Antioxidants in Science, Technology, Medicine and Nutrition

ANTIOXIDANTS in science, technology, medicine and nutrition

Gerald Scott, DSc (Oxon)

Professor Emeritus in Chemistry, Aston University, Birmingham

Table of Contents

Cover image

Title page

GERALD SCOTT, DSc (Oxon)

Copyright

Dedication

PREFACE

Chapter 1: Peroxidation in Chemistry and Chemical Technology

1.1 Peroxidation

1.2 Effect of Substrate Structure on Peroxidation Rate

1.3 Initiation of Peroxidation

1.4 Termination

1.5 Products of Peroxidation

1.6 The Technological Effects of Peroxidation

1.7 Measurement of Polymer Peroxidation

Chapter 2: The Biological Effects of Peroxidation

2.1 Causes of Peroxidation in Biological Substrates

2.2 Products of Lipid Peroxidation

2.3 Rancidification of Fats and Oils

2.4 Pathological Effects of Peroxidation

2.5 Atherosclerosis

2.6 Cancer

2.7 Inflammation

2.8 Iron Overload

2.9 Environmental Damage

2.10 Ageing

Chapter 3: Chain-breaking Antioxidants

3.1 What are Antioxidants?

3.2 The Chain-breaking Donor Mechanism

3.3 The Chain-breaking Acceptor Mechanism

3.4 The Catalytic Chain-breaking Mechanism

3.5 Applications of the Catalytic CB Process in Polymers

Chapter 4: Preventive Antioxidants, Synergism and Technological Performance

4.1 Peroxidolytic Mechanisms

4.2 Metal Deactivators

4.3 UV Absorbers and Screens

4.4 Synergism and Antagonism

4.5 Physical Aspects of Antioxidant Performance

Chapter 5: Antioxidants in Biology

5.1 Antioxidant Mechanisms in vivo

5.2 Naturally Occurring Chain-breaking Donor (CB-D) Antioxidants

5.3 Naturally Occurring Preventive Antioxidants and Synergists

5.4 Nutritional Aspects of Antioxidants

5.5 The Antioxidant Potential of Drugs

Chapter 6: Antioxidants in Disease and Oxidative Stress

6.1 Epidemiological Studies

6.2 Epidemiological Studies of Cardiovascular Disease

6.3 Antioxidant Intervention and Supplementation

6.4 The Effects of Antioxidants on CVD in Animals

6.5 Epidemiological Studies of Cancer

6.6 Effects of Antioxidants on Cancer in vivo

6.7 Antioxidants and Ageing

6.8 Parkinson’s Disease

6.9 Alzheimer’s Disease

6.10 Antioxidants in Inflammation

6.11 Antioxidants in Environmental Damage

6.12 Therapeutic Potential of Antioxidants

INDEX

GERALD SCOTT, DSc (Oxon)

Gerald Scott read Chemistry at Balliol College, Oxford, and after taking a first class honours degree in 1952 carried out research with Dr W.A. Waters on free radical reactions of biologically important polyhydroxyphenols. He joined ICI (Dyestuffs Division) where he was appointed Manager of Polymer Auxiliaries Research in 1961. In 1965 he published his first book Atmospheric Oxidation and Antioxidants covering both technological and biological materials.

In 1967, Scott left industry to take the Chair of Chemistry at the recently chartered University of Aston in Birmingham, where he developed an internationally renowned research group concerned with the oxidative deterioration and stabilisation of organic materials. His publications number some 300 scientific papers, mainly in the field of antioxidant mechanisms in recognition of which Oxford University awarded him a DSc in 1983. In 1985 he published (with Professor Grassie of Glasgow University) Polymer Degradation and Stabilisation for post-graduate students and researchers in the polymer industries. Professor Scott also edited and contributed to a series of eight books titled Developments in Polymer Stabilisation from 1979-1987, and in 1990 a book entitled Mechanisms of Polymer Degradation and Stabilisation. In 1993 Scott edited a new edition of his earlier book Atmospheric Oxidation and Antioxidants in three volumes, covering both chemical technology and biology, and in 1995 he edited (with the late Dan Gilead) Degradable Polymers: Principles and Applications.

