Spin Traps and Radical Scavengers

The hydroxyl hydrogen in phenol is particularly susceptible to abstraction by a free radical. 

The process is exothermic, suggesting that the phenoxy radical is particularly stable. Display the spin density surface for phenoxy radical. Is the unpaired electron localized or delocalized over several centers Is the unpaired electron in the a or 7t system Draw appropriate Lewis structures that account for your data. 

Phenol is a radical scavenger . Other radical scavengers include 3,5-di-tert-butyl-4-hydroxytoluene (butylated hydroxytoluene or BHT) and vitamin E. 

Examine the spin density surface for BHT radical. Is the unpaired electron localized or delocalized Examine BHT radical as a space-filling model. What effect do the bulky tert-butyl groups have on the chemistry of the species (Hint BHT radical does not readily add to alkenes or abstract hydrogens from other molecules.) 

Compare the spin density surface for vitamin E radical to those of phenoxy and BHT radicals (see also Chapter 16, Problem 2). Are there significant differences among the three If so, elaborate. What is the function of the long alkyl chain in vitamin E Examine an electrostatic potential map for vitamin E radical. Do you expect it to be soluble in aqueous (polar) or non-aqueous (non-polar) environments, or both  

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The addition of small amounts of radical scavenger (such as benzoquinone and diphenylpicrylhydrazyl) led to the appearance of induction periods in the kinetic curves. The duration of the induction periods are proportional to the concentration of the radical scavenger. The presence of atmospheric oxygen slightly slowed the polymerization. These observations indicate that the polymerization proceeds by a radical mechanism. The radicals are formed from the y-radiolysis of the monomers. By comparison to the ESR spectrum of the radicals formed by thermal initiation with azobisisobutyroni-trile in the presence of a spin trap, the radical formed is... 

Not all radicals will add to spin traps, and in some cases addition may be too slow to compete with other processes. Also in biological systems, it must be borne in mind that the spin-traps will divert the reactive radicals from their normal role. In fact, they act as radical scavengers. Scavengers are often used as probes of radical reactions, so it is useful to use spin-traps for this purpose as well as for detection by ESR spectroscopy. 

Further evidence for Type I cleavage has been obtained by the use of diamagnetic radical scavengers as spin traps for radicals produced on photolysis of benzoin and benzoin methyl ether (18). In both < ses PhCO and PhCHOR (R = H, Me) radicals were trapped and characterised from the e.s.r. spectra of the stable nitroxide radicals formed e.g. 

The radical scavenging activity of garcinol have been confirmed using the electron spin resonance (ESR) spin trapping method which observe the reaction between superoxide anion and radical scavenger more directly. 

The lack of overt, acute, toxicity of DEPMPO is consistent with the large number of studies that have been carried out with DMPO, PBN and POBN (recent examples include refs. 65-71, reviewed in refs. 3,5,7,72 and 73). It should be noted, however, that the diethoxyphosphoryl group of DEPMPO appears to have biological effects e.g. the preservation of ATP levels in isolated rat hearts ) which are independent of the radical scavenging effects of the spin trap, and this must be borne in mind in the interpretation of data obtained from in vivo or ex vivo experiments. Such metabolic effects appear to be a common problem with spin traps (see also ref 74), and is partly attributable to the high concentrations of traps that need to be employed in such studies. 

Spin trapping methods were also used to show that when carotenoid-P-cyclodextrin 1 1 inclusion complex is formed (Polyakov et al. 2004), cyclodextrin does not prevent the reaction of carotenoids with Fe3+ ions but does reduce their scavenging rate toward OOH radicals. This implies that different sites of the carotenoid interact with free radicals and the Fe3+ ions. Presumably, the OOH radical attacks only the cyclohexene ring of the carotenoid. This indicates that the torus-shaped cyclodextrins, Scheme 9.6, protects the incorporated carotenoids from reactive oxygen species. Since cyclodextrins are widely used as carriers and stabilizers of dietary carotenoids, this demonstrates a mechanism for their safe delivery to the cell membrane before reaction with oxygen species occurs. 

