Real-time tracking of ground- and excited-state responses of photoactive molecules to a change of microenvironment, from femtoseconds to milliseconds

Light triggers numerous fundamental biological processes in living organisms, by promoting (multi)chromophoric biomolecular systems to an excited state, which activates their function. The ability of these photoactive systems to absorb and use solar energy is often regulated by their microenvironment, which modulates their structural and energetic landscape and, thus, their functionality. Various stimuli-responsive photoactive materials are designed based on this principle. Understanding how photoactive molecular systems respond to light and environmental changes requires tracking changes at both the electronically excited- and ground-state levels, with ultrafast resolution across a broad range of timescales. Indeed, while light-induced excited-state dynamics involve processes as rapid as a few femtoseconds, the structural and electrostatic responses to microenvironmental changes may occur at timescales orders of magnitude slower, up to the millisecond and beyond. We demonstrate that this investigation can now be enabled by a novel and universal three-pulse ultrafast spectroscopy method that: (i) induces controlled, local changes in the microenvironment of a solution, specifically of pH, and (ii) probes the rapid spectroscopic response of photoactive molecules in both the ground and excited states, from femtoseconds to milliseconds. Controlling changes in the microenvironment and – simultaneously – bridging the ultrafast timescales of excited-state dynamics with the slower timescales of environmental and conformational changes is key to advancing our understanding of how living organisms and artificial materials regulate the interplay between light, environment, and structure in light-induced molecular processes.