title: Understanding the radical mechanism of lipoxygenases using 31P NMR spin trapping.
abstract: In this paper, we use our quantitative (31)P NMR spin trapping methods, already developed for simple oxygen- and carbon-centered radicals, to understand the radical intermediates generated by enzymatic systems and more specifically lipoxygenases. Our methodology rests on the fact that free radicals react with the nitroxide phosphorus compound, 5-diisopropoxy-phosphoryl-5-methyl-1-pyrroline-N-oxide (DIPPMPO), to form stable radical adducts, which are suitably detected and accurately quantified using (31)P NMR in the presence of a phosphorus containing internal standard. This system was thus applied to better understand the mechanism of enzymatic oxidation of linoleic acid by soybean lipoxygenases-1 (LOX). The total amount of radicals trapped by DIPPMPO was detected by (31)P NMR at different experimental conditions. In particular the effect of dioxygen concentration on the amount of radicals being trapped was studied. At low dioxygen concentration, a huge increase of radicals trapped was observed with respect to the amount of radicals being trapped at normal dioxygen concentrations.
authors.affiliation: Dipartimento di Scienze dell'Ambiente e del Territorio, Università degli Studi di Milano-Bicocca, Piazza della Scienza 1, 20126 Milano, Italy.
authors.forename: Luca
authors.initials: L
authors.lastname: Zoia
authors.forename: Raffaella
authors.initials: R
authors.lastname: Perazzini
authors.forename: Claudia
authors.initials: C
authors.lastname: Crestini
authors.forename: Dimitris S
authors.initials: DS
authors.lastname: Argyropoulos
mesh_terms: D003497:Cyclic N-Oxides / Q000737:chemistry; D005609:Free Radicals / Q000737:chemistry*; D019787:Linoleic Acid / Q000378:metabolism; D008084:Lipoxygenase / Q000737:chemistry / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D063065:Organophosphonates / Q000737:chemistry; D010759:Phosphorus Isotopes / Q000737:chemistry; D013025:Glycine max / Q000201:enzymology; D019097:Spin Trapping
chemical_list: C512236:5-diisopropoxyphosphoryl-5-methyl-1-pyrroline-N-oxide; D003497:Cyclic N-Oxides; D005609:Free Radicals; D063065:Organophosphonates; D010759:Phosphorus Isotopes; D019787:Linoleic Acid; D008084:Lipoxygenase