Some 40 international patents resulted from Professor Scott’s research, the first group concerned with antioxidants in the time-controlled degradation of plastics which are being used widely in many countries, particularly in the control of plastics litter, in agricultural applications. His second group of patents are concerned with the chemical attachment of antioxidants to polymers and are finding application in polymers subjected to aggressive conditions (high temperatures, solvent extraction, etc.). His third group of patents is concerned with a new class of antioxidants derived from spintraps (including nitric oxide) which have application in both the deterioration of organic substrates and in biological peroxidation.

Professor Scott’s research has increasingly turned in recent years to his early interest in mechanisms of biological antioxidants. He has shown that α-tocopherol and its oxidative transformation products are among the most powerful antioxidants yet examined in polymers at sub-ambient oxygen concentrations and he believes that the antioxidant mechanism of the quinonoid products is relevant to the behaviour of α-tocopheroquinone in vivo.

Professor Scott is a Fellow of the Royal Society of Chemistry, Fellow of the Institute of Materials, and a member of the New York Academy of Sciences. In 1988 he was elected honorary life member of the Japanese Materials Life Society and in 1992 he was elected to membership of the International Academy of Creative Endeavours.

Copyright

First published in 1997 by

ALBION PUBLISHING LIMITED

International Publishers

Coll House, Westergate, Chichester, West Sussex, PO20 6QL England

COPYRIGHT NOTICE

All Rights Reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, electronic, mechanical, photocopying, recording, or otherwise, without the permission of Albion Publishing, International Publishers, Coll House, Westergate, Chichester, West Sussex, England

© Gerald Scott, 1997

British Library Cataloguing in Publication Data

A catalogue record of this book is available from the British Library

ISBN 1-898563-31-4

Printed in Great Britain by Hartnolls, Bodmin, Cornwall

Dedication

Dedicated to the memory of

WILLIAM ALEXANDER WATERS

who fired me with his own enthusiasm for free radical research

PREFACE

The history of antioxidants goes back to the 19th century when it was realised that the deterioration of natural rubber was not caused by biological processes, as had previously been assumed, but by peroxidation. However, the biological terms ageing, perishing, poisoning, fatigue, etc., continue to be used by polymer technologists to the present day to describe specific aspects of rubber deterioration. It was discovered empirically that certain chemicals used in vulcanisation were able, in very low concentration, to improve the durability of rubber products. Subsequent research into antioxidants or antioxygens falls into two distinct although overlapping phases. The first, concerned with the protection of technological materials from oxidative deterioration, began in the latter part of the 19th century and reached a peak in the 1960s. The second phase began in the 1950s with the recognition of the importance of biological antioxidants in some diseases. Food chemists had earlier shown that peroxidation was primarily responsible for the rancidification of polyunsaturated oils and fats and that this was inhibited by naturally occurring vitamin antioxidants. In parallel studies, oil chemists showed that the drying of paint films, catalysed by transition metal ions was due to the oxidative cross-linking of polyunsaturated oils to macromolecules. To the chemist it is evident that the same free radical chemistry must be potentially capable of peroxidising lipids in vivo to both carbonyl breakdown products and to oligomers. However, due to the effectiveness of the endogenous biological antioxidants, the analogies between the deterioration of edible oils and the pathological effects of lipid peroxidation (e.g. in atherosclerosis) were only slowly recognised and it took the emergence of large scale epidemiological studies into antioxidant vitamin deficiency to provide the impetus for the present major upsurge of interest in free radical peroxidation in vivo.

Antioxidant research in the life sciences has not yet reached its zenith since some of the more important conclusions arising from classical chemical studies of antioxidant mechanisms have not yet crossed the disciplinary barrier. To many biochemists and materials technologists antioxidant is still synonymous with radical trap and although this is part of the truth, it is not the whole truth. In 1965, in a review of peroxidation and antioxidants, I drew attention to the importance of hydroperoxides as ubiquitous initiators of peroxidation and reviewed the available evidence on synthetic antioxidants with catalase-like activity (Atmospheric Oxidation and Antioxidants, 1965). I described the peroxidolytic antioxidants as preventive because, by preventing the formation of free radicals from hydroperoxides, they provide a complementary mechanism to the kinetic chain-breaking antioxidants. Other preventive antioxidant mechanisms identified were metal ion deactivation, UV absorption and deactivation of reactive oxygen species. Preventive mechanisms have nothing to do with radical trapping and yet they are frequently as important and sometimes more important in peroxidising systems than radical traps. The use of synergistic combinations of antioxidants acting by different mechanisms became the basis of polymer stabilisation technology in the 1960s but the concepts developed at that time have still to impact fully on antioxidant research in the life sciences.

