Two-photon fluorescent probes with large two-photon absorption (TPA) cross sections have shown wide applications in biomedical domain. However, both the species and amounts of high efficient probes are far from meeting the requirements, one main reason is that the relationship between the molecular structures and the responsive mechanisms are not clear and theoretical framework in this field is not perfect. In this work, the photophysical properties including one- and two-photon absorption and emission of three newly synthesized fluorescent probes for hydrogen polysulfide (H₂S_n) detection are investigated by density functional theory and time-dependent density functional theory with the polarizable continuum model in different solvents. Results indicate that the enhanced fluorescent intensity and enlarged TPA cross section can be found when the probes reacted with H₂S_n. Moreover, the OPA intensity is largest and its fluorescent intensity is largely enhanced when detecting H₂S_n for Pro2, this verifies its superior performance in the detection of H₂S_n than Pro1 and Pro 3. Furthermore, the inner mechanism for the increase of TPA cross section is revealed, the responsive mechanisms for photo induced electron transfer (PET) and fluorescence resonance energy transfer (FRET) processes are revealed through analyzing the energies and distributions of frontier orbitals. Our calculations provide theoretical perspectives for experimental measurements and could sever as a useful reference for developing advanced probes in biomedical fields.