摘要:

R.I.C. Reviews R.I.C. Reviews, published twice yearly, reviews areas of chemistry of interest to the chemist who has no specialist knowledge of the field under review, but who wishes to keep abreast of the growth of chemistry as a discipline. These reviews should prove useful to students in familiarizing themselves with a particular field. R.I.C. Reviews interprets the significance of chemistry in a wide context and publishes articles on the economic, social and historical aspects of chemistry, as well as on the research and applied sectors. Suggestions for future titles are welcomed. Prospective contributors should write to the Editor, enclosing a synopsis (of about 250 words) indicating the scope of their subject. The preferred length for reviews is 8000 words. Annual Subscription: €2 (R.I.C. members, € 1 )

ISSN:0035-8940    

DOI:10.1039/RR96902FX001

出版商:RSC    

年代:1969

数据来源: RSC

摘要:

ORGANIC E LECTRQC H EM ISTRY M. Fleischmann, B.Sc., Ph.D., A.R.C.S., and D. Pletcher, BSc., Ph.D., A.R.I.C. . . Chemistry Department, The University, Southampton SO9 5N H The special characteristics of electrode processes Methods of investigation, 90 Reactive intermediates in electrode reactions . . .. . . . . 94 Uncharged radicals, 95 Carbanions and carbanion radicals, 97 Carbonium ions and carbonium ion radicals, 98 Other intermediates, 100 . . .. 85 Correlations between electrochemical data and other physical pro- Some further examples of electrochemical reactions of organic perties . . .. .. .. . . .. . . . . . . 103 compounds . . .. . . . . . . . . . . . . 106 The control of electrode reactions, 106 Oxidations, 109 Reductions, 1 12 References .. . . . . . . .. . . .. . . .. 114 At the end of the last and the beginning of this century there was considerable interest in the possibility of synthesizing organic compounds by electro- chemical methods. Much of this work was summarized by Fichter.1 In some respects this is a depressing summary, as the early hopes of achieving selective reactions by electrochemical methods were clearly not realized, mainly because the electrolyses were carried out under conditions where the electrode potential, pH etc. were not controlled. Since the middle 1920s many of the advances in organic electrochemistry have been made by polarography, a technique in which a dropping mercury electrode is used to provide a highly reproducible surface in the construction of currentlpotential curves.However, polarographers have generally been reluctant to prepare and isolate the products of the electrode reaction, a factor which contributed to failure in communication between electro- chemistry and organic chemistry. In recent years, the forecast of a falling cost of electricity due to the advent of nuclear power and a realization that large scale industrial electrosynthesis is a real possibility have led to a resurgence of interest in organic electro- chemistry and research has been directed to the development of new selective routes and the improvement of existing electrosynthetic methods. At the same time it has become feasible to study the electrochemical oxidation and reduction of organic compounds in greater detail, partly because of the development of electronic control systems known as potentiostats, partly Fleischmann and Pletcher 87 because of the recognition and characterization of reaction intermediates and partly because of the considerable development in the understanding of electrode kinetics which has followed the development of numerous new techniques for studying electrode reactions.In fact, it is'now possible to study complex reaction sequences in aetail. The first section of this review (p. 88) deals briefly with the special characteristics of electrode reactions and with the most common methods of investigation, while the second (p. 94) illustrates the generation, nature and manner of reaction of intermediates of electrode reactions. The third (p.103) and fourth (p. 106) sections deal with the correlation of electrochemical data with other physical properties and with some further examples of electro- chemical reduction, oxidation and substitution reactions which illustrate features of electrosynthesis additional to those described in the second section. For obvious reasons, this review is intended to be illustrative rather than comprehensive, and for fuller expositions of various aspects of these subjects, the reader is referred to reviews of electrode mechanisms72J organic polaro- graphy,4-6 synthesis,7-llJl reductions712J3 oxidations,l4-16 substitntion,l7 and the Kolbe synthesis.18-20 Text-books on relevant topics of electrode processes include references 22-27.THE SPECIAL CHARACTERISTICS OF ELECTRODE PROCESSES It is well known that the standard free energy change of a reaction such as is given by Rd + 0, - o x O x + H, - Rd AGO = -zFEO where z is the number of electrons transferred in the process. If we refer immediately to a scale of free energies or electrode potentials (Fig. 1) it is evident that spontaneous reactions using oxygen or air as the oxidant, or hydrogen as a reducing agent, are only possible (in electrochemical terms) within the potential range limited by the reduction of oxygen and the oxidation of hydrogen. This driving force therefore amounts to roughly 0.5 eV or 42 kJ mol-1. By contrast, it is possible to carry out electrochemical reactions between +3.5 V and -2.5 V, even in aqueous solutions, if suitable electro- lytes are chosen (e.g.perchlorates for oxidations and quaternary ammonium salts for reductions). The large driving force for electrochemical processes is therefore of the order of 3 eV or 250 kJ mol-1. Strictly, it is not valid to compare the available driving force to the standard free energy change of any chosen reaction since most processes are irrever- sible; we must therefore estimate the rate constants instead. At the surface potential, 4, and in transferring a charge zFmol-1 a maximum amount of between the metal and an electrolyte solution, there is a difference in electrical work (YZ~F can be done in reaching the activated complex, where a(O< (Y < 1) is the transfer coefficient.The rate of an electrochemical reaction is therefore R.I.C. Reviews 88 - + 4.0 O 3 + 3 H 2 0 + 6 e - - 6OH- ClOh + e - -ClO, - + 3.0 - + 2.0 Ag2++ e- - Ag+ Z10-+H20+2e- 0,+2H20+4e- - C I - + 2 0 H - - 4 0 H - - + I .o radical cation - -H+ radical I I - e - 2H++2e- - H, 1 1 - e - - ov substrate -- -- 2.0 radical t anion - +H + radical 1 +e+ carbanion Zn2++ 2e- - Z n Na++ e- -Na Solvated electrons Therefore, current -- 3.0 - - 4.0 Fig. I . Representative electrode potentials or log I = const. + -.$ azF JRT at constant concentrations, where the product determines the slow step, vi is the order of the reaction with respect to species i which is present at a con- centration ci (for more exact expressions of the effect of the potential difference on the rates of electrode processes see reference 26).Equation 2 shows that we can modify the energy of activation, Ex, of electrochemical reactions to such an extent that they will take place at room temperature even if Ex is large. Fleischmann and Pletcher carbon i u m I ion I - e S + 89 Pot en t ios t a t 1 Osci I I bsco pe t m I / m Y W --L- Pulse profiles Potentiometers F a - Counter electrode Reference t Osci I lator - ---I- - - Fig. 2. Circuit diagram for a potentiostat We can summarize this first point in the following way: electrochemical reactions enable one to introduce a considerable amount of energy into molecules at low temperatures.The order of magnitude of this energy is in fact comparable to that of the strength of chemical bonds and this explains why many of the chemical changes which are observed can take place. It is not surprising that many 'high energy' chemicals which are used as oxidants or reductants in synthesis are made electrochemically and it is clearly of some interest to avoid such intermediate steps in synthesis. Methods of investigation Many methods of investigating electrode reactions which have been developed during the past decade are based on the regulation of the potential of the working electrode with respect to an unpolarized reference electrode using a feedback amplifier (Fig. 2), the current being passed between the working and a subsidiary electrode.Such feedback amplifiers are known as potentio- stats. The potential to be applied between the working and reference elec- trodes is set at the second input of the amplifier. A succession of constant potentials may be chosen to construct a current/potential curve ; alternatively, a slow linear sweep of potential with time will give this curve (potentio- dynamic method). When a dropping mercury electrode is used, the current/ potential curve is known as a polarogram. Figure 2 shows that other potential/time profiles may be imposed on the working electrode. In the case of a triangular potential/time profile with a high sweep rate [0.1-1000 V s-11, the current/potential plot may be recorded oscillographically. This technique is known as cyclic voltammetry and is discussed later.A sine wave of small amplitude applied to the input gives Lissajou figures on the oscilloscope from which the electrode impedance may be calculated; alternatively the impedance may be found using a bridge. R.I.C. Reviews 90 I .5 3.0 Fig. 3. Polarogram of anthracene in acetonitrile containing 0. I M Et,NCIO, When a potential step is applied to the input, a current/time transient is obtained; Fig. 2 shows that square pulses may be also applied. Electrode reactions may also be studied by applying a constant current and measuring the potential/time curve or the potential in the steady state. While this method requires only simple apparatus, controlled potential methods have been increasingly used in the last few years.The advantages as com- pared to constant current techniques will be apparent, since the rates of electrode processes are controlled by the potential, as may be seen from equations 2 and 3. In addition, in preparative work a measure of selectivity in the reactions may be achieved (equation 1). Equation 3 shows that the current would increase indefinitely with potential; in practice, the diffusion of the electroactive species to the electrode, and possibly also of the products away from the electrode, will become rate determining and the current reaches a limiting value. A typical current/ potential curve obtained with a dropping mercury electrode, that is by polarography, is shown in Fig. 3. The portion AC illustrates the addition of one electron to an aromatic hydrocarbon and in the region BC the electrode process is diffusion controlled.The potential at which the current is half the diffusion current is known as the half-wave potential, E,. Polarographic waves are classified as being reversible or irreversible. Jn the first case, the rate of the electrode reaction is fast compared with diffusion so that the shape of the wave is controlled by mass transfer of the reagent and product and the equilibrium at the surface Rd O x + ze-- Therefore ( 5 ) E E, + RT ~ In aox ~ zF aRd Fleischmann and Pletcher 91 where aox and aRd are the activities of the oxidized and reduced forms at the electrode surface. It will be seen that in this case E, h EO. In the second case, the rate of the electrode process is slow compared with diffusion and E, is no longer equal to EO.However, with increasing potential, the current again becomes diffusion controlled. The decision as to whether the electrode process is reversible or irreversible may be made by considering the shape of' the wave. In the second case the wave is drawn out along the potential axis compared to a reversible wake (for exact criteria, see reference 25). In the example shown, and for many other hydrocarbons, the wave AC is reversible but that for the addition of a second electron CD is irreversible. In electrode reactions, it is frequently found that a chemical process precedes electron transfer and only one species is electroactive in a potential region.For example,28 CH,(OH), HCHO + 2e- + 2HS HCHO - + H,O CH,OH ( b ) ( a ) A limiting current (which is controlled by the rate of the preceding reaction) may therefore be realized when b becomes fast compared to a. Such waves are known as kinetic waves and their distinction from diffusion controlled waves is described el~ewhere.~5 Other reaction sequences can also be investi- gated (e.g. following reaction^).^^ The steady-state current/potential data can further be analysed by plotting the initial part of the region AB in Fig. 3, according to equation 4. The linear relation between log I and + is known as a Tafel plot. The variation in position of these plots with the concentration of electroactive species gives information directly about the slow step of the electrode reaction.For measurements under non-steady state conditions, such as in cyclic voltammetry, all the information which has been described above may again be obtained23925 and further data of the reaction sequence may be derived. In the first place, the rate of diffusion in the non-steady state is greater than for steady polarization and the current can therefore reach higher values, as in section AB in Fig. 4. (This increase in current allows the study of the kinetics of faster electrode reactions than is possible by steady-state methods.) Non-steady state diffusion then becomes rate controlling and the current falls (section BC). On reversal of the potential sweep, the starting material is regenerated in the case of simple reactions and a peak, DEF, is observed in the oxidation current.Secondly, for a sequence such as R + e- - R - R ' - + X f A RX' - 3 (RX) 2 some of the intermediate is removed by the chemical reaction and the reoxida- tion peak for R'-- R + e - is therefore diminished.30 In some cases peaks for the oxidation of RX' and (RX)2 may also be observed. In general, the shape of the peaks, and in particular their height and separation and their dependence on the sweep rate, is directly related to the overall kinetics of the electrode reaction. It is R. I. C. Reviews 92 B E Fig. 4. Cyclic voltammogram of anthracene in acetonitrileZ9 containing 0. I M Et,NCIO,, sweep rate 0. I5 V s-l of considerable importance that in this way the kinetics of fast reactions succeeding a slow electron transfer may be measured.Other pulse techniques give essentially similar information and the choice of method will be governed largely by convenience although the interpretation of potential step experiments will usually require fewer assumptions than the interpretation of cyclic voltammograms and may therefore be preferable. Thus, in a single-step experiment, the initial part of the current/time transient gives the rate of the electron-transfer step and the whole transient the rate of preceding steps.27 Using a double step, say first to the cathodic and then to the anodic direction, the kinetics of a following reaction can again be deter- mined. For example, the rate constant, k, for the rearrangement of hydrazo- benzene to benzidine can be determined in this way.31 Further information about the kinetics of electrode processes may be obtained using various translating electrodes which increase the rate of mass transfer.The most widely used of these is the rotating disc electr0de.3~ Information concerning chemical reactions succeeding electron transfer may be obtained using the ring-disc electrode.33 This consists of a disc electrode closely encircled by a ring electrode. The intermediate is generated at the disc and the fraction of the intermediate which is removed at the ring is related to the rate of removal of intermediate by chemical reaction during its transit from the disc to the ring. The illustrations which have been given have assumed that the inter- mediates are not adsorbed and that the coupled reactions take place in 93 