Spin adducts

The spin adducts of free radicals and MNP or DMPO were observed by means of an ESR spectrometer. The data of hyperfine splitting constants were compiled in Tables 9 and 10 [40-42,44,45]. ESR studies on the initial free radicals revealed that the monoalkylamino radical RHN-, dialkylamino radical R2N-, and aminomethyl radical -CH2N< or aminoethylidene radical >N( CHCH3) were obtained from the corresponding primary, secondary, and cyclic tertiary amine. In case of a tertiary diamine such as TMEDA, formation of... 

Table 9 Hyperfine Splitting Constants of Spin Adduct Formed APS/Amine/MNP or DMPO System ...

Tabie 10 Hyperfine Splitting Constants of Spin Adducts Obtained from APS/Amine/MNP Systems... 

The initial radicals formed from the reaction of Ce(IV) ion and reductant systems can be trapped by MNP. The spin adducts of the initial radicals and MNP were observed by means of an ESR spectrometer. The structure of the initial radicals and the hypeiTme splitting constants of the spin adduct of the radical with MNP and tt-phenyl-N-/er/-butylnitrone (PBN) are compiled in Tables 5 and 6, respectively. 

The formation of spin adducts from radicals and MNP is as follows ... 

Figure 4.1 Time-course of free-radical production during aerobic (a) or anoxic (b) reperfusion of the isolated rat heart. Radical production was assessed using e.s.r. and quantified as the formation of a Af-tert-butyl-a-phenylnitrone (PBN) spin adduct. After a 35 min stabilization period of aerobic perfusion, hearts were made globally ischaemic for 15 min. Hearts were then reperfused, either with oxygenated buffer (a) (n = 6), or with anoxic buffer, switching to an oxygenated buffer after 10 min (b) (n = 5). The bars represent the standard errors of the means. Redrawn with permission from Garlick et af. (1987).

Poyer, J.L., McCay, P.B., Lai, E.K., Janzen, E G. and Davis, E.R. (1980). Confirmation of assignment of the trich-loromethyl radical spin adduct detected by spin trapping during carbon tetrachloride metabolism in vim and in vivo. Biochem. Biophys. Res. Commun. 94, 1154-1160. 

The presence of /3-hydrogen in the nitroxide radical may lead to disproportionation reactions. In spin-trapping experiments, N-t-butyl-a-phenyl nitrone yields rather unstable spin adducts. This type of radical can be stabilized by coordination to Nin. The Ni11 complex with N-oxy-A-r-butyl-(2-pyridyl)phenylmethanamine (923) reveals a distorted octahedral geometry with antiferromagnetic interactions between the unpaired electrons of the metal ion and the radical spins.00... 

FIGURE 9.1 EPR spectra of spin adducts recorded during the Fenton reaction in DMSO at different H202 concentrations ([FeClJ = 1 mM), (1), (2), and (3) are OH, OOH, and CH3 radicals, respectively. 

When the lifetime of the radicals is very short and direct ESR detection is not an option, spin trapping is used to detect radicals at ambient temperatures. The method is based on the scavenging of radicals, P, by a spin trap, leading to the formation of a spin adduct with higher stability in most cases, this adduct is a nitroxide radical. 

Spin trapping experiments have been performed recently in a fuel cell inserted in the ESR resonator ("in situ" cell), using DMPO and a-(4-pyridyl-l-oxide)-N-ferf-butylnitrone (POBN) as the spin traps [78,82,83], These experiments allowed the separate examination of spin adducts at the anode and cathode sides. 

HRP-catalyzed oxidation of tyrosine. ESR spectra of the spin-adducts of tyrosyl radical with 2-methyl-2-nitrosopropane were also obtained in the reactions catalyzed by MPO and LPO. 

On the other hand, microsomes may also directly oxidize or reduce various substrates. As already mentioned, microsomal oxidation of carbon tetrachloride results in the formation of trichloromethyl free radical and the initiation of lipid peroxidation. The effect of carbon tetrachloride on microsomes has been widely studied in connection with its cytotoxic activity in humans and animals. It has been shown that CCI4 is reduced by cytochrome P-450. For example, by the use of spin-trapping technique, Albani et al. [38] demonstrated the formation of the CCI3 radical in rat liver microsomal fractions and in vivo in rats. McCay et al. [39] found that carbon tetrachloride metabolism to CC13 by rat liver accompanied by the formation of lipid dienyl and lipid peroxydienyl radicals. The incubation of carbon tetrachloride with liver cells resulted in the formation of the C02 free radical (identified as the PBN-CO2 radical spin adduct) in addition to trichoromethyl radical [40]. It was found that glutathione rather than dioxygen is needed for the formation of this additional free radical. The formation of trichloromethyl radical caused the inactivation of hepatic microsomal calcium pump [41]. 

