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In this report, we present a method to characterize the kinetics of electron transfer across the bilayer of a unilamellar liposome composed of 1,2-dimyristoyl- sn -glycero-3-phosphocholine. The method utilizes synthetic phospholipids containing noninvasive nitroxide spin labels having the &gt;N-O• moiety at well-defined distances from the outer surface of the liposome to serve as reporters for their local environment and, at the same time, permit measurement of the kinetics of electron transfer. We used 5-doxyl and 16-doxyl stearic acids. The paramagnetic &gt;N-O• moiety is photo-oxidized to the corresponding diamagnetic oxoammonium cation by a ruthenium electron acceptor formed in the solution. Electron transfer is monitored by three independent spectroscopic methods: by both steady-state and time-resolved electron paramagnetic resonance and by optical spectroscopy. These techniques allowed us to differentiate between the electron transfer rates of nitroxides located in the outer leaflet of the phospholipid bilayer and of those located in the inner leaflet. Measurement of electron transfer rates as a function of temperature revealed a low-activation barrier (ΔG ‡ ∼ 40 kJ/mol) that supports a tunneling mechanism.

langmuir

Systematic Quantification of Electron Transfer in a Bare Phospholipid Membrane Using Nitroxide-Labeled Stearic Acids: Distance Dependence, Kinetics, and Activation Parameters