The invention relates to the technical field of biocatalytic synthesis, and specifically discloses an asymmetrically substituted 1,3,5-triazine compound and its preparation method and application. The asymmetrically substituted 1,3,5-triazine compound includes The following raw materials: isothiocyanate compounds, amidine compounds, and tetramethylguanidine. The asymmetrically substituted 1,3,5-triazine compound provided by the invention uses heme, horseradish peroxidase, hemoglobin or cytochrome C as a catalyst, is green, non-toxic, friendly to the environment, and has a high yield, solving the problem of Most of the existing asymmetrically substituted 1,3,5-triazine compounds use toxic catalysts in the preparation, which has the problems of being unfriendly to the environment and low yield; and the preparation method provided is superior to the traditional chemical synthesis method, and is highly efficient Green, short reaction time, mild reaction conditions, no refractory metal ion catalysis, and reduced catalyst consumption.