Shifting-Nitroxides to Investigate Enzymatic Hydrolysis of Fatty Acids by Lipases Using Electron Paramagnetic Resonance in Turbid Media

International audience While optical methods are not efficient enough for the easy, fast, and efficient detection of enzymatic activity in turbid media, the properties of the electron paramagnetic resonance (EPR) technique make it suitable for use in such media. Nitroxides which exhibit a change in...

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Bibliographic Details
Published in:Analytical Chemistry
Main Authors: Audran, Gérard, Jacoutot, Samuel, Jugniot, Natacha, Marque, Sylvain, Mellet, Philippe
Other Authors: Institut de Chimie Radicalaire (ICR), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Centre de résonance magnétique des systèmes biologiques (CRMSB), Centre National de la Recherche Scientifique (CNRS)-Université de Bordeaux (UB), Institut Lavoisier de Versailles (ILV), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)
Format: Article in Journal/Newspaper
Language:English
Published: HAL CCSD 2019
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Online Access:https://hal-cnrs.archives-ouvertes.fr/hal-02472334
https://doi.org/10.1021/acs.analchem.9b00561
Description
Summary:International audience While optical methods are not efficient enough for the easy, fast, and efficient detection of enzymatic activity in turbid media, the properties of the electron paramagnetic resonance (EPR) technique make it suitable for use in such media. Nitroxides which exhibit a change in their EPR hyperfine coupling constants upon enzymatic activity and are selective to lipases were developed under the name of shifting-nitroxides. Several fatty acids, exhibiting saturated and unsaturated chains of various lengths, were coupled with the shifting-nitroxide via an enol ester link and tested against several lipases. As the solubility of fatty acids is low in HEPES buffer, experiments were performed in turbid aqueous solution. Almost all labeled fatty acids were hydrolyzed by Candida rugosa lipase, and more selectivity is observed with Porcine Pancreas lipase type II. No activity was observed for lipase AK Amano 20, Candida antartica lipase B, and Mucor miehei lipase.