The emphasis in this book is therefore on mechanisms. A primary intention is to demonstrate the relevance of antioxidant mechanisms to current studies of antioxidants in both technology and biology. In principle it is also possible to develop from the present body of knowledge predictive theories to point the way more clearly to potential therapeutic applications of antioxidants in vivo. I make no excuse then for returning in this book to the chemical foundation of antioxidant theories which began with the very thorough science-based studies of peroxidation by Bolland, Bateman, Gee and others in the mid-decades of this century. There is no doubt that the antioxidant classification that resulted from this early work is germane to the understanding of how antioxidants function in biology and medicine. My hope is that the principles outlined and the more speculative extensions of them proposed will serve to stimulate a vigorous and fruitful debate between classical oxidation chemists and researchers in the life sciences.

I am grateful to my many collaborators past and present who have contributed to the understand of how antioxidants act and I am particularly indebted to the following who have provided me with detailed information about their own recent work: Professor Paul Addis, Dr Bruce Ames, Dr Fiorelle Biasi, Dr Norman Billingham, Dr Rod Bilton, Dr Walter Bottje, Dr Joan Braganza, Dr Richard Cottrell, Professor Evgueni Denisov, Dr Gary Duthie, Professor Fred Gey, Dr Michael Golden, Dr Edward Hall, Professor Barry Halliwell, Professor Philip James, Professor Robert Hider, Dr Frank Kelly, Dr Tim Key, Dr C.J.N. Lacey, Dr Jan Malik, Dr Michael Marmot, Dr M.J. Mitchinson, Dr Simon Maxwell, Mr J.B. Park, Dr Hilary Powers, Dr Jan Pospísil, Professor Russel Reiter, Dr Roland Stocker, Professor Alexander Tkác, Professor Walter Willett, Dr Paul Winyard, Dr R.J. Woodward and Dr Hans Zweifel. I am also grateful to the following organisations for details of their products: Dairy Crest Ltd (Colleen Amos), Kraft Jacobs Suchard Ltd (Simon Kane), Mattews Foods plc (G. Burrows), MD Foods Ltd (Søren Madsen), Safeway Stores plc (Anna Sinclair), St. Ivel Ltd (Sarah Waterfield), Van den Bergh Foods (Sarah Nolan).

I am grateful to Ellis Horwood, MBE and Rosmary Harris for the excellent collaboration in the production of this work.

Finally, I am deeply indebted to my wife, Gwen, for her help and patience.

Gerald Scott

1997

1

Peroxidation in Chemistry and Chemical Technology

1.1 Peroxidation

The reactions of dioxygen with organic materials are arguably among the most important of all chemical reactions. The oxidation of carbon-based nutrients is the basis of life energy and the combustion of hydrocarbon minerals is the primary source of domestic and industrial energy. Directed oxidation of primary oil-based hydrocarbons is currently the method of choice for the manufacture of intermediates in the chemical industry; a large number of the primary intermediates for the polymer and fine organic chemicals industries are based upon products derived from hydrocarbons by oxidation. Very often, as in the manufacture of phenol from isopropyl benzene, oxidation occurs (see Scheme 1.1) by the classical radical chain reaction first proposed by Bolland, Bateman and co-workers [1,2] and which is common to all peroxidation reactions [3,4]. In the case of cumene and other highly peroxidisable hydrocarbons, the mild oxidation conditions allow the intermediate hydroperoxide to be isolated.

Scheme 1.1 Peroxidation of iso-propyl benzene (Cumene)

In general, the more reactive is the substrate to autoxidation, the higher is the yield of hydroperoxide. However, when the desired product is not the hydroperoxide itself but one of its decomposition products, the oxidation is carried out under conditions where the intermediate hydroperoxide undergoes thermolysis to give a stable end-product. Thus, in the oxidation of cyclohexane to adipic acid or of ρ-xylene to ρ-toluic acid, the intermediate hydroperoxides are not isolated (see Scheme 1.2).

