The study investigated a long-term effect of urbanization on the water quality of Ona River in Eleyele Catchment, Oyo State Southwest Nigeria. Secondary water quality data between 1979 and 2007 were collected from Eleyele’s Water Works, Ibadan. In compliment, water samples were collected from five sampling points along River Ona. Important water parameters analyzed using standard procedures were: temperature, pH, electrical conductivity (EC), total dissolved solids (TDS), turbidity, total alkalinity, total hardness, metals (calcium, magnesium, sodium, copper, lead, cadmium, chromium, nickel, iron) and anions (nitrate, sulphate, chloride). Results showed increased concentration of nitrate and chloride and decreased dissolved oxygen level of River Ona over 28-year assessments. Present field study showed relatively high values of Pb (0.06-1.15 mg/L), Cr (0.01-1.31 mg/L), Cd (0.00-0.26 mg/L), Fe (0.40-1.96 mg/L), Ni (0.02- 0.27 mg/L) and nitrate (27.0-50.0 mg/L) compared with World Health Organization (WHO) permissible standards in drinking water. Water quality parameters were generally high between sampling point 1 and 3 due to the influx of industrial effluent from the nearby industries and indiscriminate disposal of wastes at the bank of the river.