FIeischmann and Pletcher solution.Frequently, this will not be the case but it is possible to investigate purely heterogeneous steps in precisely similar ways and indeed simultaneous heterogeneous and homogeneous pathways.34 It will be apparent that very detailed information on the reaction mecha- nisms (i.e. the rate of preceding steps, the rate of electron transfer, the rate of succeeding reactions and data concerning adsorption) can now be obtained by electrochemical methods. As yet, such investigations have been carried out only on a very small proportion of known electrode reactions and although, where this information is available, the intermediates are fully consistent with those postulated in the next section and are in line with those postulated in physical organic chemistry, in other cases the existence of intermediates must be deduced from non-kinetic data such as the nature of products, esr spectra etc.REACTIVE INTERMEDIATES IN ELECTRODE REACTIONS Early workers in the field of mechanistic organic electrochemistry believed that electrochemical reductions took place via the production of ‘active hydrogen’ at the cathode and that the potential at which an organic com- pound was reduced was dependent on the potential energy required by the ‘active hydrogen’ in order to react with the organic compound. Similarly, electrochemical oxidations were believed to proceed via the production of ‘active oxygen or hydroxyl radicals’ at the anode.In recent years it has become clear that the intermediates produced during an electrode reaction are frequently the same as those commonly encountered in other fields of organic chemistry-carbanions, carbanion radicals, radicals, carbonium ion radicals, carbonium ions and, less commonly, intermediates such as biradicals, solvated electrons, transition-metal ions, surface oxide layers and inorganic radicals. Three factors have been mainly responsible for the recognition of these intermediate species. Firstly, the growing use of aprotic solvents, such as acetonitrile, dimethyl formamide, dimethyl sulphoxide, and propylene carbonate, in which the lifetime of the intermediates is much longer than in aqueous solutions ; secondly, the development of improved analytical techniques which allow the identification of minor as well as major products and, lastly, the development of a number of techniques, both electrochemical and non-electrochemical, for the study of transient species.Of the electrochemical techniques, cyclic voltammetry has been the most widely exploited although a number of linear sweep and pulse techniques for the study of short-lived species have been described. Electron spin resonance is by far the most common non-electrochemical technique since it allows the detection and identification of very dilute solutions of free radicals.35 Short- lived radicals have been studied using electrochemical cells in the resonance ~avity.~6 The use of ir, visible and uv spectroscopy has been facilitated by the development of optically transparent electrodes37 which allow the electro- chemical cell to be placed directly in the light path.This technique permits the study of species close to the electrode while actual surface layers may be studied by total reflectance spectroscopy.38 Electrochemiluminescence39 is R.I.C. Reviews 94 another spectroscopic technique which has been used for the study of anion and cation radicals. The species Rf and R- are produced together, either by the use of alternating current or by placing two electrodes in close proximity, and are allowed to react to produce excited molecules, R*, which can then emit light which is characteristic of the system.In this section some of the electrochemical methods of producing inter- mediates are described and some illustrations of their reactions are given. It is important to realize that the behaviour of the intermediates will be different when they are adsorbed on the surface of the electrode from when they are free in solution; this is one of the factors which causes the electrode material to be important in controlling the products of an electrode reaction. Thus the Kolbe reaction RCOOH - RCOO' + e- + H+ - CO, + R' p R - R product R++ e--- has been shown to give widely differing products on platinumls and carbon40-42 electrodes. Moreover, although the Kolbe synthesis is believed to proceed via radical intermediates, no esr spectra have been obtained for them, presumably because the radicals are strongly adsorbed on the electrode.Uncharged radicals Radicals are intermediates in many electrode reactions, both oxidations and reductions, and a number of examples are cited as equations below. (R = alkyl, 4 = phenyl.) 43 45 46 47 RCOOH - R'+ CO, + H+ + e- A l (CH,), - CH; + Al(CH,), + e- OH 0' 0 0' OH 0 -CH (COOEt) , - CH(COOEt),+ e - + 2 C = 0 + H++ e - - q5,C'OH 49 R'+ I - RI + e----- Fleischmann and Pletcher 95 radical + RH RH + alkene I I R - C - C ' I I \ metal al kyl 50 Fig. 5. Main modes of decomposition of uncharged radicals With the exception of a few radicals such as the diphenylpicrylhydrazyl radical, these intermediates are very reactive species and can react in a number of ways.The main modes of disappearance are summarized in Fig. 5. The most common mode of disappearance is dimerization as illustrated by the normal Kolbe synthesis, the formation of pinacol from the reduction of acetone47 and the formation of diphenyl from the reduction of tetraphenyl- ammonium ions.48 However, under suitable conditions, high yields of pro- ducts other than the dimer can be obtained, for example, 4RMgl .-- RCOOH 51 4e- + 4Mg2++ 41-+ PbR, e- + H+ + CO, + R' CH2=CH-cH=CH2=_ dimer RCH,-CCH=CH-CH;- although by-products formed by competing reactions are almost always observed. At the same time, it is important to realize that the products and reaction routes will depend on the concentration of radicals present, i.e.will be dependent on the electrode potential, concentration of electroactive species and other electrolysis parameters. For example, a high concentration of radicals will favour dimerization. Other typical radical processes which have been demonstrated to occur during electrode reactions are the initiation of p~lymerization~~ and chain reactions. An example of a chain reaction is the reduction of benzyl iodide on mercury in aqueous ethanol53 where several times the coulombic yield of the product, benzylmercuric iodide, was obtained. The direct chemical reaction is very slow. Thus the mechanism postulated is R. I. C. Reviews 96 I-.- +H- solvent- +H2 So'Vent +2- Carbanions and carbanion radicals In aprotic solvents, aromatic hydrocarbons may be shown (e.g.by polaro- graphy or cyclic voltammetry) to reduce in two one-electron steps to the carbanion radical and di~arbanion.5~ In the absence of proton donors or other addition agents, the radical anions are relatively stable. On the time scale of an electrochemical experiment, which is determined by the transition of the species through the boundary layer in contact with the electrode (approx. 0.1 s), the process may be reversed by changing the potential and the parent hydrocarbon will be regenerated.29 On a longer time scale, dimeri- zation or disproportionation will take place; the dianions formed will abstract protons relatively easily even from aprotic solvents so that dihydro- aromatic or dihydrodiaromatic compounds are formed at the potential at which the first electron is added. Thus anthracene gives 9, lo-dihydroanthra- cene55 and phenanthrene gives 9,9', lO,lO'-dihydrodiphenanthrene.56 The dicarbanion produced directly at more negative potentials, naturally, also protonates readily; in this case, reversal of the polarization in cyclic volt- ammetry can lead to the radical 4H*57 so that the protonation clearly takes place in two steps.scheme t +- +e- +- +-- +-a +H - +H - +H ' + + + 2 - When a proton donor [e.g. water or benzoic acid154 is added to the aprotic media, the first reduction wave increases at the expense of the second wave until the two waves merge to form a single, two-electron wave.The reduction H'+e- - +H- w 2 #H-+H+- has been postulated, the single wave being obtained because the species 4H- has a higher electron affinity (i.e. it is more readily reduced) than the parent hydrocarbon. A similar scheme may be written for the reduction of the hydrocarbon in the presence of carbon dioxide55 Fleischmann and Pletcher 97 + + e--- +-- + .- + CO, - $COO - + 'COO- + e- - +-coo- CH CH,' + -coo-+co, - + ( c o o - ) 2 and when naphthalene is reduced in the presence of carbon dioxide, the product is 1,4-dihydro- 1,4-dicarboxynaphthalene. It is also possible to react these aromatic hydrocarbon carbanions with methyl iodide to give the dime thy1 di hydroderivative .55 The carbon dioxide addition is a general reaction of carbanions.For example, when stilbene or benzyl chloride is reduced in the presence of carbon dioxide the products are 1,2-diphenylsuccinic acid55 and phenylacetic acid5* respectively. Aliphatic carbaiiions are also common intermediates and they usually occur at more negative potentials in reactions producing radicals. For example, R-+CI- RCI + 2e--- +C-H -C-H+ +CH = CH++ 2e-- C-H, - C-HCN CH, = CHCN + 2e-- CH, = CHCN + C-H,-C-HCN + 2Hf- \C = + 2e- + H+ -- CH3\ C--OH 59 CH 3' and in most cases the final products arise by proton abstraction from the solvent. Thus, the reduction of ethyl chloride and acetone at very negative potentials gives ethane and isopropanol respectively. Compounds, such as acrylonitrile, which contain an activated alkenic group60 may be reduced to form a dianion NCCH2CH2CH2CH2CN which will react with another molecule of acrylonitrile to form the hydrodimer, adiponitrile.This reaction is the basis for a new industrial plant. 1,3-Buta- diene61 and c+unsaturated acids62 are other examples of the many alkenic compounds which reduce in the same way and it is also possible to form crossed hydrodimers from mixtures of activated alkenes.63 Carboniunz ions and carbonium ion radicals It might be expected that in an aprotic solvent, aromatic hydrocarbons would also oxidize in two one-electron steps to give a carbonium ion radical and a carbonium ion 49 55 +H - $H'++ e - +H'+ - +++ H++ e- +H~++ e - or +H.+- 98 R.I.C. Reviews However, the cation species are much more unstable than the corresponding anion species and this simple behaviour is found only for relatively few aromatic hydrocarbons such as 9, IO-diphenylanthra~ene~~.~~ and 1,8- dithionaphthalene.65 The behaviour found for most hydrocarbons consists of an ece mechanism, i.e.the first electron transfer is followed by a very fast chemical reaction, possibly dimerization or reaction between the cation radical and the solvent, to give a species which is oxidized further. The first electron transfer has been shown to be reversible by using very rapid cyclic techniques where the rate of potential change is fast compared with the following chemical reaction.66 The products which have been reported for the oxidation of hydrocarbons such as anthracene are many and varied and perhaps this is not surprising in view of the reactivity of the cation inter- mediates.However, in some cases such as hexamethylbenzene oxidized in acetonitrile good yields of a substituted amide have been reported67 C H 3 ) 3 C H 3 CH 3 + H+ + 2e- \ CH3 CH3 CH3 CH3 CH 3 CH,CN + CH3@H3 CH, CH3 CH, CH, CH3C+= N -CH2 CH3 CH, CH3 CH3 CH2 = CH- CH2R -CH2 \\ CH CO. NH.CH vlH3 - Hydrolysis Moreover, there now seems general agreement that the electrochemical substitution of aromatic species takes place via carbonium ion intermediates.68 Thus, whether the reaction studied is halogenation, cyanation, methoxylation or acetoxylation the mechanism seems to be +H - #I++ H++ 2e- x- +x and not a mechanism involving formation of radicals followed by radical substitution, since substitution only takes place at potentials where the hydrocarbon is oxidized, even when the substituting anion is oxidized to radicals at lower potential^.^^ Furthermore, the distribution of product isomers coincides more closely with a reaction between a carbonium ion and an anion rather than attack on an aromatic system by a radical.70 Aliphatic carbonium ions may be prepared by the oxidation of carboxylic acids,71 alkyl iodides,72 aliphatic hydrocarbons73 and primary amines.74 RCOOH --- R++C02+2e-+H+ CH,I - CH~++ I+ + 2e- =CH - CfHR + H++ 2e- RCH2NH2 - RCH2N'+H2 + e - - RC+H2 +NH2 Fleischmann and Pletcher 3 3 4 .3 99 ROMe ROCOCH H O R + alkene - -H+ CH CN 3 CH c += N RL CH ,CON H R RX 4 R Fig.6. Probable reactions of aliphatic carbonium ions in common solvents A summary of the probable reactions of aliphatic carbonium ions in common solvents is shown in Fig. 6. Carbonium ions formed in electrode reactions show the usual carbonium ion rearrangement. Thus, when neopentyl iodide is oxidized in acetonitrile, N-tert-pentyl acetamide can be isolated as well as N-neopentyl acetamide. 72 CH, lcHlcN CH3--C=N -CH,-CMe3 CH,-C=N CH3- C-CH, I C2H5 Other intermediates + It is also interesting to note that non-classical carbonium ions can be prepared electrochemically. For example, the anodic oxidation of exo- norbornene-2-carboxylic acid gives the non-classical carbonium ion.75 Two interesting biradicals which have been prepared electrochemically are benzyne76 and dichl~rocarbene.~~ Since both are extremely reactive, their preparation is inferred by products from reactions with trapping agents.The small yield of product may be due to low efficiency in the preparation of 100 R. I.C. Reviews the biradicals but is also likely to be due to competing trapping reactions, e.g. reaction with the solvent. The biradicals were prepared by reduction in acetonitrile, dichlorocarbene from carbon tetrachloride and benzyne from o-dibromobenzene. The trapping agents were tetramethylethylene and furan respectively. 2C I- + CC 1; CC 14 + 2e-- N O - NO; + e- Me2C=CMe2 - Me ‘c-c /Me ClO, - C l O i + e- /CH3 ‘CHj There is a number of oxidations where inorganic radicals have been postulated as intermediates.Thus, in acetonitrile, the anodic limit has been found to be strongly dependent on the inert ele~trolyte,~~ which suggests that the initial reaction is oxidation of the anion present. The following mechanism, with perchlorate as the inert electrolyte, has been proposed :78 C104+CH3CN - CH2CN+ HC104 Similarly, the methoxylation of dimethyl formamide in a nitrate base electrolyte is believed to involve the nitrate radical :79 Me/ ‘c’ ‘Me + HNO3 / CH3 NO; + HCON ‘CH3 - HCON Clearly, the intermediates in the oxidation and reduction of substituted hydrocarbons will not always be carbanions and carbonium ions. For example, the intermediates from nitrogen containing organic compounds will frequently have the charge situated on the nitrogen.However, their reactions and properties do not differ greatly from the carbon species. Examples of such charged nitrogen species are shown below. The solvated electron is a probable intermediate in the electrochemical reduction of ben~ene.8~98~ It is only possible to reduce benzene in solvents such as ammonia, hexamethylphosphoramide, ethylenediamine and amines, Fleischmann and Pletcher 8 101 80 R3N - e - + R 3 N + s R3N+H 81 CH,CONHR - e-+ CH,CON+HR SOIVenf_ CH,CON+H,R 82 + e-+ H+ NH2 NH NH - NH in which solvated electrons may be formed. Therefore, it seems likely that in the presence of Group I metal ions the reduction mechanism is Li ( s h ) + e- - Li+(sol.) + e-(soh.) - w products Metal ions in lower or higher valence states may also be used as inter- mediates in reductions and oxidations.For example, M g ( p and A1(1)8~ produced by the anodic oxidations of the metals will reduce 2-methoxy- phenyl mesityl ketone, and nitrosobenzene, azoxybenzene and azobenzene respectively. 