Abstract: Milk as raw material was centrifuged to remove the lipids and then precipitated the proteins by appending with different concentration of calcium chloride (Ca Cl2) in different heating conditions. After being eliminated the lactose by cooling crystallization, cyclic adenosine phosphate (c AMP) extracted from milk was detected by high performance liquid chromatography ( HPLC) . Finally, the optimization condition of c AMP extraction technology was determined. As the results showed, comparing with the c AMP concentration dealed with different Ca Cl2 concentrations there was no significant difference between the 0.8mg/m L Ca Cl2 samples and 2.0mg/m L (p>0.05) samples. However, the c AMP concentration of 0.8mg/m L Ca Cl2 samples was higher than that of the 1.2mg/m L and 1.6mg/m L Ca Cl2samples (p<0.05) . While considering the heating effect on c AMP concentration, the 80℃ group showed the higher production than the other three groups (p<0.05) . In combination with the two factors referred before, the optimal technique process was the 1.6mg/m L Ca Cl2with80℃, so the c AMP concentration could reach as much as (66.86±3.21) μg/m L. Therefore, we could get the optimum condition of the extraction and isolation of c AMP from milk:1.6mg/m L Ca Cl2, 80℃. This process was great significance for milk comprehensive utilization as well as increasing the added values of milk.