Scheme 1.2 Oxidation of ρ-xylene to ρ-toluic acid

A common feature of many industrial autoxidation processes is the use of transition metal ions, frequently cobalt carboxylates, which in small amount catalyse the radical breakdown of the intermediate hydroperoxide by reactions (1) and (2), thus reducing the activation energy of this process and increasing the rate of the overall reaction:

(1)

(2)

Catalysis by low concentrations of transition metal ions is a common feature of all oxidation processes and is particularly important in biological systems due to the ubiquitous presence of iron in both complexed and ionic form.

Many peroxidation processes are autoinhibiting; that is the rates of oxygen absorption and hydroperoxide formation become slower as the reaction proceeds. In the case of iso-propyl benzene, it was discovered empirically that the addition of a small amount of a base markedly increased the yield of hydroperoxide [5]. It was suggested by Roberston and Waters [6] that the autoinhibition of alkyl aromatic oxidation was due to the formation of a small amount of phenol by the acid catalysed process shown in Scheme 1.1. Phenol is a weak antioxidant which functions by reducing the intermediate alkylperoxyl:

(3)

and is thus a retarder of the radical chain reaction which leads to the accumulation of hydroperoxide. The phenoxyl radical is relatively stable and readily undergoes dimerisation to dimers and trimers (reaction 3(b)) which are themselves weak chain-breaking antioxidants [7]. However, it is unlikely that reaction 3(a) is primarily responsible for autoinhibition, because phenols without ortho tertiary alkyl groups are not efficient antioxidants since they undergo chain transfer with oxidisable substrates like cumene (reaction 3(c)). It is much more likely that acid catalysed decomposition of cumene hydroperoxide is the main cause of inhibition in Scheme 1.1. This is a general antioxidant mechanism which will be discussed in more detail in Chapter 4, but it should be noted that, following the pioneering work of Oberright et al on the mechanisms of sulphur antioxidants in petroleum hydrocarbons [8], measurement of the ratio between the ionic decomposition products of cumene hydroperoxide (phenol + acetone) and the homolytic decomposition products (a-cumyl alcohol + a-methylstyrene + acetophenone) is frequently used as a diagnostic technique to identify and quantify the significance of peroxidolytic antioxidants in peroxide initiated peroxidations [9–16]. Autoinhibition is very general in peroxidation since low molecular weight carboxylic acids are widely formed by breakdown of hydroperoxides. Most important of these are formic acid (see Scheme 1.1) which has pKa = 3.8 and malonic acid (pKa = 1.9). Even benzoic acid (pKa = 4.2) and acetic acid (pKa = 4.8) are sufficiently acidic to provide a mild antioxidant effect, although benzoic acid and to a lesser extent other carboxylic acids can also induce radical decomposition of hydroperoxides at higher concentrations [17]:

(4)

In industrial practice, alkali is added continuously to autooxidising cumene in order to maintain the pH at 7.0 [18] and thus inhibit the acid catalysed decomposition of the hydroperoxide.

Many industrial chemicals undergo peroxidation under ambient conditions, particularly in the presence of light. The explosion danger of peroxidised diethyl ether during distillation is well known and warnings about their removal before distillation is an essential part of the safety advice given to chemistry students. Similar problems are encountered with other dialkyl ethers. For example, tetrahydrofuran is widely used in polymer characterisation (e.g. in size exclusion chromatography) and unless hydroperoxides are removed before using this solvent, their thermal degradation products may confuse the outcome of the analysis.

1.2 Effect of Substrate Structure on Peroxidation Rate

For most carbon-based substrates at ambient oxygen pressures, the reaction of an alkyl radical with dioxygen, reaction (5), occurs with zero activation energy. Consequently, the rate-controlling step in the radical chain oxidation process is the rate of reaction of alkylperoxyl with the substrate. Two alternative reactions have to be considered.

(5)

(6a)

(6b)

The first, reaction 6(a), is the rate of hydrogen abstraction of the most labile hydrogen atom in the substrate and the second, reaction 6(b) is the addition of alkylperoxyl to a reactive double bond. Both reactions may occur together if the activation energies are similar. For example, allyl benzene and indene copolymerise with ground state oxygen to give a low molar mass copolymer of oxygen and olefin [19]. This is illustrated for indene in Scheme 1.3 [20]. where it can be seen that hydroperoxide formation constitutes a chain-transfer process. 1,2-substituted double bonds do not normally participate in reaction 6(b) but olefins conjugated, either with carbonyl groups, with other olefinic double bonds or with aromatic rings may copolymerise with oxygen [19].