3AI++2C6H5N0 +4H20 - 60H-+ 3A13++ c6H5N- H H l l NC6H5 such as AI - AI++ e- or CO(III) will oxidize alkenes or aromatic compounds87 in a sequence cO2+ - co3+fe- and in this case the metal ion acts as a catalyst regenerated at the surface of the electrode. Similar oxidations may be carried out with metal oxides pro- duced on the electrode surface.87 Mn2++ 2 H 2 0 - Mn02 +4HS+ 2e- Mn02 + Rd - Mn2++ Ox + 2e- An intermediate which is likely to be of considerable importance is the superoxide ion generated by the reduction of oxygen in non-aqueous soh- ti0ns.8~7~~ 0; 0 2 + e - - This species will react with organic substrates leading, for example, to the R.I.C. Reviews 102 formation of organic peroxides in a reductive process.Electrochemical auto- catalysis can also be observed.g0 The reactions which have been listed in this section have been formulated as though the intermediates are dissolved in solution. In fact, in many cases these intermediates will be adsorbed on the electrode surface and this will be particularly true for radical intermediates, for example as shown by the cis addition of methyl radicals, generated in the Kolbe reaction, to butadiene.91 Again the formation of methanol by oxidation of acetate ions in basic solu- tions is best explained by the biradical reactiong2 For the complete oxidation of hydrocarbons to carbon dioxide on platinum metal catalysed fuel-cell electrodes, the participation of OH radicals adsorbed on the surface has also been postulatedg3 and in these mechanisms one can indeed see an application of the original ideas of the mechanism of electro- oxidation.Again, it is possible to cite reduction reactions when using catalytic- ally active electrode materials such as platinum black where the original mechanisms involving hydrogen would be more appropriate (cf. hydro- genat ion). 94 AGRd h - -/- AG:;. CORRELATIONS BETWEEN ELECTROCHEMICAL DATA AND OTHER PHYSICAL PROPERTIES For any electrode reaction, a thermodynamic cycle, such as that shown in Fig.7 for the reduction of R, may be drawn up. From the cycle, it may be seen that - FA (6) where AGRd is the overall free energy change for the reduction R + e-+R-, AGEI. and AGE;. are the free energies of solvation of R and R- respectively and A is the electron affinity of R. Since the formal electrode potential, EO, (the potential on the scale of the normal hydrogen electrode) is given by EO = - AGRd/F and for a reversible reduction Eo E= EFd, it may be con- cluded that (7) Ey = A + - 1 [AGE,. - AGri.1 F Similarly for an anodic reaction we can derive the expression Fig. 7. Thermodynamic cycle for the reduction of compound R 103 Fleischmann and Pletcher where I is the ionization potential of R and AGS.is the free energy of solva- tion of R+. It is important to note that these expressions only hold for reversible electrode reactions in the absence of fast following chemical reactions removing R+ or R- and in the absence of complex formation between R+ or R- and the solvent, or ion-pair formation between R+ or R- and the base used directly to correlate Ey with ionization potentials95 and EFd with electrolyte ions. With these reservations in mind, equations 7 and 8 may be electron affinities.95 The results of such correlations are shown in Figs 8 and 9 pyrene, chrysene, phenanthrene and triphenylene. Both EFd and Ey were for a series of aromatic hydrocarbons-anthracene, 172-benzanthracene, obtained in acetonitrile and the Ey were obtained at high voltage sweep rates in order to eliminate the following reaction which R+ would undergo.To within experimental error, linear plots are obtained for both correlations. Therefore, it must be concluded that the free energy terms in equations 7 and 8 are either reasonably constant or vary linearly with potential. It would not be surprising if these free energy terms remain constant for series of similar large molecules. As the electron affinities and ionization potentials may be calculated for aromatic systems, for example by simple Hiickel theory, the correlations may also be made between E, and the orbital energy, i.e. with the level of the lowest unfilled or highest filled molecular orbital.96~97 For alternate hydro- carbons these levels are symmetrically placed with respect to the non-bonding orbital and this fact also allows correlation with the frequency for the first electronic tran~ition.~~ It has also been shown that EFd and Ey for the discharge of aromatic hydrocarbons may be correlated with the frequency of the charge-transfer transition of the molecules in the presence of donor or acceptor molecules respectively (e.g.hexamethylbenzene99 and tri- fluorenoneloo). These correlations are also based on a relation between the frequency of the transition and I or A . A number of such correlations for a variety of compounds has appeared in the literature and, despite the fact that in some cases half-wavepotentials for irreversible processes or processes including a coupled reaction have been applied, good correlations have been obtained. For irreversible cases, the term AGEi.- FA determines the energy of the final state of the reaction while AGEI. - F+ determines the energy of the initial state. There will therefore be a linear relation (in the first approxima- tion) between these energy terms and the free energy of activation, i.e. a is an example of such a relationship since + changes the free energy of activa- ‘linear free energy correlation’. Indeed, equation 4 relating log I to azF+/JRT tion in a linear manner. Correlations between Ey and A will therefore still be observed provided AGEi. and AGE,. remain constant, the reactions have similar pathways and the heats of adsorption of R and R- are also independent 104 R .I. C. Reviews I .6 I .4 + 1.2 E px (V) Fig. 8 (upper) and Fig. 9 (lower). Correlations of €yx with ionization potential and Ey with electron affinity for a series of aromatic hydrocarbons of the nature of R, since these heat terms also determine the energies of the initial and final states. Similar linear free energy relationships can be obtained between half-wave potentials for compounds RAX and the structure of the compounds as expressed by the Hammett-Taft po or p*o* terms, i.e. (AE+)X == (E+)X - (E&)H = fn,R'X for a benzenoid series ( i e . A is an aromatic series) or (AE+)x = (E+)x - (E+)H = P*~,RQ*X (9) Fleischmann and Pletcher I .8 (10) 105 for a non-benzenoid series, where (E,)x and (E& are the half-wave poten- tials of the substituted and unsubstituted molecule respectively (substituent X) and p n , ~ , the reaction constants, measure the susceptibility of R to the effect of X and are independent of the nature of X, while OX, the substituent constants, depend on the position and nature of X but are independent of R.These substituent constants are determined by solution phase reactions. Linear correlations of the type 9 and 10 therefore relate the electrode process to other known reactions and indicate uniformity of reaction mechanism; their uses have been reviewed elsewhere.lo1 102 As has already been explained (p. go), the development of electronic potentiostats allows the systematic application of this idea, for example, I 04 SOME FURTHER EXAMPLES OF ELECTROCHEMICAL REACTIONS OF ORGANIC COMPOUNDS This section is divided into two parts; the first illustrates the importance of controlling the electrode potential and solution composition to give selective reactions and the second illustrates further reaction types, many of which could be applied to synthesis.CHl-NH, 2e- t 2Hi-, @OH OH The control of electrode reactions As early as 1898 it was shown that nitrobenzene could be reduced to phenyl- hydroxylamine at low negative and to aniline at more negative potentials. CH= NH CH=NOH VH + H,O / / OH Q"" 2e-+2H+- OH I05 I 06 R.I.C. Reviews It is interesting to note that the stereochemistry of an electrode reaction can also depend on the potential.For example, in the reduction of benzil the ratio of cis to trans stilbenediol formed is potential dependent.107 Furthermore, the course of a reaction may be changed by applying a pulse profile such that several different reactions take place successively and repetitively. An example is the alternate generation of carbanions and carbo- nium ions leading to electrochemiluminescence. An application to synthesis is the variation in the reduction mechanism of nitrobenzene caused by reoxidizing the intermediate, phenylhydroxylamine, before it has time to azoxybenzene NO / 0 1 (y="o (y-yJ t H H hydrazobenzene H2N m N H 2 benzidine '08 diffuse away from the electrode surface.Thus, by choosing the potential for the reduction step it is possible to accumulate the coupling product, azoxy- benzene, or its reduction product. In effect, by applying a suitable square- wave potential/time profile, benzidine may be prepared directly from nitro- benzenelog and it is clear that many other products could similarly be made directly from nitrobenzene by choosing suitable pulse profiles. It will be apparent that the duration of the pulses is an additional control variable. A question of key importance for achieving a selective reaction is whether the desired product is itself electroactive at the potential at which the reaction is carried out. The half-wave potential for the substrate and product under the experimental conditions will be a useful guide. For example, it is clear that it will always be difficult to find selective oxidation routes for the forma- tion of phenols or acetates since these are usually more easily oxidized than the parent aromatic hydrocarbon.On the other hand, cyanation of aromatic compounds is a favourable reaction since the products have more positive half-wave potentials. Polarographic data will not necessarily be an accurate guide for the course of a reaction since large scale preparations are carried out on a longer time scale and can, therefore, be affected by disproportiona- Fleischmann and Pletcher 107 tions. Thus, dihydroaromatic compounds may be formed at the potential at which only one electron is added to the hydr~carbon.~~ The control of electrode reactions by the correct choice of the solvent has already been implied, for example in the discussion of Fig.6 and in the discussion of the protonation of radical anions in aprotic solvents. In aqueous solution, the pH is of key importance and determines whether protonated or unprotonated species will be electroactive. These effects are also apparent in aprotic media where the presence of base (pyridine) can have a marked effect on the electrode mechanism. Examples which have been observed include the simple, two-electron oxidation of anthracene in the presence of pyridine to give a stable product in absence of pyridine formation e - ' y ? ' ' C H M e M e 0 M e o a c ' y H Me dimer -Meon 3::; & M e 0 = I O M e 108 O M e CH ,CHMe M e 0 Q O M e OMe R.I.C.Reviews Oxidat ions In addition to the reactions listed earlier, other oxidation processes can be formulated as proceeding through carbonium ion intermediates, for example, ring closures, expansions and contractions CH,CH,CH,COO- -2e- -'02- CH,CH,CH; r CH2- CH, 4- other products 40 75 l R \ / Me0 CH2 and the methoxylation of furans may be formulated as 1 Me0 O : M e On the other hand, many other oxidation reactions may be formulated as proceeding through radical intermediates. For example, the Kolbe and the related Brown-Walker reactions have in recent years been used for several 109 Fleischmann and Pletcher interesting syntheses which proceed via radicals.Examples are COOH 114 2 EtOOC (CH,),COO- (EtOOC),CH - - CH, - C H (COOE t)2 formation - OEt -Ze--2Cok EtOOC(CH 2)GCOOEt Other examples include the polymerization of vinyl acetate, vinyl chloride and methyl acrylate initiated by radicals formed by the Kolbe reaction.116 Radicals may be prepared by the anodic oxidation of the sodium derivative of diethyl malonate,46 ethyl acetoacetate,46 ethyl phenylacetate46 and nitro- paraffins117 as well as less common anionic species. In the absence of reactive substrates, the radicals will dimerize, but clearly such radicals are particularly useful in synthesis, as in their reactions with alkenic bonds. A reaction scheme for the oxidation of diethyl malonate in the presence of vinyl ethyl ether118 is OEt OEt I I cH(COOEt), a dH(COOEt), CH2=CHoEt- (EtOOC),CH-CH,-tH I EtOOC I-'- I + EtO G O E t OEt (E t OOC), - CH - CH ,-CH (OEt), - So'Vent (EtOOC)2CH-CH2-CH- OEt EtO' EtO' 110 R. I .C. Reviews This scheme illustrates that radicals may dimerize, hydrogen abstract or be oxidized further to give carbonium ions which may react with the solvent, cyclize, or lose a proton, and is an example of the build up of complex mole- cules from simple starting materials. It is possible to use other alkenes such as cyclohexene, styrene or butadiene and the other carbanion species listed above .I18 Several examples of elimination reactions have been reported during electrochemical oxidations. Examples include the oxidation of p-substituted phenols.OH r- 1 - HBr MeOH 0 1 CH,=CH-CHO+4H++ H2O- CH,-CH=CH2+4Hg2++ U A number of indirect oxidations, where inorganic intermediates generated at the electrode react with substrates [cf. Mg+, Al+ in reductions; MnOz, Co3+ in oxidations], have been reported. Examples include the electro- chemical modification of the Clauson-Kaas methoxylation of furan where the bromine is generated at the anode,l21 M e 0 O k M e the oxidation of propylene where the mercury(I1) is regenerated electro- chemically, 122 the oxidation of propylene to propylene oxide by electrochemically generated hyp~chloritel~~ and the oxidative reaction of alkenes with carbon monoxide and methanol 121 I I9 Fleischmnnn and Plefcher 2 H g r 11 1 in the presence of platinum carbonyls to give methyl esters of a,,B unsaturated acids1Z4 C6HSCR =CH, +CO +-OMe - C6H,CR= CHCOOMe Another example of an anodic reaction which is extensively used industrially is the perfluorination of aliphatic hydrocarbons in anhydrous hydrogen fl~oride.1259~26 The mechanism of these reactions, which clearly must be complex, is not fully understood.Reductions Processes analogous to those described for the reduction of aromatic hydro- carbons are also observed in heterocyclic series. Thus the reduction of pyrimidine is explained by the scheme127 H H+ I29 R.Z. C. Reviews 112 In heterocyclic series, ring expansion has also been observed131 131 and several different ring closures have been reported, amongst which are H OH I H I 4e:i2+2 COO H COOH CH = C(COOH), I I Non-heterocyclic ring closures which have been observed include I32 I33 I34 I33 which presumably arises from intramolecular radical coupling.An example which may be attributed to carbanionic attack on an activated alkenic bond I35 mo ,++ CH, -CH-CH,-CH-CH, -H,O NH ___ N' H2 L 'OH CH3-C == CH -C-CH, 6e-+5H+ CH3-CH-CH2-CH-CH3 c HN'OH HONH NH2 HONH - ? = + I I CH2 CH = CHCOOEt 2H+ I is the intramolecular hydrodimerization, , CH.! II =CHCOOEt I ,CH CH2 'CH ___L I m CH2CooEt CHI ,C-H - C-HCOOEt CH2COOEt \ CH2 CH2 \ /CH =CHCOOEt CH2 Fleischmann and Pletcher 113 The reactions of activated alkenes, such as hydrodimerizations, may be used for a wide variety of organic syntheses many of which would lead to industrially useful products.What is more, such reactions can often be carried out in aqueous media using tetraalkylammonium p-toluenesulphonate (McKee’s salts) as electrolyte and solubilizing agent. As the work up and extraction of products from electrosynthesis will frequently be simple compared to conventional processes because of the absence of reagents, it is likely that selective electro-organic syntheses will find increasing application as additional and alternative reaction routes. REFERENCES 1 F. Fichter, Organische elektrochemie. Dresden: Steinkopf, 1942. See also M. J. Allen, Organic electrode processes. New York : Reinhold, 1958.2 C. L. Perrin, Prog. phys. org. Chem., 1965, 3, 165. 3 P. 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ISSN:0035-8940    

