The mechanisms of how dyes and catalysts for solar-driven transformations such as water oxidation to form O₂ work have been intensively investigated, however little is known about how their independent photophysical and chemical processes work together. The level of coordination between the dye and the catalyst in time determines the overall water oxidation system's efficiency. In this computational stochastic kinetics study, we have examined coordination and timing for a Ru-based dye-catalyst diad, [P2Ru(4-mebpy-4'-bimpy)Ru(tpy)(OH₂)]⁴⁺, where P2 is 4,4'-bisphosphonato-2,2'-bipyridine, 4-mebpy-4'-bimpy is 4-(methylbipyridin-4'-yl)-N-benzimid-N'-pyridine, a bridging ligand, and tpy is (2,2':6',2''-terpyridine), taking advantage of the extensive data available for both dye and catalyst, and direct studies of the diads bound to a semiconductor surface. The simulation results for both ensembles of diads and single diads show that progress through the generally accepted water oxidation catalytic cycle is not controlled by the relatively low flux of solar irradiation or by charge or excitation losses, rather is gated by buildup of intermediates whose chemical reactions are not accelerated by photoexcitations. The stochastics of these thermal reactions govern the level of coordination between the dye and the catalyst. This suggests that catalytic efficiency can be improved in these multiphoton catalytic cycles by providing a means for photostimulation of all intermediates so that the catalytic rate is governed by charge injection under solar illumination alone.

Serial femtosecond crystallography has opened up many new opportunities in structural biology. In recent years, several approaches employing light-inducible systems have emerged to enable time-resolved experiments that reveal protein dynamics at high atomic and temporal resolutions. However, very few enzymes are light-dependent, whereas macromolecules requiring ligand diffusion into an active site are ubiquitous. In this work we present a drop-on-drop sample delivery system that enables the study of enzyme-catalyzed reactions in microcrystal slurries. The system delivers ligand solutions in bursts of multiple picoliter-sized drops on top of a larger crystal-containing drop inducing turbulent mixing and transports the mixture to the X-ray interaction region with temporal resolution. We demonstrate mixing using fluorescent dyes, numerical simulations and time-resolved serial femtosecond crystallography, which show rapid ligand diffusion through microdroplets. The drop-on-drop method has the potential to be widely applicable to serial crystallography studies, particularly of enzyme reactions with small molecule substrates.