Scheme 1.3 Copolymerisation of indene with oxygen [ 20]

The driving force for both 6(a) and 6(b) is the stability of the radical produced. Many polyunsaturated allylic and benzylic compounds form delocalised radicals and autoxidise slowly at ambient temperature even in the absence of light. However, polar factors are also involved in the transition state since alkylperoxyl is electrophilic and it was elegantly shown by Russell many years ago that the relative oxidisabilities of the para substituted alkyl benzenes obey the Hammett relationship [21]. This is shown for substituted toluenes in Table 1.1. In general, substrates containing alkyl are activated to oxidation, whereas those containing halogens, nitryl, nitro and carboxyl groups are deactivated [21].

Table 1.1

Relative reactivities of substituted toluenes toward ROO· at 90°C (per hydrogen atom) [21]

*Assumes constant rate of termination

The effect of electron delocalisation in the transition state is much more important than the effect of polarity. Alkanes and saturated fatty acids (e.g. stearic, C18:0 and palmitic, C16:0), unlike their polyunsaturated analogues (e.g. C18:2) do not oxidise at a significant rate at ambient temperatures and pressures. In general it is necessary to go to high temperatures (>150°C) with the use of transition metal ion catalysts to obtain appreciable oxidation rates in the case of saturated hydrocarbons. The introduction of just one double bond or an aromatic ring a- to methylene changes the position and it then become possible to isolate the intermediate hydroperoxides (see iso-propyl benzene above). The relative rates of oxidation of oleic, C18:1,(n-9), I, linoleic, C18:2(n-6), II, linolenic C18:3,(n-6), III and arachidonic, C20:4,(n-6), IV acids is 1:10:20:40. (Note that fatty acids are described by the number of carbon atoms:number of double bonds and, in parenthesis, the position of the nearest double bond to the terminal methyl.)

(I)

(II)

(III)

(IV)

It is evident that the effect of two activating double bonds adjacent to methylene in II is significantly greater than the addition of a further 1,4-diene unit in III. The electron is delocalised over 5 carbon atoms in both cases and the activating effect of additional 1,4-diene groups is essentially statistical.

Some branched-chain substrates peroxidise intramolecularly by hydrogen abstraction by peroxyl from a neighbouring reactive methylene group. This is particularly true of the a-olefin polymers and copolymers. Polypropylene has been shown to form a sequence of hydroperoxide groups at vicinal tertiary-alkyl carbon atoms [22–24]. These are formed by the mechanism shown in Scheme 1.4. Propagation by hydrogen abstraction from a tertiary carbon atom is about six times faster than from a secondary carbon [25] and this is particularly facilitated in the transition state in Scheme 1.4 by intramolecular hydrogen bonding [23]. Furthermore, the vicinal hydroperoxides are less stable than isolated hydroperoxides due to internal hydrogen bonding which induces homolysis of the O-O bond, thus increasing radical formation. Finally, the rate of termination of tertiary peroxyl radicals is up to three orders of magnitude slower than the rate of termination of secondary peroxyl radicals (see Section 1.4). This combination of factors results in a much more rapid peroxidation of polypropylene than of polyethylene and the peroxidation kinetic chain length is approximately 100 for PP but only 10 for PE [26,27]. An important consequence of the greater oxidative sensitivity of PP than PE is that equilibrium hydroperoxide concentrations in the former are much higher than in the latter so that it is possible to follow the concentrations of hydroperoxide in PP during photooxidation whereas in PE concentrations are too low to be measured by standard spectroscopic methods [22].

Scheme 1.4 Formation and decomposition of vicinal hydroperoxides in polypropylene

In the light of the above, it is not unexpected that blends of PE and PP decrease in photostability as the proportion of PP in the blend increases [28] and PP can be looked upon as a prooxidant for PE [22]. However, this is partly an effect of phase separation between the two components since a rubbery terpolymer of ethene, propene and a diene monomer improves both the compatibility of PE/PP blends and their resistance to photodegradation in spite of the fact that the terpolymer actually oxidises more rapidly [29]. Similar sensitising effects by polyunsaturated polymers have been observed in the impact-modified polymers which are generally polyunsaturated rubbery polymer in polystyrene [30–32] and PVC [33]. It is clear, then, that morphological effects can be as important as chemical peroxidisability in determining polymer durability and this will be discussed further in Section 1.6.4.