DOI:10.1039/RR9690200087

出版商:RSC    

年代:1969

数据来源: RSC

摘要:

THE CHEMISTRY AND PHYSICS OF ENZYME CATALYSIS S. Doonan, B.Sc., Ph.D. . . . . . . .. . . . . .. 117 Deportment of Chemistry, University College, Gower Street, London W C I Enzyme structure . . Mechanism and catalysis . . . . . . .. . . .. .. 123 Possible factors involved in enzyme catalysis . . . . . . . . 127 Studies of individual enzymes . . . . . . . . .. . . 131 Lysozyme, 13 1 Chymotrypsin, 136 Carboxypeptidase A, 138 Conclusion . . . . . . . . .. .. . . . . .. 140 Acknowledgments . . .. . . .. .. . . .. . . 141 References . . .. . . . . . . .. . . .. .. 141 Ten years ago, in conclusion to a review of current knowledge concerning the mechanism of enzymic catalysis, Lumryl stated that ‘there is much ill-defined swamp and little firm ground’. Progress in our understanding of the mode of action of enzymes in this decade has been spectacular and it is now possible, for a small number of enzymes, to describe both the mechanisms of the reac- tions which they catalyse and the factors involved in the catalysis of these reactions : the necessary distinction between mechanism and catalysis will be discussed below.In the present review, the nature of the problem of enzyme catalysis will be defined, some of the more important factors which have been proposed to account for the activity of enzymes will be discussed and, finally, the results of recent chemical and physical investigations of selected enzymes will be described briefly to illustrate these general considerations. More extensive coverage of these topics may be found in recent reviews by Doonan, Vernon and Banks2 and Koshland and Neet.3 ENZYME STRUCTURE Enzymes are globular proteins and hence are polymers constructed from the twenty L-a-amino acids whose characteristic side chains are listed in Table 1, plus the imino acid proline. The amino acids are joined together by their amino and carboxyl functional groups to produce a polypeptide chain of the type shown, where R1, R2 etc. represent the distinguishing side chains: I I I H2NCHCONHCHCO... . . NHCHCONHCHCOOH I Rl R n R2 R”- I A free amino group occurs at one end of the chain (the N-terminus) and a free carboxyl group at the other (the C-terminus). In addition to amino-acid residues, some enzymes contain either a metal ion or a small organic molecule which is essential for enzymic activity. Examples are the zinc ion in carboxypeptidase and the pyridoxal-5’-phosphate cofactor of aspartate aminotransferase ; these enzymes are discussed on Doonan 117 9 pages 138-140 and 124-126 respectively.However, even in these cases, it is the protein part of the enzyme which is primarily responsible for its catalytic capabilities and it is with the relationship between protein structure and enzymic catalysis that this review is concerned. ~~ ~~ ~ R Side chain H - C H3- cH3>CHCH2- CH3 CH3CH2 >CH - CH3 HOCH, - CH3CH - I OH Table 1. Structures of the side chains of the amino acids occurring in enzymes NH2CHCOOH R I Amino acid Glycine Alanine Valine Leucine lsoleucine Serine Th reon i ne Cystine Cysteine (Half cystine) Methionine NHzCHCOOH R I Amino acid Phenylalanine Tyrosi n e Tryptophan Aspartic acid Asparagine Glutamic acid Glutamine Lysine Arginine Histidine SCH, - I SCH2 - HSCH2 - CH3SCH2CH2 - The imino acid proline has the structure 'i H Protein structure is conveniently considered at two levels.Since the struc- tures of the constituent amino acids are known, and it can be assumed that the amino-acid residues are joined by peptide bonds, then once the numbers of residues of each type and the sequence in which they occur in the poly- peptide chain are known, the covalent structure of the protein molecule is completely specified. This is usually referred to as the primary structure.More important for the correlation of structure and activity, however, is the HO 0 CH2 - a ! - c H 2 - - CH2COOH - CHzCONHz - CHlCH2COOH - CH2CH2CONHl - CH2CHzCH2 N HC N H2 II +NH CH; /- 0 COOH 118 R Side chain R. I.C. Reviews conformation of the polypeptide chain and the arrangements of the individual amino-acid side-chains in space. These two aspects of protein structure will be considered separately. The chemical methods for 'the determination of the primary structures of proteins have been exhaustively reviewed.4 In outline, the basis of currently used methods is the Edman degradation process5 in which the N-terminal amino acid is removed from the polypeptide chain by the sequence of reac- tions shown below.The terminal amino acid is isolated and identified as the I I NHPh PI1 - N = C = 5 + NH2CHCONHCHCO.. . .- P" 9 Ph - NHCNHCHCONHCHCO . . . . I I R2 Rl HN, I R2 II s R, R,-CH -CO I \ /NPh C I I 3 phenylthiohydantoin derivative, whilst the remainder of the peptide chain is left intact. Repeated application of the degradation process yields the primary sequence of the polypeptide. In practice, technical difficulties limit the application of this technique to polypeptides containing about 20-30 amino-acid residues. For example, the structures of the 18-residue peptide apamin6 and the 26-residue peptide melitin,7 both isolated from bee venom, have been determined by repeated application of the Edman degradation technique.Since the amino-acid chains of enzymes are very much longer than those of the peptides mentioned above, and may contain from 100 to 1000 amino- acid residues, their primary structures are not immediately determinable by application of the Edman technique. In general, the polypeptide chain is first degraded into smaller peptides, these smaller fragments are separated from one another, and their primary structures are determined. The problem of the order in which these smaller fragments occurred in the native protein must then be solved; this is done by the method of overlaps, illustrated in Fig. 1. The chain is represented by a series of amino-acid residues (R11, R12 etc.) and the dotted lines represent points of hydrolysis.The first hydrolysis yields a series of peptides PI, PZ etc. whose structures may be determined using the Edman method. It is then necessary to decide in which order these 119 Doonan I Fig. I . The method of overlaps applied t o the determination of the primary structures of proteins. peptides occurred in the original structure; that is, whether the original sequence was PlP2P3P4.. . or P I P ~ P ~ P ~ . . . or P3P4PlP2.. . and so on. A second hydrolysis is carried out, under different conditions from the first, to give a new set of peptides Pi, Pi, Pi etc. These new peptides are examined for portions occurring in two or more of the first set. For example in Fig. 1, peptide Pi will contain amino acids both from PI and P2.Similarly Pi con- tains parts of P2 and P3 and Pi contains parts of P3 and P4. Hence the original sequence must have been P1P2P3P4 . . . . In practice, three or four different methods of hydrolysis are usually required to provide all the overlaps. The methods of hydrolysis used depend on the selectivity of enzymes which catalyse the hydrolysis of peptide bonds. For example, the enzyme trypsin catalyses the hydrolysis of peptide bonds involving the amino acids lysine and arginine whilst the similar enzyme chymotrypsin is most active in the hydro- lysis of peptide bonds involving the aromatic amino acids phenylalanine, tyrosine and tryptophan; an account of methods used in the hydrolysis of proteins is given in section V of reference 4.Primary sequence determinations are technically difficult and require a large amount of starting material. For these reasons, complete structures are known for only a few rather small enzymes which are available in large quantities. An example is the enzyme chymotrypsin which is disussed in detail later (pp. 136-138). The primary sequence has been determined by Hartley8 and by Keil and S6rm.g The primary structure, as presented by Matthews et aZ.10 is given in Fig. 2; the amino acids are represented by the first three letters of their names except in the cases of isoleucine, asparagine and gluta- mine which are abbreviated to ile, asn and gln respectively. The enzyme con- tains 241 amino-acid residues (the numbering system in Fig.2 is that for a 120 R. I.C. Reviews NI Fig. 2. The primary structure of a-chymotrypsin (from reference 10). c precursor, chymotrypsinogen, which has four more amino-acid residues than chymotrypsin) in three polypeptide chains. The chains are joined together by disulphide bridges from the amino acid cystine (Table 1); similarly there are three intra-chain disulphide bridges. Primary sequences such as that in Fig. 2 tell us very little about the activity of enzymes. For example, chemical evidence (p. 136) has shown that two important amino-acid side-chains for the activity of chymotrypsin are those of histidine-57 and serine-195. These residues are widely separated in the primary structure and it is not possible to see from the sequence given how they contribute to the activity of the enzyme. In order to solve this problem it is necessary to determine the conformation of the peptide chain and the orientations of the amino-acid side-chains in space.Information of this sort can only be obtained by x-ray diffraction analysis. Doonan 121 Fig. 3. Conformation of the peptide chain of a-chymotrypsin. The determination of the structures of small molecules by x-ray crystallo- graphy is now largely a routine matter, but this is not the case in protein structure analysis due to the size and complexity of protein molecules. Indeed, the classical methods of x-ray diffraction analysis are not applicable in these cases. The method of isomorphous replacement was developed by Kendrew and his associates to overcome these problems, and was used to determine the complete three dimensional structure of the oxygen-carrying protein myoglobin.16 Lack of space precludes even an outline description of x-ray diffraction studies of proteins, but the subject has been extensively reviewed.l2> l 3 An example of the type of results obtained is shown in Fig.3. This figure R.I.C. Reviews 122 gives a simplified diagram of the structure of chymotrypsin determined by Matthews, Sigler, Henderson and Blow.lo Only the conformation of the peptide chain is shown together with the side chains of residues histidine-57 and serine-195 (which in this structure carries a tosyl group). It can be seen that the conformation of the peptide chain brings these two important amino- acid side-chains into close proximity.A detailed discussion of the relationship of this structure to the activity of chymotrypsin is given later (pp. 136-138). At the present time complete three-dimensional structures of only a very small number of proteins have been determined and very few general observa- tions about the relationships between primary and higher order structures can be made. In general, the non-polar amino-acid side-chains are found in the interior of the molecule where they are well shielded from the solvation shell of the protein; the polar side chains are on the surface of the molecule and interact directly with the solvation shell. The main forces stabilizing the struc- ture appear to be van der Waals’ interactions between non-polar side chains, and hydrogen bonds which are formed between the carbonyl oxygen and amide hydrogen atoms of peptide bonds which are brought into the correct juxtaposition by the folding of the polypeptide chain.Determination of the three-dimensional structure of a protein requires prior knowledge of the primary structure and since the latter is more easily determinable, it would obviously be of great advantage if the three-dimensional structure could be calculated from the primary structure. It is widely held that a connection between these two levels of structure must exist. The genetic material of any cell, that is, the nucleic acids which it contains, is thought to specify only the primary sequences of the proteins which the cell produces.Once the proteins are synthesized, they then fold spontaneously into the correct three-dimensional configuration. If it is assumed that the configura- tion achieved is that of maximum thermodynamic stability then, in theory at least, it is possible to calculate this configuration. However, recent evidence2 tends to suggest that the formation of a particular three-dimensional structure is determined by kinetic rather than thermodynamic factors. If this is so, then there is no possibility, in the foreseeable future, of predicting the complete structure of a protein from its known primary sequence. This being the case, the structural information which is required for the interpretation of enzyme activity must be obtained by the difficult and time- consuming techniques outlined above, and the rate at which new structures are forthcoming is likely to be small.However, the few structures which have been reported provide a wealth of information for the correlation of enzyme structure with activity, the problem to which the remainder of this review is devoted. MECHANISM AND CATALYSIS In outline, the events occurring during an enzyme catalysed reaction may be described in terms of three distinct stages, namely: (a) combination of the molecule or molecules (usually termed substrates) which are to undergo reaction with a particular region on the enzyme surface (the active site) to form the so-called Michaelis complex ; (b) covalency changes in the substrate ; (c) diffusion from the protein of the products of reaction.Given such a Doonan 123 sequence of events, then the term mechanism should properly be used to describe the interactions which the substrate makes with amino-acid side- chains of the protein to form the Michaelis complex, the number and struc- tures of the intermediates lying on the reaction pathway between the Michaelis complex and the formation of products, and the order in which the products diffuse from the enzyme surface. A simple, generalized scheme for an enzyme catalysed reaction is given below, where E, S, and A and B represent the enzyme, substrate and products respectively. Mechanism then describes the structures of the species ES, EAB and EA, the interactions which the substrate and products make with the protein in these species, and rates of the unit steps in the reaction sequence.In these terms, the mechanism of action of a variety of enzymes is, at least in part, understood; the qualification is necessary since in no case are the values of all the rate constants associated with unit steps