) method is developed for estimation of Isosorbide 5-Mononitrate in bulk drug and tablet dosage form. The method employed, with reverse phase phenomenex® Luna 5µ C18 (2) 100A (250 × 4.60 mm) column in an isocratic mode, with mobile phase of methanol: water: acetonitrile in the ratio 55:28:17 (%v/v/v). The flow rate was 1.0 ml/min and effluent was monitored at 217 nm. Retention time was found to be 4.391±0.015 min. The method was validated in terms of linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ) etc. in accordance with ICH guidelines. Linear regression analysis data for the calibration plot showed that there was good linear relationship between response and concentration in the range of 1- 9 µg/ml respectively. The LOD and LOQ values for were found to be 2.5 and 10 ng/ml respectively. No chromatographic interference from tablet excipients and degradants were found. The proposed method was successfully used for estimation of Isosorbide 5-Mononitrate in tablet dosage form.