The invention provides Chinese chestnut polysaccharide pharmaceutical composition with an anti-cancer effect and a preparation method of the pharmaceutical composition and relates to the pharmaceutical field. The pharmaceutical composition is prepared from Chinese chestnut polysaccharide as well as pharmaceutically acceptable carriers or auxiliary materials. The preparation method of the pharmaceutical composition comprises the steps as follows: a Chinese chestnut material is subjected to extraction with a solvent extraction method, and a crude Chinese chestnut extract is obtained; the crude Chinese chestnut extract is subjected to deproteinization and decolorization treatment, dialysis is performed with a dialysis bag, and a Chinese chestnut dialysis liquid is obtained; the Chinese chestnut dialysis liquid is subjected to gel column chromatography, water is used as an eluent, an eluate containing polysaccharide components is collected, and Chinese chestnut polysaccharide is obtained;the Chinese chestnut polysaccharide is mixed with pharmaceutical adjuvant. The pharmaceutical composition takes the Chinese chestnut polysaccharide as the main active component, has significant cancercell inhibiting and anti-cancer effects and almost has no side effects.