River Hindon is an important river catering the demand of highly populated and industrial cluster of western Uttar Pradesh, India. Water quality of river Hindon is constantly deteriorating at an alarming rate due to various industrial, municipal and agricultural activities taking place along the course of the river. The present study investigates the seasonal and spatial variation in the water quality parameters at eight different sites for two consecutive years (2013-2015) along the selected stretch of the river. The physicochemical parameters such as pH, temperature, turbidity, total hardness (TH), total dissolved solids (TDS), total solids (TS), dissolved oxygen (DO), biological oxygen demand (BOD), chloride, nitrate, phosphate and fecal Coliform were analyzed for the river water quality. One way analysis of variance (ANOVA) was used to investigate the statistically considerable spatial and seasonal difference. Results of ANOVA suggest that there exist a statistically significant seasonal variation in the water quality of river with respect to pH, temperature, nitrate, sulphate, phosphate and DO. Whereas, the significant spatial variation was shown by TDS, TS, TH and chloride. Water quality index (WQI) was calculated for each site using the National sanitation foundation water quality index (NSFWQI) method. WQI results in the present study reveals that the water quality varies “bad” to “very bad” category at all sites in pre monsoon season. In post monsoon water quality comes under “bad” category at all sampling locations. The significant seasonal variation (p< 0.05) was recorded between the WQI of pre monsoon and post monsoon.