The invention relates to a triple optical anti-counterfeiting ink with fluorescence, delayed fluorescence and room temperature phosphorescence, as well as an anti-counterfeiting method and an application thereof, which relate to the optical anti-counterfeiting ink. The anti-counterfeiting ink is composed of 1-5 parts of a nitrogen-enriched fluorescent carbon dot material, 720 to 880 of urea, 280 to 320 of biuret, and 25000 to 30000 of an organic solvent. The anti-counterfeiting method comprises: providing a triple optical anti-counterfeiting system having fluorescence, delayed fluorescence androom temperature phosphorescence; obtaining a fluorescent anti-counterfeiting mark under an excitation light source of 250-460 nm, the confidential information stored by the triple anti-counterfeiting ink and the interference information stored by the fluorescent anti-counterfeiting ink; after the excitation light source of 250-320 nm is turned off, obtaining the delayed fluorescent anti-counterfeiting mark, and displaying the confidential information; and after the excitation light source of 330-460 nm is turned off, obtaining the room temperature phosphorescent anti-counterfeiting mark, wherein the color thereof is obviously different from that of the delayed fluorescence. The anti-counterfeiting ink and anti-counterfeiting system are used in ink-jet printers, anti-counterfeiting marks,and information hiding.