AbstractTwo four‐coordinate organoboron N,C‐chelate complexes with different functional terminals on the PEG chains are studied with respect to their photophysical properties within human MCF‐7 cells. Their excited‐state properties are characterized by time‐resolved pump‐probe spectroscopy and fluorescence lifetime microscopy. The excited‐state relaxation dynamics of the two complexes are similar when studied in DMSO. Aggregation of the complexes with the carboxylate terminal group is observed in water. When studying the light‐driven excited‐state dynamics of both complexes in cellulo, i. e., after being taken up into human MCF‐7 cells, both complexes show different features depending on the nature of the anchoring PEG chains. The lifetime of a characteristic intramolecular charge‐transfer state is significantly shorter when studied in cellulo (360±170 ps) as compared to in DMSO (∼960 ps) at 600 nm for the complexes with an amino group. However, the kinetics of the complexes with the carboxylate group are in line with those recorded in DMSO. On the other hand, the lifetimes of the fluorescent state are almost identical for both complexes in cellulo. These findings underline the importance to evaluate the excited‐state properties of fluorophores in a complex biological environment in order to fully account for intra‐ and intermolecular effects governing the light‐induced processes in functional dyes.