The invention discloses a preparation method of a polysaccharide derivative and vanadium complex, which comprises the following steps of: firstly, reacting NaOH-alkalifying polysaccharide with chloroacetic acid for 3-4 hours at the temperature of 40-80 DEG C to synthesize the polysaccharide derivative; then reacting the polysaccharide derivative with a vanadium solution for 12-24 hours at the room temperature to obtain the vanadium complex of the polysaccharide derivative. The polysaccharide derivative and vanadium complex is applied to medicines for treating diabetes mellitus. In the invention, the polysaccharide derivative is adopted as raw material and reacts with VOSO4 in an aqueous medium under the neutral condition to obtain the polysaccharide derivative and vanadium complex through filtration, drying and crushing. In the invention, the complex preparation is completely carried out at the room temperature, has simple process and is easy to realize industrial production.