The present study was planned to investigate the effect of oxytocin administration to Sahiwal cow, local breed of Pakistan on milk composition with respect to lactation stages (mature milk, peak production and end production). Sixteen cows were divided into two groups. One group was subjected to intramuscular injection of oxytocin (20IU) and other group kept as control. Milk samples were collected from both groups at different lactation stages and evaluated for various parameters. The pH, fat, protein, total solids and ash increased, lactose and acidity decreased, whereas solid not fat were not affected by the lactation stages. Oxytocin administration to cows resulted in decreased of milk fat, lactose, protein, total solids, solids not fat and increased in ash content. Phosphorous and sodium concentration increased while copper, zinc and potassium decreased with lactation stages. Magnesium, calcium and chloride content first decreased at peak production stage then increased at the late lactation stage. Administration of oxytocin also resulted in increased concentration of sodium, chloride and decreased in potassium content at all lactation stages whereas increase in copper content in oxytocin treated milk at end production was observed. The concentration of fatty acids (from C4:0 to C14:0) increased upto peak production and then decreased at the end of lactation stages. The palmatic acid (C16:0) increased while stearic acid (C18:0) and oleic acid (C18:1) decreased with the progress of lactation stages. There was no effect of oxytocin on the C4:0, C8:0, C10:0, C14:0 and C18:0 fatty acids while C6:0, C12:0, C16:0 and C18:1 fatty acid decreased on oxytocin administration. Electrophoretic study indicated maximum intensity of casein in milk at mid lactation stage, whereas whey content was found to be the highest at end production stage of milking. Casein (as1, as2 and β-CN) fractions and whey protein fractions (Ig, BSA, β-Lg and α-La) showed lighter bands in oxytocin injected milk as compared to control. Lactoperoxidase and acid phosphatase decreased while lipase activity increased with the progress of lactation. Alkaline phosphatase activity first increased then decreased at the end of lactation. Thiocyanate content also increased with lactation. Acid phosphatase increased and alkaline phosphatase and lipase decreased while there was no effect on the lactoperoxidase activity when the oxytocin was injected for a longer period. Thiocyanate content increased with the administration of oxytocin.