The invention discloses a preparation method and application of novel perylene drug molecules with bioactivity, and belongs to the technical field of medicine synthesis. The novel perylene drug molecules with bioactivity are characterized by having structural formula shown in the description, wherein R is O or S. The invention also relates to the preparation method of the novel perylene drug molecules with bioactivity. The novel perylene drug molecules with bioactivity prepared herein have good inhibitory activity against human cervical carcinoma Hela cells, human breast cancer MDA-MB-231 cells and human liver cancer HepG2 cells, have low toxicity to normal human cells, and are potentially applicable as antitumor drugs.