Previous reports have described the importance of the substituents in the position 4 of nitrobenzodiazoles to fine-tune their optical properties. Therefore, we synthesized a subset of nitrobenzoselenadiazoles where the position 4 was functionalized with different nucleophiles and aromatic groups to assess whether those would influence phototoxicity. Benzoselenadiazoles including amines, phenols, thiophenols and aryl groups (compounds 6-9, Figure ) were synthesized by nucleophilic substitution under basic conditions or by conventional Pd-catalyzed Suzuki couplings (full synthetic and characterization details in Supplemental Information). Spectral characterization confirmed that amino-substituted analogues presented an effective push-pull dipole as indicated by their longer excitation wavelengths (Figure ). Amino-substituted nitrobenzoselenadiazoles showed the highest phototoxicity in cells compared to all other substituents and to the control compound 4F (Figure ). We also examined whether the incorporation of additional heavier atoms in nitrobenzoselenadiazoles -such as iodine, already reported for the successful conversion of other fluorescent structures into photosensitizers- would further improve their phototoxic potential. We employed the commercially available mono-iodinated diamine 10 to synthesize 9-diethylamino-6iodo-7-nitrobenzoselenadiazole (11a) as described above (Figure ; full synthetic details in Supplemental Information) and compared its photosensitive properties to the parent non-iodinated compound (4a). Both compounds 4a and 11a presented similar excitation wavelengths around 500 nm, with the non-iodinated derivative showing higher ratios of phototoxicity and being more suitable for the preparation of photosensitive metabolites. Finally, we examined the mechanism by which amino-substituted nitrobenzoselenadiazoles induce oxidative death and confirmed that, upon light irradiation, they predominantly produce singlet oxygen over radical anions (approximately in a 75:25 ratio, Figure ). Amino-substituted nitrobenzoselenadiazoles are therefore one of the smallest photosensitizer structures reported to date.