This study conducted using 40 healthy lactating mixed breed cows aged 1.8-4 years from the date of 1-10-2018 to 1-1-2019. These cows selected from Sohag Governorate;40 cows reared on small farms feed on balanced ration; 20 cows from them were at the first time of parturition, 10 cows were at the second time of parturition and 10 cows were at the third time of parturition. 10 cows used as a control animals brought from farmers houses and feed on unbalanced food. Blood and milk samples collected during three stages of lactation (first, second and third month of lactation). On each serum sample insulin, liver enzymes (SGOT, SGPT and ALP), total bilirubin, kidney function tests (urea, creatinine and uric acid), some minerals includes (Ca, phosphorus, Cl, Na, and K), some heavy metals include (Cu, Fe, lead, cadmium and Se), some antioxidants include (GSH, CAT and SOD), some hormones include (TSH in serum and milk leptin hormone) and glucose level were measured. A significant effect of the lactation stages were observed on SGOT, SGPT, urea, creatinine, Na, Ca, phosphorus, Cl, K, Cu, Fe, leptin, TSH, GSH, CAT, SOD, glucose, insulin, while there were no significant differences during the three lactation stages on serum ALP, total bilirubin, uric acid and Se.