Lactoferrin is a kind of key basic proteins in bovine milk, which has attracted more and more research attentions. In the present study, lactoferrin was isolated from fresh bovine milk by defatting and casein-removing treatments, subsequently purified by SP Sepharose Big Bead ion exchange chromatography, Superdex 200 gel chromatography, and ultrafiltration The purified form of lactoferrin was obtained with the purity of 9471%. The denaturation temperature of lactoferrin was determined to be 731℃ by differential scanning calorimetry. Circular dichroism studies on the influence of heat treatment -63℃ 30min, 72~75℃ 20s, 85℃ 10min, 95℃ 10min- on the secondary structure of lactoferrin showed that,the α-helix conformation of lactoferrin was gradually transformed to β-sheet conformation with the increase of the treatment temperature, indicating that the protein molecules became better ordered and more compact with significantly increased trends of protein denaturation.