The simulated two-photon spectra of Nile Red shown in figure 3 reveals larger differences between the PE and PDE models. The spectra are generated from the five lowest singlet excited states. The gas-phase spectrum is composed of three intense transitions located at 1.5, 1.9, and 2.1 eV. Both embedding models predict that the lowest-energy transition is redshifted by 0.1 eV when including the protein and solvent. Moreover, the intensities of this lowest excited state are almost identical. When moving towards higher energies, the spectrum based on the PDE model retains the basic characteristics of the gas-phase spectrum, although (due to broadening) the two peaks from the gas-phase now appear as one peak with a shoulder. In contrast, the spectrum based on the PE model shows no clearly defined structure and instead shows a broad featureless band starting from 1.6-2.1 eV. A correct description of the two-photon transition probability requires an accurate description of not only the initial and final states involved in the transition, but in principle, also all other excited states. Thus, the artificial stabilization of higher-lying states and resulting overly large density of states at higher energies (see figure ), is the root cause of the broader spectra found when using PE to model the environment. Overall, we find that the two-photon absorption spectrum is clearly more sensitive to the environment description than the onephoton absorption spectrum and that the PDE model yields a more physically reasonable result.