Belonging to the field of pharmaceutical compounds, the invention relates to 11, 20-dicarbonyl Jiyuan rubescensin a and L-amino acid-14-ester trifluoroacetate thereof, and a preparation method and usethereof. The preparation method includes: adopting Jiyuan rubescensin a as the raw material and performing oxidation with a Jones reagent, thus obtaining 11, 20-dicarbonyl Jiyuan rubescensin a, the structure of which is shown as the specification; then carrying out 14-position esterification reaction with N-BOC-L-amino acid to obtain N-BOC-L-amino acid-11, 20-dicarbonyl Jiyuan rubescensin a ester(III), then removing BOC protective group in trifluoroacetic acid, and then conducting salt forming to obtain L-amino acid-14-(11, 20-dicarbonyl Jiyuan rubescensin a)ester trifluoroacetate, which hasthe following general formula. The compound has good stability and anti-tumor activity, and provides the basis for screening antitumor drugs against esophageal cancer, gastric cancer, primary liver cancer, pancreatic cancer, cardiac carcinoma, colorectal cancer, bladder cancer, breast cancer, acute myelogenous leukemia and the like.