1.3 Initiation of Peroxidation

1.3.1 Autoinitiation by ground-state dioxygen

Substrates which contain reactive methylene groups are very readily hydroperoxidised by the chemistry discussed in the last section and, particularly in the presence of transition metal ions, it is experimentally difficult to exclude reactions (1) and (2) from the initiation process. Moreover, ground-state triplet oxygen is a relatively unreactive molecule, and direct involvement in initiation reactions is normally difficult to prove due to the widespread occurrence of hydroperoxides which are notoriously unstable to light and heat. However, styrene has no reactive methylene groups to form hydroperoxides and the normal hydroperoxide initiated rate equation: r ∝ (O2abs)½ which reflects that the rate of peroxidation is not followed in styrene peroxidation. Instead it is constant (r = (1.4·10⁵)½). The most probable explanation is that this represents the rate of direct attack of ground state oxygen at the vinyl group [34] and a similar explanation has been proposed for the self-initiation of dihydrophenanthrene peroxidation [35].

1.3.2 Peroxidation induced by ionising radiation

High energy radiation can break carbon-carbon and carbon-hydrogen bonds in organic substates [36] and hydrogen-oxygen bonds in water [37] (Chapter 2) and the resulting free radicals are important initiators for peroxidation. This process has assumed considerable practical importance in polymers as a means of cross-linking through carbon but more recently, due to the increasing application of electron beams in the sterilisation of polymer artifacts for medical applications, it has been found to have an important influence on the initiation of peroxidation in the hydrocarbon polymers. The following chemical sequence is believed to be involved over a very short time-scale (given in parenthesis) in the formation of macroradicals;

(7)

(8)

(9)

The subsequent reactions of macroradicals with oxygen depend on the availability of oxygen to the carbon radical centres, and the rate of peroxidation is markedly affected by oxygen diffusion rates and the presence or absence of antioxidants in the polymer. A practical consequence of this is that there is normally a pronounced lag time between irradiation and the onset of mechanical deterioration. Thus, in the case of polypropylene syringes sterilised by electron beam irradiation, there may be no discernable change in mechanical properties for many weeks before catastrophic disintegration occurs.

Competition between chain scission and cross-linking is very much dependent upon the structure of the polymer and the availability of oxygen [38], but the chemistry of peroxidation products follows very much the same pattern as that described in Section 1.2 for thermal oxidation of polymers. Consequently, initiation by high energy irradiation is a convenient technique for the study of the peroxidation chain reaction and termination processes at ambient temperatures [38]. However, in semi-crystalline polymers, some peroxyl radicals are long-lived due to incarceration in the crystalline phase and these play very little part in the subsequent propagation process [39,40].

1.3.3 Initiation by reactive oxygen species

In contrast to ionising radiation, light reaching the earth’s surface (i.e. of longer wavelength than 295 nm), does not normally break C-C or C-H bonds in the absence of a suitable chromophore to form chemically reactive photoexcited states. However, singlet oxygen, (¹ΔgO2 or ¹O2) is formed readily under photoxidative conditions by energy tranfer to ground state oxygen from photoexcited states of sensitisers (Sens) such as rose bengal or methylene blue and some inorganic pigments. This topic has been comprehensively reviewed by Rabek [41] and the reader is directed to this book for an encyclopaedic overview of the voluminous literature on the effects of light in chemistry. The generation of singlet oxygen is described by reactions (10) and (11):

(10)

(11)

Singlet oxygen is very reactive toward most olefins. Unlike ground state oxygen, it is not itself a diradical but it reacts instead like a dienophile. Thus in the case of a 1,2-dialkyl olefin, it adds to the double bond with simultaneous hydrogen abstraction in an ene reaction [42–45]:

(12)

The hydroperoxides formed in reaction 12 are effective intititors for peroxidation.

1,3-Dienes readily form cyclic peroxides (endoperoxides) [46] with almost zero activation energy and the very fast reaction of ¹O2 with diphenyliso-benzifuran (DBPF) without physical quenching [46,47] provides a useful quantitative method of measuring ¹O2 concentrations in solution;

(13)

High yields of hydroperoxides can be produced when singlet oxygen, made by microwave discharge or by reaction of hydrogen peroxide with hypochlorous acid [48], is reacted with olefins; but yields are much lower when produced by photosensitisation of ground state oxygen due to the photolysis of the hydroperoxide itself [49].