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 has been widely used for superoxide detection in various in vitro systems [16] this method was applied for the study of microsomal reduction of nitro compounds [17], microsomal lipid peroxidation [18], xanthine-xanthine oxidase system [19], etc. As DMPO-OOH adduct quickly decomposes yielding DMPO-OH, the latter is frequently used for the measurement of superoxide formation. (Discrimination between spin trapping of superoxide and hydroxyl radicals by DMPO can be performed by the application of hydroxyl radical scavengers, see below.) For example, Mansbach et al. [20] showed that the incubation of cultured enterocytes with menadione or nitrazepam in the presence of DMPO resulted in the formation of DMPO OH signal, which supposedly originated from the reduction of DMPO OOH adduct by glutathione peroxidase. 

The last few years have seen numerous applications of spin trapping to biological systems, and in these the trapping of hydroxyl radicals has assumed some importance. This work has been confined almost exclusively to nitrone scavengers 4 the fact that the hydroxyl adduct [6] of DMPO is much more persistent than that [7] of the commonly used nitrone, benzylidene-t-butylamine-N-oxide ( phenyl t-butyl nitrone ,3 or PBN) [3], may be due to a fragmentation reaction, with subsequent oxidation of the cr-hydroxybenzyl radical, as shown. 

The pre-eminent advantage of C-nitroso-compounds as spin traps is that in the spin adduct the scavenged radical is directly attached to the nitroxide nitrogen. Consequently, the esr spectrum of the spin adduct is likely to reveal splittings from magnetic nuclei in the trapped radical, and these will greatly facilitate its identification. A simple example is presented in Fig. 2, which shows the spectrum of the spin adduct of the methyl radical with 2-methyl-2-nitroso-... 

The tri-t-butylnitrosobenzene, TNB, is monomeric even in the solid state, but the principal advantage of this scavenger, exemplified in the mechanistic studies described in Section 3 (p. 47), is that it functions as an ambident spin trap (Terabe and Konaka, 1973). Thus, primary alkyl radicals add to form nitroxides in the normal way, but with t-alkyl radicals, addition occurs at oxygen, alkoxyaminyl radicals (ArNOR) being formed. Secondary alkyl radicals give mixtures of both species (Fig. 5). The alkoxyaminyl radicals have a lower g-value than the nitroxides (ca. 2.004 vs. 2.006) and their spectra are therefore centred at slightly higher field positions than those of the nitroxides. 

A simple extension of the competition technique is to the comparison of scavenger efficiencies. Thus pairs of spin traps have been allowed to compete for a variety of radicals, including t-butoxyl, phenyl, and primary alkyl. Much more revealing, however, is the type of experiment in which the bimolecular trapping of a radical is allowed to compete with some other reaction of that radical whose absolute rate constant is known. In this way, the rate constant for the trapping reaction itself is accessible. 

Rate constants for spin trapping of alkyl radicals measured by the procedures outlined here, are collected with other spin-trapping rate data in Table 5. It will be seen that most nitrone and nitroso traps scavenge reactive radicals of diverse types with rate constants generally in the range 10s-10 1 mol-1 s l. Of the nitroso-compounds, the nitroso-aromatics (except for the very crowded TBN) are particularly reactive, whilst MBN and DMPO are the most reactive nitrones. Much of the data for spin trapping by nitrones has been accumulated by Janzen and his colleagues, who have discussed in a short review how steric and electronic factors influence these reactions (Janzen et ai, 1978). 

As data for the rates of spin-trapping reactions are accumulated, so it becomes possible to use the competition experiment in reverse , i.e. to determine rates of rearrangement, fragmentation, atom transfer, etc. which can compete with spin trapping. An attempt to estimate rates of decarbonylation of acyl radicals depended on this approach (Perkins and Roberts, 1973). Although the results obtained were intuitively reasonable, they depended on the assumption that the rate of scavenging of acyl radicals by MNP would be no different from that measured for the butoxycarbonyl radical. This still awaits experimental verification. Another application, reported recently, was to the rates of rearrangement (23) of a series of (o-(alkoxycarbonyl)-alkyl radicals... 

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