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

• Published in: Analytical Chemistry
• Main Authors: Audran, Gérard, Jacoutot, Samuel, Jugniot, Natacha, Marque, Sylvain, Mellet, Philippe
• Other Authors: Institut de Chimie Radicalaire (ICR), Aix Marseille Université (AMU)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Centre de résonance magnétique des systèmes biologiques (CRMSB), Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS), 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
• Subjects: [SDV]Life Sciences [q-bio]; Rugosa; antartic*
• Online Access: https://hal-cnrs.archives-ouvertes.fr/hal-02472334; https://doi.org/10.1021/acs.analchem.9b00561

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.