One-electron oxidation and reduction of glycosaminoglycan chloramides: A kinetic study

Abstract

Hypochlorous acid and its acid-base counterpart, hypochlorite ions, produced in inflammatory conditions, may produce chloramides of glycosaminoglycans, the latter being significant components of the extracellular matrix (ECM). This may occur through the binding of myeloperoxidase directly to the glycosaminoglycans.The N-Cl group in the chloramides is a potential selective target for both reducing and oxidising radicals, leading possibly to more efficient and damaging fragmentation of these biopolymers relative to the parent glycosaminoglycans. In this study,the fast reaction techniques of pulse radiolysis and nanosecond laser flash photolysis have been used to generate both oxidizing and reducing radicals to react with the chloramides of hyaluronan (HACl) and heparin (HepCl).The strong reducing formate radicals and hydrated electrons were found to react rapidly with both HACl and HepCl with rate constants of (1-1.7) x 10 8 M -1 s -1 and (0.7-1.2) x 10 8 M -1 s -1 for formate radicals and 2.2 x 10 9 M -1 s and 7.2 x 10 8 M -1 s -1 for hydrated electrons, respectively. The spectral characteristics of the products of these reactions were identical and were consistent with initial attack at the N-Cl groups, followed by elimination of chloride ions to produce nitrogencentred radicals, which re-arrange subsequently and rapidly to produce C-2 radicals on the glucosamine moiety supporting an earlier EPR study by Rees et 3 This is the first study therefore to conclusively demonstrate that reducing radicals react rapidly with glycosaminoglycan chloramides in a site-specific attack at the N-Cl groups, probably to produce a 100 % efficient biopolymer fragmentation process. Although less reactive, carbonate radicals, which may be produced in vivo via reactions of peroxynitrite with serum levels of carbon dioxide ,also appear to react in a highly site-specific manner at the N-Cl group. It is not yet known if such site-specific attacks by this important in vivo species lead to a more efficient fragmentation of the biopolymers than would be expected for attack by the stronger oxidising species, the hydroxyl radical. It is clear however that the N-Cl group formed in inflammatory conditions in the extracellular matrix does present a more likely target for both reactive oxygen species and for reducing species than the N-H groups in the parent glycosaminoglycans. Graphical abstract4

Efficiencies of fragmentation of glycosaminoglycan chloramides of the extracellular matrix by oxidizing and reducing radicals: potential site-specific targets in inflammation?

“…This was supported by the lack of any demonstrable reaction of these species with the parent molecule, HA, and also by the absorption spectra of transient species attributable to carbon-centred free radicals formed on the glucosamine moiety [41]. The latter observation provided support for the 1,2 hydrogen shift mechanism for HACl proposed by Davies and co-workers as shown in Scheme 2 in which a chloride ion is eliminated by one-electron reductants to form a nitrogen-centred radical in the first instance followed by rapid re-arrangement to produce the C-2 radical on the glucosamine moiety and/or C-4 radicals on the uronic acid moiety [22].…”

“…However, evidence from time-resolved studies appears to support the formation of C-2 radicals since the transient spectra observed are consistent with those expected for carbon-centred radicals associated with an adjacent nitrogen atom such as RCONRCH 2 . (where R can be H-or CH 3 -) [41]. It seems probable therefore that the 100% fragmentation yield observed for HACl occurs via the C-2 radical on the glucosamine moiety with the subsequent β-scission being 100% effective.…”

“…Although reaction (9) must produce N-centred radicals in the first instance, these must be short-lived as only carbon-centred radicals could be detected [22,41].…”

Free radical studies of components of the extracellular matrix: contributions to protection of biomolecules and biomaterials from sterilising doses of ionising radiation

“…Hence, for the more biologically relevant carbonate radical, a much higher degree of selectivity of attack with glycosaminoglycans is to be expected, as confirmed in fragmentation studies (Sibanda et al 2013a). The reactivities of both oxidizing and strongly reducing radicals with the chloramide derivatives of hyaluronan and heparin were investigated using both pulse radiolysis and laser flash photolysis techniques (Sibanda et al 2013b). The strong reducing agent, the formate radical was produced by both pulse radiolysis and by laser flash photolysis.…”

The kinetics and mechanism of the oxidation of pyruvate ion by hypochlorous acid

“…[33][34][35][36][37][38][39][40] Due to its outstanding oxidative power, this species can significantly alter the activities and availabilities of biomolecules.…”