Another approach to this problem is a search for the other more effective spin traps. Frejaville et al. [23] demonstrated that the half-life of spin-adduct of superoxide with 5-(diethoxyphosphoryl)-5-mcthyl-l -pyrrolinc-/V-oxide (DEMPO) is about tenfold longer than that of DMPO OOH. Despite a much more efficiency of this spin trap, its hydrophilic properties limit its use for superoxide detection in lipid membranes. Stolze et al. [24] studied the efficiency of some lipophilic derivatives of DEMPO in the reaction with superoxide. These authors demonstrated a higher stability of superoxide spin-adducts with 5-(di- -propoxypho-sphoryl)-5-methyl-1 -pyrrolinc-A -oxidc (DPPMPO) and 5-(di- -butoxyphosphoryl)-5-methyl-... 

To study mechanisms C—E, it seems reasonable to employ both, electrochemical approaches and EPR-spectroscopy. It is important to be aware of the electrochemical properties of nitrones if used as spin traps for production of spin adducts (SA) is possible not only via homolytic process (C) but also via ionic processes shown in Scheme 2.77. In the case of (B), protonation can protect the... 

In contrast to this, it has been concluded that the formation of spin adducts on ultraviolet irradiation or mild oxidation of indole nitrones, in the presence of a N -heleroaromatic base, proceeds according to the Forrester-Hepburn mechanism (522). 

Under similar reaction, the action of XeF2 on PBN and DMPO led to similar spin adducts, proving the radical cation mechanism (524). Unlike this reaction, the formation of fluoro-containing spin adducts when using a weaker oxidant such as /V-lluorodibenzsulfonamide [(PhSC>2)2N-F] is believed to involve the Forrester-Hepburn mechanism (524). 

A study of the polymerization kinetics of methyl methacrylate, in the presence of PBN, and of molecular-mass properties of the obtained polymers shows that the systems react by the pseudoliving mechanism (699). In the first stages of the polymerization process, PBN reacts with oligomeric radicals, forming stable nitroxyl radical-spin adducts A-, see Scheme 2.207. 

Nitroxyl spin adducts (A ) react with the growing radical chain leading to the formation of labile end groups (Scheme 2.208). 

In order to identify organic free - radicals present at quantifiable concentrations during the sonication of PCBs, we employed Electron Spin Resonance (ESR) with a spin trap, N-t-butyl-a-phenyl-nitrone (PBN). PBN reacts with the reactive free - radicals to form more stable spin-adducts, which are then detected by ESR. The ESR spectrum of a PBN spin adduct exhibits hyperfine coupling of the unpaired election with the 14N and the (3-H nuclei which leads to a triplet of doublets. The combination of the spin-adduct peak position and peak interval uniquely identifies the structure of a free-radical. 

This article concerns a simple expedient whereby short-lived reactive free radicals may be transformed into more persistent paramagnetic species, thus enabling esr techniques to be applied to systems in which the concentration of the reactive radical remains below normal detection limits. The principle is a simple one. It depends upon the addition to the reaction system of a small quantity of a diamagnetic substance (the spin-trap ) having a particularly high affinity for reactive radicals the product of this trapping reaction must be a particularly persistant free radical (the spin adduct ) whose concentration will build to readily detectable levels (>ca. 10—7—10-6 M). The general reaction is represented by equation (1). 

From the foregoing discussion it is evident that, whilst some nitroxide spin adducts may be sufficiently persistent to be isolated, others decay more or less rapidly by one or more of a variety of pathways. This range of behaviour must be recognized as a considerable difficulty, especially in situations where it is not possible to make spectroscopic observations directly on the reacting sample but only when a period of time has elapsed after the experiment is completed this is frequently the case in radiolysis studies. 

Spin-adduct spectra often reveal splittings to substituent atoms other than hydrogen. Indeed, since the spin-trapping technique provides a convenient route to many nitroxides containing structural features likely to be of spectroscopic interest, it has frequently been used to this end. Chlorine and bromine splittings, as well as those from fluorine, have been encountered and many nitroxide spectra have been reported in which there is splitting from a second nitrogen, from phosphorus, or even from a metal atom. 

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