in the process known. It must be emphasized that such a description of an enzyme catalysed reaction tells us what happens but not why these events occur at rates which are so much greater than those of similar processes in non-catalysed reactions ; this is the problem of catalysis. Catalysis is not simply derivable from mechanism and the information required to explain the catalytic effect of an enzyme is not included in a description of its mechanism of action.This distinction between mechanism and catalysis may be illustrated by a specific example, namely the enzyme aspartate aminotransferase which catalyses reaction 2. The enzyme from pig heart muscle has been purified and COOH I COOH I COOH I CHNH, co COOH I CHNH, I 1 CH2 + I CH2 CH2 I I I COOH aspartic acid co e ‘ + ( 3 4 2 I CH2 I COOH glutamic acid CH2 I COOH oxaloacetic acid CHO COOH a - ketoglutaric acid characterized,14J5 and a complete analysis of the steady-state kinetics of the enzyme catalysed reaction has been carried 0ut.l6J7 This enzyme depends for its catalytic activity on the presence of one or other of the vitamin B g deriva- tives pyridoxal-5’-phosphate, I I 0 or pyridoxamine-5’-phosphate.124 R. I.C. Reviews 0 II H o Z c H 2 ~ O H I These species are tightly bound to the enzyme surface and are referred to as cofactors for the enzyme. There is abundant evidence (for a summary see reference 18) that four intermediates, excluding Michaelis complexes, lie on the reaction pathway and that the mechanism of the reaction may be repre- sented by equations 3 and 4, where E-CHO and E-CHzNHz represent the k2 + H 2 0 - H,O - L kl COOH I E-CHO + NH2CH I CH2 E - CHl- N =C I COOH + H2O COOH I d k5 Y k6 - H,O - H,O - k 7 k8 +H,O L +H20 - k l l A I CH2 I CH2 CH2 I COOH I COOH E - CYNH, + CO I I COOH COOH I E - CH=N -CH I CH2 I k I2 -H,O CH2 I COOH Doonan CH2NH2 CH3 COOH k3 I k4 CH2 E-CH=N-CH I I COOH COOH I E-CH2NH2 + CO I CH2 I COOH k9 L 1 kI0 COOH E - CHO + NH2CH I COOH I CH2 E - CH2- N=C I CH2 I I COOH I CH2 I CH, I COOH 125 Factor ~~ Table2.Comparison of rate constants in the enzyme catalysed and non-enzymic transamina- tion reactions Enzymic reaction Rate constant Value (s-1) Non-enzymic reaction 2.0 x 10-6 250 I100 I00 1.25 x los - 1 . 1 x 107 1 . 1 x 109 - 9.1 x 10-6 0.8 x 10-6 k3 k4 k9 k o 900 forms of the enzyme containing pyridoxal- and pyridoxamine-5’-phosphates respectively. The rate determining steps in reaction sequences 3 and 4 are the prototropic shifts characterized by rate constants k3, k4, kg and klo; values of these four rate constants have been determined2J6J7 and are given in Table 2.After the initial demonstration by Snell and his co-workerslg of a trans- amination reaction when pyridoxal was incubated with amino acids in the absence of the enzyme, an extended investigation of the non-enzymic reaction between the natural substrates and cofactors of aspartate aminotransferase was carried out.16 The reaction mechanism in this model system parallels precisely that of the enzyme catalysed reaction; that is, the sequence of events is the same as that given in equations 3 and 4 but with E-CHO and E-CH2NH2 replaced by the free cofactors.Moreover, the values of nine of the 12 rate constants in the reaction sequence were evaluated; the values of k3, kg and klo are given in Table 2. Comparison of the figures in Table 2 yields information of considerable interest since they provide a direct measure of the catalytic effect of the protein part of aspartate aminotransferase on individual steps in the reaction seq- uence; these catalytic factors can be seen to range from 107 to 109. Hence the situation arises where the presence of the catalytic protein enhances the rates of processes which are mechanistically similar by very large factors. There is nothing in the formulation of the mechanism of the enzyme-catalysed reaction given in equations 3 and 4 which accounts for such a rate enhancement; thus the problem of catalysis is not solved by a statement of mechanism.The description of the mechanism of enzyme catalysed transamination given above is incomplete insofar as the groups in the protein which interact with the bound substrate have not been specified. It might, then, be assumed that once these groups have been identified, the origin of the catalytic effect of the protein will become obvious from the complete mechanistic description. This, however, is not the case. If attention is focused on the prototropic shift in reaction sequence 3, it may reasonably be supposed that the process involves suitably positioned acidic (-AH) and basic (-B) amino-acid side- chains in the protein which function as proton donors and acceptors.Accord- ing to the currently accepted formalism of enzymology the process would then be written as in Fig. 4, where the continuous line represents the enzyme surface, @ denotes the phosphate group and R is the substrate side chain. This more complete formulation of the reaction mechanism does contain 126 R. I . C. Reviews L 1 Fig. 4. The rate-determining step in enzymic transamination. some information concerning catalysis but only in the relatively trivial sens.: that the reaction is acid-base catalysed; it does not give any indication as to why the rate enhancement factors are so large. General acid and general base catalysis are well known in physical organic chemistry but the rate enhance- ments observed rarely exceed a factor of 10.For example, in the imidazole catalysed hydrolysis of acetyl tyrosine ethyl ester, the rate of the general base catalysed hydrolysis was found to be only tenfold greater than that of the non-catalysed hydrolysis ;20 this rate enhancement is several orders of magni- tude less than that observed for the catalysis of hydrolysis of this substrate by the enzyme chymotrypsin (see later). In general, the problem remains as to why acid-base catalysis by the side-chain groups in the active site of enzymes is so remarkably effective. One explanation of this effectiveness, which has frequently been assumed, arises from a misconception of the significance to be attached to formal mechanistic diagrams such as that in Fig. 4.Such diagrams are frequently interpreted in terms of a synchronous process (the push-pull hypothesis21), that is, in terms of a concerted proton donation by -AH and abstraction by -B. Large catalytic factors are then assumed to arise from the acid and base groups acting in concert rather than in a sequential fashion. In fact the arrows in Fig. 4 simply indicate which bonds are made and broken in the passage from one enzyme-substrate intermediate to the other and contain no informa- tion about the timing of these events. Concerted acid-base catalysis has not been conclusively demonstrated for any enzyme-catalysed reaction, and even if such processes were concerted there is no reason to suppose that acid and base groups acting together would be more effective in catalysis than the same groups acting sequentially.POSSIBLE FACTORS INVOLVED IN ENZYME CATALYSIS In the previous section a distinction was drawn between mechanism and catalysis in enzymic reactions and in particular it was argued that acid-base catalysis cannot account for the large rate enhancement effects of enzymes. A similar point must be made in connection with the possible catalytic signifi- cance of formation of covalent enzyme-substrate intermediates. This is a feature of the mechanism of action of a variety of enzymes,3 but again one which cannot contribute significantly to their catalytic effects. An example should help to clarify this point. Doonan 127 Several enzymes have been studied (chymotrypsin, trypsin, elastase, throm- bin, subtilisin) which catalyse the hydrolysis of ester and amide linkages (reaction 5, where X is -OR’ or -NHR’).There is good evidence,2J2 particu- larly in the case of chymotrypsin, that the reaction pathway includes an inter- RCOOH + HX RCOX + H,O- i BH R R ( 5 ) mediate formed between the acyl group of the substrate and the hydroxyl group of a serine residue in the protein. The process may be written as in is B: - X - B H + +H20 k 2 histidine residue R (see later). has equation a basic 6 group where which -OH represents been o - c = o the identified R I serine side as the chain imidazole r hydroxyl 0 - H + side + group R C = O OH 1 I chain and of -B (6) a A hypothetical alternative scheme, still involving group B but not covalent intermediate formation is shown in equation 7.Here, the reaction sequence involves direct attack of a water molecule on the substrate. It is evident that, for the formation of a covalent intermediate to contribute to catalysis, both kl and k2 in equation 6 must be much greater than k3 in equation 7; this could only be the case if the serine hydroxyl group was both a better attacking group than water and a better leaving group than -X (-OR’ or -NHR’). In general, serine hydroxyl groups in proteins are not good nucleophiles or leaving groups, and, given the reaction mechanism in equation 6, the problem of catalysis by chymotrypsin and similar enzymes remains.Another possible catalytic factor which may be proposed on the basis of reaction schemes such as those in Fig. 4 and equations 3,4, 6 and 7 is the so- called proximity effect.3 These reaction sequences involve formation of complexes between either two catalytic groups and one substrate molecule or one catalytic group and two substrate molecules. It might be supposed that the formation of such ternary complexes would lead to an improved collision probability (i.e. an increased relative concentration of reactants) with con- comitant rate enhancement. An analogous effect is well known in physical organic chemistry in the phenomenon of ‘intramolecular catalysis’.2392* To give a single example, hydrolysis of the propanol thiolester of 4-(4’-imidazoly1)- butyric acid proceeds with intramolecular catalysis by the imidazole group (equation 8) and is about 3 x lo6 times as fast as similar reactions in which 128 R.I.C. Reviews + PrSH imidazole groups do not p a r t i ~ i p a t e . ~ ~ The analogy between such reactions and those occurring on the surface of an enzyme is obvious, and it might be thought that the proximity effect would lead to similar large rate enhance- ment factors in enzyme-catalysed reactions. Koshland26~~7 has calculated, however, that the proximity effect will lead to a decrease in rate for many enzyme-catalysed reactions and for those cases in which rate enhancement is to be expected the calculated factor is far less than that which is observed in practice. Koshland and Neet3 explain this effect in terms of the increased relative concentrations of substrates and reactive groups being partially or completely offset by the very low absolute concentration of enzyme active sites. Thus, in general, although proximity effects may contribute significantly to catalysis for some enzymes the effect is not of dominant importance.It is clear from the examples given that a complete mechanistic description of an enzyme-catalysed reaction provides information about the types of catalytic processes involved but does not answer the question as to why the catalysis is so great compared with that observed in non-enzymic model systems. The problem is to explain why the mutual reactivity of a particular set of substrates and catalytic groups is so much greater on the enzyme surface than in free solution.For example, in enzymic transamination (Fig. 4) why are the groups -AH and -B so effective in donating and removing protons from the bound substrate in the environment provided by the active site of the enzyme. Similarly, in the case of catalysis by chymotrypsin (equation 6) an explanation must be provided for the enormous increase in nucleo- philicity of the serine hydroxyl group in the enzyme-substrate complex compared with the nucleophilicity of hydroxyl groups in free solution; in addition the possibility that the carbonyl group of the substrate is activated in the enzyme-substrate complex must be considered. Recent evidence2 suggests that the fundamental factor in enzyme catalysis is the physical environment of the active site and the effects which a particular environment produces on the properties of catalytic groups and substrates in the active site.Two main ways in which environmental effects could modify the behaviour of substrates and catalytic groups may be proposed, both of which have analogies in physical organic chemistry; these are strain and solvent effects. The strain effect1928 arises from attractions (ionic, non-polar and hydrogen bonding) between the substrate and protein side chains which physically distort the substrate molecule, thus enhancing its reactivity. That large rate enhancements can arise from such physical distortions is well known. For example, Haake and Westheimerzg have shown that the base catalysed hydro- lysis of the monoanion of ethylene phosphate (equation 9) proceeds 107 times Doonan 129 C H I - 0 CH20PO:- 0 (9) +OH--- 0- CH2- 0 CH,OH (10) , P , f OH-- 1 CH3-0 ' 4 0 C H 3 - 0 I CH30POi-+CH30H CH ,Cl + I- (1 1) (CH3)zNCHO ' + ,P, 0- faster than the hydrolysis of the corresponding anion of dimethylphosphate (equation 10); this rate difference was attributed solely to bond strain in the 5-membered ring system of ethylene phosphate.Allowing for differences in position of bond cleavage, a rate difference of at least 108 was calculated for hydrolysis of the P-0 bond in the two cases. It is easy to see how such an effect could contribute to enzyme catalysis and its possible contribution to catalysis by the enzyme lysozyme will be discussed later.The second environmental effect, and the one which may prove to be by far the most important factor in enzyme catalysis, depends on the ability of enzymes to exclude water from their active sites on formation of enzyme- substrate complexes. Thus the catalytic groups and bound substrates are removed from the bulk solvent (water) and transferred to an environment which is a function of the particular amino-acid side-chains forming the active site region; this environment can then be 'tailor-made' to suit the requirements of the particular reaction. Again, recourse to physical organic chemistry provides examples of the profound rate changes which frequently accompany changes in solvent systems.Water, and other hydrogen-bonding solvents, are poor media for many organic reactions due to their ability to solvate ionic species. For example, an anion in aqueous solution is strongly solvated so that