6533b7d0fe1ef96bd125b4ea

RESEARCH PRODUCT

Synthesis of novel organic-based fluorophores – Implementation to the design of fluorogenic enzyme substrates based on the principle of in situ synthesis

subject

description

Detection and fluorescence imaging of biologic systems requires the implementation of efficient, robust and easy-to-use tools. Conventional fluorogenic probes currently used in microbiology lack efficiency since they are based on the single chemical modification of a fluorophore bearing an optically tunable reactive group, which often leads to incomplete fluorescence quenching. The main goal of my Ph.D thesis was to develop novel fluorogenic enzymatic substrates based on the "covalent assembly" principle. This approach also named "in situ synthesis" is based on the use of domino reactions to form a fluorescent moiety starting from a "caged" non-fluorescent molecule. In our case, the bioanalyte that triggers the reaction is present in bacteria, that we want to detect. This strategy provides many advantages (improved signal-to-noise ratio, easy exemplification to a wide range of analytes, …). In this context, the first goal of my thesis was to synthesize original fluorescent hetero-xanthene dyes to assess their stability under physiological conditions. Novel sulfone- and Si-pyronin derivatives were obtained. Synthesis of the corresponding "caged" precursors to the most stable compounds was then undertaken for their use as a peptidase-responsive probes. This work is described in the first and second chapter of this manuscript. Faced with difficulties to implement "covalent assembly" probe design principle to Si-pyronins, another class of fluorophores, namely quinoxalinone was explored. In situ formation of these fluorescent heterocycles triggered by an enzyme is presented in the third chapter. Finally, the last chapter was devoted to the synthesis of a conventional probe derived from a bis-sulfonyl-bis-aniline recently reported in the literature.