The present invention pertains to the field of chemical medicine, particularly to 4-substituted coumarin derivatives and preparation methods and applications thereof. The invention provides 4-substituted coumarin derivatives with a structural formula as shown in Formula I. The invention also provides preparation methods and applications for the above 4-substituted coumarin derivatives. The compounds provided in the invention have strong anti-tumor activity with IC50 for plural tumor cell lines between 0.01-5 nM, and it also performs better to inhibit microtubule polymerization and has diversified biological activities and low toxicity, providing new options for drug-sensitive and drug-resistant tumor cells.