the negative charge is effectively distributed over the ion and solvation shell ; thus its nucleophilicity is drastically reduced. In dipolar aprotic solvents, however, where solvation is much lower, an anion is a much stronger nucleophile.30 The S N ~ reaction shown in equation 11 has been studied in a variety of solvents31 and some relative rate data are given in Table 3. CH, I +CI-- Table 3. Relative rates for reaction I I at 25"31 CH3NHCHO NHzCHO CH30H Solvent Relative rate 12.5 I 54.3 1.2 x 106 A change from methanol to the dipolar aprotic solvent dimethyl formamide results in a rate increase of lo6 fold.Formamide and methyl formamide, which are relatively good hydrogen bonding solvents, do not cause a com- parable increase in rate. Similarly, anions in dipolar aprotic solvents act as much stronger bases than the same anions in polar hydrogen bonding solvents. For example, the R.I.C. Reviews 130 methoxide ion catalysed racemization of (+)-2-methyl-3-phenylpropionitrile is 109 times faster in dimethyl sulphoxide as solvent than the same reaction in methanol;32 since the rate limiting step in this reaction is the removal of a proton from the asymmetric carbon atom, this rate difference reflects the increased basicity of the methoxide ion in dimethyl sulphoxide as solvent.An even larger effect has been observed in the racemization of 1-phenylmethoxy- ethanol by t-butoxide ions in dimethyl sulphoxide which proceeds 1012 times faster than the reaction in methanol catalysed by methoxide ions.33 From this very small selection of examples of the effect of solvent changes on reaction rates (further examples are discussed in, for example, reference 30) it can be seen that the rate enhancements observed are of the same order of magnitude as the catalytic effect of enzymes. Hence, given the ability of an enzyme to exclude water from the active site and thus effectively to change the solvent in which the reaction is taking place, it is reasonable to propose that environmental effects are largely, if not entirely, responsible for enzyme catalysis. Lysozyrne Lysozyme catalyses the hydrolysis of /3(1 + 4) glycosidic linkages in poly- saccharides of the type shown in Fig.5. The natural substrate is a poly- saccharide, present in the cell walls of certain bacteria, which consists of alternating units of N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG) ;42 hydrolysis occurs exclusively between NAM and NAG residues and not between NAG and NAM. The enzyme also catalyses the hydrolysis CH2OH CH2 OH CHIOH NHCOCH, NHCOCH, NHCOCH, STUDIES OF INDIVIDUAL ENZYMES Many studies of the mechanism of action of enzymes have been reported,34 but in only a small number of cases is information pertinent to a description of catalysis available.The reason for this is that the information required can only be derived from a knowledge of the complete three-dimensional struc- tures of enzymes. Such structure determinations, which, as has already been pointed out, present very formidable problems in the application of x-ray crystallography, have so far been reported for a rather small number of enzymes (lysozyme35J6 ribonuclease,37$3* chymotrypsin,loJg carboxypepti- dase40 and papain41). Of these, lysozyme, chymotrypsin and carboxy- peptidase are perhaps best understood and these three examples will be used to illustrate the general considerations of the previous section. CH2OH CH20H NHCOCH, NHCOCH, R = - H (N- acetylglucosamine) ,CH3 or R=-CH, (N - acetylrnuramic acid) COOH Fig.5. Substrates for lysozyme. 131 Doonan x1-x2 dimer 0.003 X1-Xz-X3 trimer I 4 4 X1-Xz-X3-X4 X1-Xz-X3-X4-X5 Table 4. Hydrolysis of oligosaccharides of NAG by lysozyme44 Main cleavage points J. I Relative rare 8 4000 30 000 Saccharide tetramer pentamer hexamer C 9 ? $ 4 xl-x2-x3-x4-X5-x6 - C of chitin,43 the polysaccharide in which only NAG occurs. Oligosaccharides containing two to four residues of NAG are hydrolysed very slowly, the pentamer much more rapidly and the enzyme is maximally active with the hexamer ; with the pentamer and hexamer, cleavage occurs principally between residues four and five (Table 4).44 Phillips and his coworkers35J6 have determined the complete three-dimen- sional structure of lysozyme and also that of a complex between lysozyme and the inhibitor (or rather, very poor substrate), tri-NAG.From these studies, the active site of the enzyme has been located and a plausible hypothesis for the mechanism of action and catalytic effect of the enzyme has been advanced. Lysozyme is a compact molecule with a pronounced cleft running along one side; in the complex between the enzyme and tri-NAG, the trimer is bound into this cleft (shown diagramatically in Fig. 6a) with the reducing end near the centre. Since the trimer inhibits the binding of hexa-NAG to the enzyme, Phillips and his coworkers assumed that it occupied part of the binding site of the hexamer and attempted to fit further NAG residues into their model of the enzyme-tri-NAG complex.It was found that three more residues could be fitted (Fig. 6b) provided that residue D was distorted some- what from the normal chair conformation. Assuming that the model repre- sented the Michaelis complex between lysozyme and hexa-NAG, it was possible to locate the catalytic groups in the active site. Rupley's work on the hydrolysis of oligomers of NAG suggested that cleavage occurred between residues D and E. This postulate was strongly supported by attempts to build asp-52 / 0- \O .1 ? C ? 6 F f I t b) Binding of tri - NAG Binding of hexa - NAG Arrangement of catalytic groups Fig. 6. Diagrammatic representation of the substrate binding site of lysozyme. 132 R. I . C. Reviews a NAM-NAG polymer into the active site.It was found that NAM residues could not occupy positions C and E because of interactions between the lactyl side chain on C3 of the hexose ring and amino-acid side-chains; the hexamer could only be fitted if it had the sequence NAG. NAM .NAG. NAM . NAG. NAM. From the known specificity of the enzyme, possible sites of hydrolysis were therefore limited to the links between residues B and C or D and E. Since in the unreactive complex with tri-NAG sites A, B and C are occupied, it seems that hydrolysis must occur between residues D and E. Moreover, two likely catalytic groups, namely the carboxyl side chains of residues glutamic-35 and aspartic-52, are located on either side of the linkage between residues D and E (Fig.6c). Close examination of the lysozyme-hexa-NAG model has yielded informa- tion about the mechanism and catalysis of the hydrolytic reaction. Of greatest significance are the micro-environments of the catalytic side chains of aspartic- 52 and glutamic-35. The former is located in a polar region of the enzyme whereas the environment of glutamic-35 is essentially non-polar ; under the conditions of pH at which lysozyme is active, aspartic-52 will exist as the carboxylate ion whilst glutamic-35 will be in its uncharged protonated form (Fig. 6c). The carboxyl group of glutamic-35 is situated about 0.3 nm from CH3CONH I CH3CONH C 0'" \ asp Fig. 7. The mechanism of action of lysozyme (continued ovedeof). 133 Doonan 10 CHlOH I 0- CH20H A - B - C CH3CONH I CH,CONH C 04 \ - a* P 0-H A - B-C Go :- I CH,CONH I 0GC\ asp CH,CONH C i 04 \ the glycosidic oxygen atom between residues D and E and is, therefore, ideally positioned to participate in acid catalysis of the hydrolytic reaction.On the basis of the information given above, and the established mechanisms of non-enzymic hydrolysis of glycosides, Vernon45 has proposed the scheme shown in Fig. 7 for the enzyme-catalysed hydrolysis of hexa-NAG. Glutamic-35 protonates the glycosidic oxygen atom, after which hetero- lysis of the bond between the protonated oxygen and C1 of residue D occurs with the generation of a carbonium ion. The dimer EF leaves the active site and is replaced by a water molecule. The water molecule reacts with the carbonium ion to generate the product tetra-NAG which then diffuses away from the enzyme.Arguments in support of this mechanism rather than other 134 R. I. C. Reviews possibilities involving bimolecular cleavage of the glycosidic bond or intra- molecular participation by the acetamido group on C2 of residue D have been discussed .2 Given the mechanism in Fig. 7, it remains to explain the rate enhancement produced by the enzyme, the estimated magnitude of which is 1010 fold or greater.46 Of the factors which may contribute to catalysis, that arising from protonation of the glycosidic oxygen atom by glutamic-35 is the most difficult to assess. The catalytic factor from this source will depend on the proton donating and accepting properties of the acidic group and glycosidic oxygen atom respectively in the particular environment provided by the enzyme; there is, of course, no way of measuring this.However, the acid strength of the glutamic residue in its essentially non-polar environment would be expected to be low, and consequently acid catalysis is not likely to contribute a large rate enhancement factor to the overall reaction. Similarly, the distortion of residue D which occurs on binding of the substrate to the enzyme surface is a contributing, but not a dominant, factor in catalysis. Model building studies suggest that substrate binding is accompanied by distortion of the ring of residue D towards the so-called half-chair conformation in which atoms C1, C2, C4 and the ring oxygen are coplanar; this is the preferred conforma- tion of carbonium ions derived from pyranosyl ring systems since it enables the positive charge to be distributed over both C1 and the ring oxygen atom.47 Distortion of the substrate on binding lowers the energy barrier between the enzyme-substrate complex and the transition state of the reaction, thus increasing the reaction rate.It has been argued,2 however, that the rate enhancement due to ring distortion is unlikely to be more than 104 fold, which still leaves the major part of the catalysis to be explained. The feature of the active site of lysozyme which is responsible for the major part of the catalytic activity of the enzyme is the negative charge on aspartic-52, the side chain oxygen atoms of which approach to within about 0.3 nm of C1 and the ring oxygen atom of residue D.It seems that the energy required for formation of the carbonium ion intermediate, the rate limiting step in the reaction sequence, is provided by the electrostatic interaction between the negative charge on aspartic-52 and the developing positive charge on the carbonium ion. It can be shown that the energy of interaction between two point charges separated by 0.3 nm in vacuo is about 5 eV*,45 whereas the difference in free energy of activation required to produce the estimated rate increase in the enzyme catalysed reaction is only about 0.5 eV; obviously on this simple model the electrostatic energy is more than enough to account for catalysis by the enzyme.The point-charge model overestimates the energy of interaction since both of the charges will be dispersed to some extent-that on aspartic-52 over both carboxyl oxygen atoms and that of the carbonium ion over C1 and the ring oxygen-and in addition the dielectric constant of the environment of the charges will be greater than unity. Even allowing for these effects, the energy involved is certainly large enough to account for the catalytic effect of the enzyme. A requirement for this catalytic effect to operate is the exclusion of bulk solvent (water) from the active site in the enzyme-substrate complex: * 1 eV is approximately equal to 1.602 x 10-19J. 135 Doonan water, by virtue of its high dielectric constant, would reduce the electrostatic interaction to a value less than 0.1 eV thus providing negligible rate enhance- ment.In summary, the major part of the catalytic activity of lysozyme derives from the energy of interaction of a negative charge in the enzyme and a developing positive charge on the substrate, the function of the enzyme being to provide a physical environment in which these charges will form readily and interact strongly. Ch ymo t rypsiiz Chymotrypsin is an enzyme which catalyses the hydrolysis of a variety of esters and amides (equation 5). The mechanism of action of the enzyme has been the subject of extensive investigation~~~9~~ and was in large part under- stood before the three-dimensional structure was determined from x-ray diffraction studieslOJ9 (see p.122); these studies have, however, led to an understanding of the catalytic properties of the enzyme. Recognition of the importance of a serine residue for the activity of the enzyme arose from the work of Balls and J a n ~ e n ~ ~ who showed that the enzyme is completely inhibited by treatment with diisopropylphosphofluoridate (DFP) in stoicheiometric amounts. Subsequent work50 showed that DFP reacts with a unique serine residue (serine-195 in the sequence published by Hartleys) and with none of the other 28 serine residues in the enzyme. These observations were taken to show that serine-195 is involved in the activity of the enzyme. It has now been established with some certainty that for sub- strates other than esters or amides of aromatic amino acids, hydrolysis occurs by the so-called double displacement reaction sequence (equation 12) ;22948 that is, the hydroxyl group of serine-195 (shown as E.OH in equation 12) H O E.OH -k RCOX (12) + HX E.OCOR 2 E.OH + RCOOH displaces group -X from the substrate and is itself acylated, after which hydrolysis of the acyl enzyme intermediate liberates the free enzyme and carboxylic acid.Although the evidence that reaction sequence 12 is applicable to the hydrolysis of esters and amides of aromatic amino acids is less com- plete,2 it will be assumed for the purposes of the present discussion that equation 12 is a general representation of chymotrypsin catalysed reactions. The second reactive amino-acid side-chain to be implicated in chymo- trypsin activity was the imidazole ring of histidine-57.Studies of the depen- dence of reaction rate on pH showed that activity was controlled by a group with pKa of about 7 and it was assumed that this group was the side chain of a histidine residue.51 Further, Schoellmann and Shaw52 showed that inhibi- tion of the enzyme by the substrate analogue tosyl-L-phenylalanylchloro- methane resulted from reaction of the reagent with an imidazole side chain, and Smillie and H a r t l e ~ ~ ~ identified this as the side chain of histidine-57. On the basis of these