Singlet oxygen is relatively unimportant as an intitiator of photooxidation in saturated hydrocarbons [50], since it is physically quenched by more abundant molecules in the environment (e.g. water). However it may play a significant role in the peroxidation of unsaturated compounds, particularly in biological systems [46,51].

Ozone (O3) also reacts rapidly with olefins [45,52] but very slowly with tetrahedral carbon. However, it has been shown [53] that aryl activated methylene groups do react slowly with ozone to give hydroperoxides. Ozonolysis is of considerable importance in tyre technology, where ozone cracking occurs at very low ozone concentrations. The chemistry of the ozonolysis of olefins is ionic rather than radical chain and has been very thoroughly investigated by Criegee [54] and by Bailey [55]. Its main significance to free radical peroxidation is the formation of dialkyl peroxides and particularly hydroperoxides which may under appropriate conditions act as initiators for free radical peroxidation. This chemistry is summarised in Scheme 1.5.

Scheme 1.5 Reactions of ozone with olefins

The technological effects of ozone have been extensively reviewed [3,52,56,57] and it should be noted that peroxides are primary products of ozonolysis of many unsaturated compounds in vivo as well as in vitro and it is likely that these species intensify asthmatic diseases in heavily industrialised atmospheres where damaging concentrations of ozone are frequently reported. This will be discussed in Chapter 2.

Superoxide (O2·-) is readily formed by a variety of reducing agents/sensitisers in aqueous media, reaction 14, but it is not a powerful oxidising agent due to resonance stabilisation [52,58], and it frequently acts as a reducing agent for iron in biological systems, for example in the Fenton reaction, reaction 15(b):

(14)

O2·- has very low free radical reactivity [58] but, when protonated, it gives the much more reactive hydroperoxyl radical, HOO·. This radical is more organo-soluble than superoxide, and like the latter it is readily reduced to hydrogen peroxide. In the presence of the reduced transition metal ions (notably iron), hydrogen peroxide is reduced to the highly reactive hydroxyl radical in the Fenton reaction:

(15)

Superoxide and hydrogen peroxide have not been found to play a very important role as initiators of peroxidation in technological media, almost certainly because the necessary hydrophilic conditions do not normally exist for the reduction of ground state oxygen to occur. However O2·- is extremely important as a source of hydrogen peroxide and hydroxyl radicals in biological systems and the consequence of hydrogen peroxide reduction in vivo will be discussed in in Chapter 2. Alkyl hydroperoxides on the other hand are the most common source of radicals formed in an analogous reduction process in hydrophobic systems by reactions (1) and (2) and this is of great significance in technological media, notably hydrocarbon oils [59], rubber [60] and plastics [60], where organo-soluble transition metal salts are frequently contaminants from their working environment. However some transition metal ions (notably Cu²+) may undergo inversion from prooxidants to effective antioxidants as concentrations are increased (Chapter 3). Macroalkyl hydroperoxides are also very sensitive to UV light and are the primary source of radicals during the photooxidation of commercial polymers (Section 1.6.3).

1.3.4 Initiation by physical stress

Mechanical shear can break carbon-carbon bonds in solids [61] or in highly viscous liquids [62] leading to macroalkyl formation by carbon-carbon bond scission. Mechanooxidation is particularly important in polymer technology during processing (Section 1.6.2(c)). Stress is also an important initiator in rubbers subjected to mechanical deformation (Section 1.6.2(d)) and the technologically important phenomena of fatigue and ozone cracking [63,64] are associated with this stress-induced peroxidation. Recent evidence also suggests that stress may accelerate the rate of photooxidation of polyolefins [65].

It also seems likely by analogy with polymers that increased radical formation in stressed muscles may in part be a mechanochemical process although little research has been carried out to examine this possibility. However, Symons et al. have shown that peroxyl radicals are formed during bone fracture [66,67] and they attribute this to the breakage of collagen strands embedded in the bone. It seems likely that analogous physical processes may occur in severely fatigued muscles. Mechanooxidation seems to be a potentially important area for future peroxidation studies in vivo.

