The application discloses a MOF-based high-order multi-photon excitation fluorescent material and a preparation method and application thereof, relates to the technical field of multi-photon excitation fluorescent materials, and the preparation method comprises the following steps: 1) preparation of ZrTc; 2) preparation of Au NRs; 3) preparation of Au NR / ZrTc; and 4) preparation of Au NR / ZrTc@FA. The MOF-based high-order multi-photon excitation fluorescent material prepared by the application has good biocompatibility and low dark toxicity, can realize enhanced two / three-photon fluorescence imaging under near-infrared two-region laserirradiation, and can be applied to cancercell specific targeting and deep tissuefluorescence imaging.