observations, a possible reaction mechanism may be written as in equations 13 and 14, where -OH and >N: represent the hydroxyl group of serine-195 and one of the ring nitrogen atoms of histidine- 57 respectively.R.I.C. Reviews 136 The sequence of reactions in equations 13 and 14 does not, of course, account for catalysis by the enzyme. The rate limiting steps in the sequence are presumably the nucleophilic attacks by serine on the substrate in 13 and by water on the acyl enzyme intermediate in 14; hence catalysis must arise from some factor which increases the nucleophilicity of these species. It was pointed out earlier (p. 130) that the most effective nucleophiles are unsolvated anions; hence it is reasonable to assume that the enzyme functions by pro- ducing the alkoxide ion of serine-195 (in reaction 13) and a hydroxide ion (in reaction 14) under conditions where they are essentially unsolvated, and that the role of histidine-57 is to assist in the deprotonation reaction producing these ions.The central problem is to explain how the base strength of histi- dine-57, in the particular environment of the enzyme active site, is increased to the extent that it can effectively deprotonate the serine residue or a water molecule. The solution to this problem has come from the x-ray diffraction studies carried out by Blow and his colleagues.l0J9 They found that residues histi- dine-57 and serine-195 are properly positioned to interact in the active site, but, in addition, histidine-57 is in close contact with the carboxyl side chain of aspartic-102,5* a residue which had not previously been implicated in the activity of the enzyme.These three residues are arranged in the enzyme in such a way that a pair of protons can interact with four binding sites (namely the carboxylic oxygen, two nitrogen atoms of the imidazole ring and the serine oxygen) to produce a system with the extreme forms shown in Fig. 8; his his v Fig. 8. Charge relay system at the active site of a-chymotrypsin. Doonan 137 these species may be considered either as tautomers in equilibrium or as canonical forms of a resonance structure depending on whether proton movements occur. If the form on the right of Fig. 8 predominates, then the serine oxygen atom will carry a large fraction of a negative charge; that is, the coupling of aspartic-102 and histidine-57 will increase the base strength of the histidine residue sufficiently to deprotonate serine-195.In fact, aspartic- 102 is situated in a non-polar environment in which its ionization will be suppressed and hence the form of the aspartic-histidine-serine system in which aspartic-102 is uncharged and serine-102 exists as the alkoxide ion will predominate. Thus the particular arrangement of aspartic-102 and histidine-57 constitutes a device for activating serine-195 in reaction 13 and, by analogy, the water molecule involved in the hydrolysis of the acyl enzyme inter- mediate (reaction 14). Generation of a negative charge on swine-195 is a necessary but not a sufficient condition for catalysis of reaction 13. The second requirement is that binding of the substrate to the active site displaces water from the region of serine-195, thereby desolvating the partial negative charge formed by inter- action with histidine-57.The fact that serine-195 exhibits high nucleophilicity only towards substrates and substrate analogues which bind at the active site supports the idea that binding promotes desolvation of the active serine residue. The reactivity of the desolvated alkoxide ion should then be great enough to account for the catalytic effect. I (15) I Rn- I Carboxypeptidase A The mechanism of action and catalytic features of carboxypeptidase A are not so well understood as are those of lysozyme and chymotrypsin, but recent x-ray diffraction studies40 have revealed interesting features of the interaction between the enzyme and its substrate.The active site of an enzyme is fre- quently pictured as a rigid region of the enzyme surface in which the catalytic groups are correctly orientated to react with the substrate once the latter has become attached to the appropriate binding sites. It was suggested some time ago, however, that in some cases the catalytic groups may be forced into the correct orientation as a result of substrate binding; that is, the active site should be considered as a flexible structure which can be moulded into the correct 'shape' by the act of binding the substrate.55~56 Evidence that this effect plays an important part in the mechanism of action of carboxypeptidase A has been 0btained.~0 Carboxypeptidase-A catalyses the hydrolysis of the C-terminal peptide bond of peptides and proteins (reaction 15).The enzyme contains one gram I . . . CHCONHCHCOOH + H,O - . . .CHCOOH + NHzCHCOOH I Rn-1 R" R n atom of zinc per mole of protein, and it has been shown that the zinc ion is essential for catalytic a~tivity.5~ Similarly, a tyrosine residue has been impli- cated in the activity of the en~yrne.5~959 Lipscomb and his coworkers40 have described the events which occur on binding of a poor substrate, glycyl-L-tyrosine, to the active site of the enzyme R.I.C. Reviews 138 \ \ I I I I * I I I OH / I ____/ - / NH,CH,C Go NHCHCOO- I I1 /I\ OH - _ I , active enzyme - substrate complex Fig. 9. Substrate induced conformational changes in carboxypeptidase-A. inactive enzyme- substrate complex (Fig.9). The tyrosine residue of the substrate fits into a 'pocket' in the enzyme surface and is held in position by interaction between the carbonyl oxygen atom and the protein-bound zinc ion. An interaction occurs between the negatively charged carboxyl group on the substrate and a positively charged arginine residue in the protein (arginine- 145) which causes the latter residue to move by about 0.2 nm towards the substrate. This small movement has a profound effect on other regions of the enzyme chain, and in particular it causes the hydroxyl group of residue tyrosine-248 to move by about 1.20 nm from a position pointing into the solvent shell of the enzyme to a new position directly above the peptide bond of the substrate (Fig.9). This appears to be a direct demonstration of Koshland's induced fit the0ry.~~?56 These induced conformational changes also explain the observation that carboxypeptidase-A does not catalyse the hydrolysis of peptides in which the terminal carboxyl group is amidated even though such amidated peptides bind readily to the enzyme. Amidated peptides lack a negative charge on the terminal carboxyl group and hence cannot initiate the conformational changes which lead to the catalytically active conformation of the active site. The events which occur after formation of the active enzyme-substrate complex are a matter of speculation. It seems likely that the function of tyrosine-248 is to protonate the peptide nitrogen atom thus facilitating either unimolecular or bimolecular cleavage of the peptide bond ; polarization of the peptide carbonyl group by binding to the zinc ion will also assist in either mode of bond cleavage.Crystallographic studies4() have shown that the carboxyl group of residue glutamic-270 is orientated directly towards the peptide bond of the substrate. This group could participate in the reaction by nucleophilic attack on the carbonyl group to produce a mixed anhydride, subsequent hydrolysis of which would yield the desired product (Fig. 10). Alternatively, glutamic-270 may participate by stabilization of an inter- mediate carbonium ion formed by unimolecular cleavage of the peptide bond ; the situation would then be very similar to that found with lysozyme.Doonan 139 I I / I :H-COO- - 1 Rn I Rn-1 O Rn-1 0 I I --L - CH - c -OH NH~CHCOO- I1 Rrl I I Zn2+ Zn2+ A Fig. 10. A possible reaction mechanism for carboxypeptidase-A. Whichever of these two mechanisms is operative, the general requirements for catalysis seem clear. Firstly, the environments of the tyrosine hydroxyl group and the peptide nitrogen atom must be such that proton transfer from one to the other is essentially complete. Secondly, the carboxyl group of glutamic-270 must be shielded from solvent water so that the effectiveness of the group either as a nucleophile or in stabilizing a carbonium ion by electro- static interactions is maximized. A precise description of these catalytic factors must await more detailed information about the active site of the enzyme.CONCLUSIONS The central theme of this article has been that a description of catalytic factors requires more information than is included in the most detailed statement of the mechanism of an enzyme-catalysed reaction. In particular, information is required about the physical environments of catalytic groups in the active site and the way in which environmental factors change the chemical behaviour of these groups. This sort of information is only available at the present time for a very small number of enzymes, but it may reasonably be hoped that the principles outlined above to explain catalysis by lysozyme, chymotrypsin and carboxypeptidase will be found to be of much more general application.R. I . C. Reviews 140 5 P. Edman, Proc. R . Anst. chem. Inst., 1957, 434; Ann. N. Y. Acad. Sci., 1960, 88, 602. ACKNOWLEDGMENTS Figure 2 is reproduced by courtesy of Dr D. M. Blow and the editor of Nature; Figure 3 by courtesy of Dr Blow and the Medical Research Council. REFERENCES 1 R. Lumry, in The enzymes (ed. P. D. Boyer, H. Lardy and K. Myrback), vol. 1, chapter 4. New York: Academic Press, 1959. 2 S. Doonan, C. A. Vernon and B. E. C. Banks, Prog. Biophys. molec. Biol., 1969, in press. 3 D. E. Koshland and IS. E. Neet, A. Rev. Biochem., 1968,37, 359. 4 C. H. W. Hirs (ed.), Methods in enzymology. New York: Academic Press, 1967. 6 R. Shipolini, A. F. Bradbury, G. L. Callewaert and C. A. Vernon, Chem. Commun., 7 G.L. Callewaert and R. Shipolini, unpublished work. 8 B. S. Hartley, in Structure and activity of enzymes, (ed. T. W. Goodwin, J. I. Harris and B. S. Hartley), p. 47. London: Academic Press, 1964. 9 B. Keil and F. Sbrm, in Structure and activity of enzymes (ed. T. W. Goodwin, 652. 1967, 679. J. I. Harris and B. S. Hartley), p. 37. London: Academic Press, 1964. 10 B. W. Matthews, P. B. Sigler, R. Henderson and D. M. Blow, Nature, Lond., 1967, 214, 11 J. C. Kendrew, H. C. Watson, B. E. Strandberg, R. E. Dickerson, D. C. Phillips and 5, 528. V. C. Shore, Nature, Lond., 1961, 190, 666. 12 L. Stryer, A . Rev. Biochem., 1968, 37, 25. 13 D. C. Phillips, Prog. Biophys. molec. Biol., 1969, in press. 14 B. E. C. Banks, S. Doonan, A. J. Lawrence and C.A. Vernon, Eur. J . Biochem., 1968, 15 B. E. C. Banks, S. Doonan, J. Gauldie, A. J. Lawrence and C. A. Vernon, Eur. J. 16 B. E. C. Banks, M. P. Bell, A. J. Lawrence and C. A. Vernon, in Pyridoxal catalysis: enzymes and model systems (ed. E. E. Snell, A. E. Braunstein, E. S. Severin and Yu. M. Torchinsky), p. 191. New York: Interscience, 1968. Biochem., 1968, 6, 507. 17 B. E. C. Banks, A. J. Lawrence and C. A. Vernon, Eur. J. Biochem., 1969, in press. 18 S . F. Velick and J. Vavra, in The enzymes (ed. P. D. Boyer, H. Lardy and K. Myrback), vol. 6, chapter 15. New York: Academic Press, 1962. 19 D. E. Metzler, M. Ikawa and E. E. Snell, J. Am. chem. Soc., 1954, 76, 648. 20 J. Kirsch and W. P. Jencks, J. Am. chem. SOC., 1964, 86, 837.21 C. G. Swain and J. F. Brown, J. Am. chem. SOC., 1952, 74, 2538. 22 M. L. Bender and F. J. Kkzdy, A. Rev. Biochem., 1965, 34,49. 23 M. L. Bender, J. Am. chem. Soc., 1957, 79, 1258. 24 B. Capon, Q. Rev. chem. SOC., 1964, 18,45. 25 T. C. Bruice, J. Am. chem. SOC., 1959, 81, 5444. 26 D. E. Koshland, J. theor. Biol., 1962, 2, 75. 27 D. E. Koshland, J. cell. comp. Physioi., 1956, 47 suppl. 1, 217. 28 W. P. Jencks, in Current aspects of biochemical energetics (ed. N. 0. Kaplan and E. P. Kennedy), p. 273. New York: Academic Press, 1966. 29 P. C. Haake and F. H. Westheimer, J. Am. chern. SOC., 1961, 83, 1102. 83, 3678. 30 A. J. Parker, Q. Rev. chem. SOC., 1962, 16, 163; Chem. Rev., 1969, 69, 1. 31 A. J. Parker, J. chem. Soc., 1961, 1328. 32 D. J. Cram, B. Rickborn, C. A. Kingsbury and P. Haberfield, J. Am. chem. Soc., 1961, 33 D. J. Cram, C. A. Kingsbury and B. Rickburn, J. Am. chem. SOC., 1961, 83, 3688. 34 P. D. Boyer, H. Lardy and K. Myrback (eds), The enzymes, vols 1-8. New York: Academic Press, 1959-63. 35 C. C. F. Blake, G. A. Mair, A. C. T. North, D. C. Phillips and V. R. Sarma, Proc. R. 36 C. C. F. Blake, L. N. Johnson, G. A. Mair, A. C. T. North, D. C. Philips and SOC., 1967, B167, 365. 37 G. Kartha, J. Bello and D. Harker, Nature, Lond., 1967, 213, 862. V. R. Sarma, Proc. R . SOC., 1967, B167, 378. 38 H. W. Wyckoff, K. D. Hardman, N. M. Allewell, J. Inagami, D. Tsernoglou, L. N. Johnson and F. M. Richards, J. b i d . Chem., 1967, 242, 3749. 39 P. B. Sigler, D. M. Blow, B. W. Matthews and R. Henderson, J. mol. Biol., 1968, 35, 143. Doonan 141 40 G. N. Reeke, J. A. Hartsuck, M. L. Ludwig, F. A. Quiocho, T. A. Steitz and W. N. Lipscomb, Proc. natn. Acad. Sci. U.S.A., 1967, 58, 2220. 41 J. Drenth, J. N. Jansonius, R. Koekoek, H. M. Swen and B. G. Wolthers, Nature, Lond., 1968, 218, 929. 42 M. R. J. Salton and J. M. Ghuysen, Biochim. biophys. Acta, 1959,36, 552; 1960,45, 355. 43 L. R. Berger and R. S . Weiser, Biochim. biophys. Acta, 1957, 26, 517. 44 J. A. Rupley, Proc. R. SOC., 1967, B167, 416. 45 C. A. Vernon, Proc. R. Soc., 1967, B167, 389. 46 L. N. Johnson, D. C. Phillips and J. A. Rupley, Brookhaven Symp. Biol., 1968, in press. 47 R. V. Lemieux and G . Haber, Can. J. Res., 1955, 33, 128. 48 T. C. Bruice and S. J. Benkovic, Bioorganic mechanisms, vol. 1, chapter 2. New York : Benjamin, 1966. 49 A. K. Balls and E. F. Jansen, Adv. Enzymol., 1952, 13, 321. 50 R. A. Oosterbaan, M. van Adrichem and J. A. Cohen, Biochim. biophys. Acta, 1962, 63, 204. 51 D. E. Koshland, D. H. Strumeyer and W. J. Ray, Brookhaven Symp. Biol., 1962,15, 101. 52 G. Schoellmann and E. Shaw, Biochemistry, 1963, 2, 252. 53 L. B. Smillie and B. S. Hartley, Fed. Eur. Biochem. Socs, 1964, abstract A 30. 54 D. M. Blow, J. J. Birktoft and B. S . Hartley, Nature, Lond., 1969, 221, 337. 55 D. E. Koshland, Proc. natn. Acad. Sci. U.S.A., 1958, 44, 98. 56 D. E. Koshland, J. cell. comp. Physiol., 1959, 54, 235. 57 B. L. Vallee, J. A. Rupley, T. L. Coombs and H. Neurath, J. biol. Chem., 1960,64,235. 58 0. A. Roholt and D. Pressman, Proc. natn. Acad. Sci. U.S.A., 1967, 58, 280. 59 R. T. Simpson, J. F. Riordan and B. L. Vallee, Biochemistry, 1963, 2, 616. 142 R. I.C. Reviews

