The invention discloses a preparation method of a chitosan-based derivative polymer point with a fluorescent tracing function. The preparation method is characterized by comprising the following steps of: taking N-alkyl-chitosan, anhydrous citric acid and N-2-hydroxy ethylenediamine, uniformly mixing and dispersing N-alkyl-chitosan, anhydrous citric acid and N-2-hydroxy ethylenediamine into a mixed solution of polyethylene glycol 400 and deionized water, performing reaction, dialyzing reaction products with water, and then drying, thereby obtaining chitosan-g-citric polymer points. The chitosan-based derivative polymer point has relatively good carbon dot fluorescence material biocompatibility, is low in toxicity, is good in water solubility, is free from caking and agglomeration in water, is applicable to fluorescence imaging and is suitable for serving as a drug carrier material.