1.4 Termination

In most peroxidations, reactions 6(a) and 6(b) are rate determining in the radical chain process. Consequently, at ambient pressures, the main termination process is reaction (16). However, at low oxygen concentrations, and particularly in the case of hydrocarbons that give relatively stable alkenyl radicals termination through cabon-centred radicals, reactions (17) and (18) may play a significant role even at ambient oxygen pressures [1,68]:

(16)

(17)

(18)

Tertiary alkylperoxyl radicals are much more stable than secondary since they cannot undergo facile disproportionation, reaction (16). Dialkyl peroxides are major products of the bimolecular reactions of tert-alkylperoxyl, shown typically for cumylperoxyl (RcOO·) in Scheme 1.6 [69].

Scheme 1.6 Products of α-cumylperoxyl termination [ 69]

The addition of a small molar proportion of tetralin to cumene dramatically reduces the peroxidation rate of the latter, and Russell [70] found that the rate of reaction of RcOO· with tetralinperoxyl (RtOO·) was over a hundred times faster than the rate of reaction of RcOO· with itself:

(19)

As little as 2 mole % of tetralin in cumene results in 100% cross-termination by reaction (19) [70]. It is clear then that the rate of termination may dominate the overall rate of peroxidation and this was seen to be of considerable importance in the peroxidation of branched-chain polyolefins (Section 1.2).

The equations describing the steady state rates of oxidation of moderately reactive hydrocarbons under ambient oxygen pressures and of highly reactive hydrocarbons under conditions of reduced oxygen pressure are (i) and (ii) respectively [2]:

(i)

(ii)

where ri is the rate of intitiation and the k subscripts indicate the equations in the text. In the case of very oxidisable olefins, e.g. 2,6-dimethylheptan-2,5-diene, V, the rate of reaction 6(a) approaches that of reaction 5. Consequently, termination through the delocalised radical, VI, supervenes to relatively high oxygen pressures [71,72].

(20)

The presence of a small amount of a relatively stable carbon radical such as VI or triphenylmethyl may determine the overall rate of oxidation. Thus, stable carbon-centred free radicals are effective retarders for the oxidation of many substrates at low oxygen pressures, due to reactions 17 and 18 [73]. It should be noted that this is not inhibition in the accepted sense of the word, since carbon-centred radicals always react with oxygen with continuation of the kinetic chain and this is more correctly called retarded cooxidation. It will be seen later (Section 1.6.2) that the extended conjugated polyene system produced in PVC during thermal degradation readily scavenges alkylperoxyl to give a delocalised polyene radical which can sacrificially co-oxidise with the main substrate. The same mechanism is probably involved in the antioxidant mechanism of ß-carotene and this will be discussed in Chapter 5.

1.5 Products of Peroxidation

1.5.1 Hydroperoxides and their decomposition products

The major route to the formation of the end products of peroxidation is through the hydroperoxide. This can occur by unimolecular thermolysis and photolysis (reaction 21) or by reduction to give an alkoxyl radical (reaction 22):

(21)

(22)

By far the most effective reducing agents in reaction (22) are the lower valency transition metal ions, notably Fe²+ and Cu+ which are the major source of prooxidant species in both technological and biological systems.

The alkoxyl radical is highly reactive although not as reactive as the hydroxyl radical formed in the analogous Fenton reaction (reaction 15) and it can undergo further reduction, oxidation (or disproportionation), alkyl elimination and addition to a carbon-carbon double bonds [74].

Oxidation of C-H bonds by alkoxyl initiates the radical chain (reaction 23) and is the primary reason why hydroperoxides and transition meal ions are ubiquitous prooxidants in technological media:

(23)

Reaction 24(a) is a termination step when it occurs in a molecular cage reaction with hydroxyl when there is a hydrogen on the a carbon atom:

(24)

However a small proportion of the radicals escape from the cage even in the case of secondary hydroperoxides. Fragmentation of alkoxyl, reaction (25), is the predominent process occurring with tertiary alkoxyl. This is a propagation process which in the case of polymers also leads to molar mass reduction:

(25)

The products are carbonyl compounds: initially aldehydes and ketones and ultimately carboxylic acids. Reactions (23a) and (25) are in competition and, whichever, predominates depends on the oxidisability (by hydrogen abstraction) of the substrate and the stability of the eliminated carbon-based radical (·R") [74]. Reaction (26) is also a propagation reaction which