Two-photon sensitive protecting groups for biological application

Thesis

Caged compounds are a class of photosensitive reagents used to stimulate cells with spatial control down to a sub-cellular level, and millisecond temporal control. They comprise of biologically important molecule which is modified with a photolabile protecting group. In the absence of light, caged compounds are physiologically silent but irradiation with light induces the release of biologically active species. Illumination under two-photon conditions is particularly advantageous as it enables restriction of the photolysis volume to ~1 fL and it provides deeper penetration into scattering samples.

This thesis reports the development of new protecting groups for two-photon uncaging in neuroscience. Mechanistically, the deprotection in these novel groups is designed to operate via an intramolecular photoinduced electron transfer (PeT) between the absorbing unit (electron-donor) and the release module (electron-acceptor). The modular design of these cages ensures separation of absorption and release steps, and allows each process to be tuned and optimized independently.

Chapter 1 provides an introduction to the two-photon absorption phenomenon and a historic overview of the uncaging technique. It also discusses recent advances in the development of two-photon sensitive probes used in neuroscience.

Chapter 2 describes the exploration of molecular designs for novel protecting groups. A two-photon absorbing dye (electron-donor; fluorene dye) and three different release units (electron-acceptors; nitrobenzyl, pyridinium and phenacyl) were identified as suitable building blocks for the current project. Efficiency of the intramolecular electron transfer between chosen units was evaluated using model dyads which constitute covalently linked electron-donor and acceptor species.

Chapter 3 is devoted to the synthesis and photophysical evaluation of nitrobenzyl-based protecting group.

Chapter 4 describes the preparation of pyridinium-derived protecting group and demonstrates PeT-mediated release of tryptophan and GABA under one- and two-photon excitation. Hydrolytic instability of pyridinium esters is highlighted.

Chapter 5 reports the synthesis, hydrolytic stability and one-photon uncaging efficiency of phenacyl-based derivatives.

Chapter 6 discusses properties of developed caged compounds and compares them with other compounds reported in literature. It contains overall conclusions and outlook for the current project.

Chapter 7 details the experimental procedures and the characterization of compounds synthesized during this work.

Authors: Korzycka, KA

• Publication date: 2015
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: photoinduced electron transfer; photolysis; uncaging; two-photon absorption; GABA
• Subjects: Organic chemistry; Neuroscience; Photochemistry and reaction dynamics; Organic synthesis; Chemistry & allied sciences; Synthetic organic chemistry