The invention provides a chitosan microsphere. The chitosan microsphere comprises, by mass, 25-99% of chitosan and 1-75% of a hydrophilic polymer, and the hydrophilic polymer comprises at least one of polyvinyl alcohol, gelatin, cellulose and polyoxyethylene. The chitosan microsphere improves the moisture absorption capability and the moisture retention capability of chitosan, and solves the problem of traditional powder hemostasis materials. The invention also provides a preparation method of the chitosan microsphere. The method comprises the following steps: mixing a chitosan solution with a hydrophilic polymer solution until the chitosan solution and the hydrophilic polymer solution are uniform in order to prepare a mixed solution; and adding the mixed solution to a negatively charged receiving solution in a dropwise manner through adopting an electrostatic spraying technology, standing to obtain a microsphere suspension, separating to obtain microspheres, washing the microspheres, and drying the washed microspheres to obtain the chitosan microsphere. The preparation method has the advantages of simplicity and mild conditions, and the dimensions of the prepared chitosan microsphere are uniform and controllable.