The invention discloses an alkannin naphthazarine mother nucleus acetoxylation derivative as well as a preparation method and application thereof, belonging to the technical field of medical chemistry. The structural formula of the alkannin naphthazarine mother nucleus acetoxylation derivative is shown in the specifications, wherein R is alkane, olefin, cyclane, cycloolefine or arene which contains 1-7 carbon atoms, or is alkane, cyclane, cycloalkane or arene which contains atoms; R1=R2=COCH3; and R3=R4=H or R1=R2=R3=R4=COCH3. Compared with alkannin, the invention has the advantages that: an in-vitro tumor cell suppression test indicates that the cytotoxicity is lowered, an animal in-vivo antitumor test indicates that the derivative has a certain tumor growth suppressing action, and the derivative can be used as a pre-medicament applied to the treatment of malignant tumors or the treatment of malignant tumors. Meanwhile, the preparation method related to the invention has the advantages of readily-available raw materials, short synthetic route and mild condition.