ISSN:0035-8940    

DOI:10.1039/RR9690200117

出版商:RSC    

年代:1969

数据来源: RSC

摘要:

The Seventh P. F. Frankland Memorial Lecture CHEMISTRY AND NUTRITION The Late Alastair Frazer, C.B.E., M.D., Ph.D., D.Sc., F.I.Biol., F.R.C.P. British Nutrition Foundation. London SE I . . . . .. . . . . . . . . . . 144 . . . . . . . . . . .. . . 144 What is food? What does food do? Some factors responsible for the development of modern methods of food production and distribution . . . . . . . . . . 145 . . . . . . . . . . . . . . . . 146 Population growth, 145 Increased urbanization, 145 Changed ways of living, 146 Raw materials Main nutrient constituents, 147 Natural toxicants, 149 Residues of chemical aids used in agriculture, 150 The role of the chemist in relation to raw materials, 152 The development of foods from raw materials .. . . . . . . 153 Modification of nutritional value of a food, 153 Formation of potentially deleterious toxic substances in food, 154 Safety evaluation of food additives, 156 Contaminants introduced into foods, 157 Packaging materials, 157 The role of the chemist in relation to food production, 158 Effects on the consumer . . . . . . . . . . . . . . 158 Availability of nutrients, 158 Metabolism in foods, 159 Genetic factors, 159 Pathology and drug therapy, 160 The role of the chemist in the study of the consumer, 161 Conclusion . . . . . . . . . . . . . . . . . . 161 References . . . . . . . . . . . . . . . . . . 162 The seventh P. F. Frankland Memorial Lecture was delivered at the University of Birmingham on 8 November 1968 by Dr Alastair Campbell Frazer.Dr Frazer was born in London in 1909 and was educated at Lancing and S t Mary’s Hospital Medical School. He lectured in physiology and pharmacology at the latter institu- tion from 1929 until 1941. From 1943 until 1967 he was Professor of Medical Biochemistry and Pharmacology in the University of Birmingham. On leaving Birmingham, he became Director-General of the British Nutrition Foundation, a post he held until his death in June 1969. In addition, he was a consultant t o the UKAEA, president of both BFMIRA and BIBRA, scientific adviser t o the Ministry of Agriculture, Fisheries and Food, and Chairman of the Food Section of IUPAC. Frazer 143 I greatly appreciate the honour of being invited to give the seventh Frankland Memorial Lecture, since it enables me to follow in the footsteps of a number of friends and colleagues whom I greatly respect and admire.It also gives me particular pleasure because for nearly 25 years, from 1943 until 1967, I occupied an established Chair in this University in which Frankland worked. Percy Faraday Frankland was Professor of Chemistry first at Mason College and then in the University of Birmingham from 1894 until 1918, and during that time he built up a world reputation for his Department, which has been further enhanced by those who followed him. He was a remarkable man, whose characteristics and personality were fully and affectionately described by the late Dr Leslie Lampitt in the first lecture in this series in 1949.1 Birmingham owes much to Frankland.I owe a great deal to Birmingham, which gave me the opportunity of developing some new ideas in biochemical education and research. In the development of the Department of Medical Biochemistry and Pharmacology, I had the active help and co- operation of the successors of Professor Frankland-first the late Professor Sir Norman Haworth and subsequently Professor Maurice Stacey. They provided me with a series of first-class chemists without whose help the Honour School of Medical Biochemistry and the graduate training facilities in my department would never have been created. I have no doubt that Frankland himself would have appreciated the need for more adequate development of chemistry and biochemistry in relation to medicine, especially in such fields as pharmacology, toxicology, nutrition and metabolic diseases, with which I have been associated.They have much in common, especially as regards the background sciences needed for their development. Here I will deal only with chemistry and nutrition, although, as will become apparent as we proceed, this subject cannot be entirely divorced from some aspects of pharmacology, toxicology and metabolic diseases. WHAT IS FOOD? This seemingly simple question calls for careful consideration and analysis. Contrary to the impression given by naturists, the great majority of foods are not consumed in a form that nature provides. Animal or vegetable raw materials require slaughter or harvesting, respectively, followed by various procedures which lead to the development of food.Furthermore, a variable time interval is needed to ensure that the food is in an acceptable state. Freshness may be an important feature in a vegetable, such as a lettuce, but it often does not enhance the palatability of meat, game, cheese or wine, which improves with the passage of time. Thus, food consists of substances derived from raw materials by the application of various procedures over an appro- priate time interval. Most food is unstable and it will only remain acceptable for a limited time period; if it is eaten too soon, it may be unpleasantly immature, but, if too late, it may be found to have deteriorated. WHAT DOES FOOD DO? Food has many effects on the body.Most of them are beneficial and necessary, but some may be unwanted or even harmful. The detailed study of these various effects and their definition in chemical, physical or biochemical R.I.C. Reviews 144 terms forms an important part of the subject matter of nutrition. It is not sensible to separate food chemistry from nutrition; both fields are inter- dependent and should be integrated as closely as possible. A substance is only a food when it is eaten and assimilated; the definition of the properties of a food is inadequate if it does not include the effects the food might have on the consumer. I propose to consider the role of chemistry in the field of food and nutrition in three steps : first, in relation to the raw materials from which food may be produced; second, in connection with the ways in which foods are developed and, third, with reference to the possible influence of the characteristics of the consumer on the effects that food may have on the body.Before doing this we must briefly consider the factors that have changed food production and distribution from a simple cottage handicraft into a major undertaking that calls for the full collaboration of science, industry and government. Three main factors have been responsible for this change: population growth, increased urbanization, and alteration in ways of living. SOME FACTORS RESPONSIBLE FOR THE DEVELOPMENT OF MODERN METHODS OF FOOD PRODUCTION AND DISTRIBUTION Population growth The world’s population has doubled in the last 40 years and it is likely to double again by the end of the century unless some check is applied.Not more than half of the present world population is adequately fed; by the year 2000, the proportion is likely to be much less than this, unless steps are taken to improve matters. Food production today is not sufficient to overcome the wastage and faulty distribution of food supplies. It is important that every- thing possible should be done to increase yields and food production generally, to reduce wastage, to improve distribution and storage, and to make better use of land. Much can be done by improved methods of husbandry, but it is also necessary to make full use of scientific advances, such as the use of fertilizers, pesticides and other agricultural aids, and to develop new methods of large-scale food manufacture and better methods of packaging, storage and distribution.While many of these advances will help to improve the world food situation, they may also raise problems on their own account which have to be taken into consideration. Increased urbanization A time there was, ere England’s griefs began, When every rood of ground maintained its man; For him light labour spread her wholesome store, Just gave what life requir’d, but gave no more. So wrote Goldsmith2 in the middle of the 18th century. The flight from the country into the towns has vastly increased since that time. The changes in population in Britain and the percentage that was urbanized between 1801 and 1951 are indicated in Table 1.It has been said that every year between now and the end of the century another urban area the size of Bristol will be created. This trend towards living in towns greatly affects the provision of adequate food supplies. Clearly, the amount of land for food production is progres- Frazer 145 Population (m.) Urban (%) 29 50 75 RAW MATERIALS Table 1. Urbanization in Great Britain I 80 I -I 95 I 10.5 21 37 49 80 Year 1801 1851 1901 1951 sively diminishing. At the same time, there has been a demand that more of our daily food should be produced in this country. If these two lines of action are to be followed, further intensification of farming and food production in Britain is inevitable.This will require the full use of all scientific aids. It is only possible to achieve these various objectives without lowering standards in food and nutrition if there is a continuing and coordinated effort in research and development covering the whole field, a matter discussed in some detail by Sir Gordon Cox in the fifth Frankland Memorial L e ~ t u r e . ~ Changed ways of living Urbanization itself results in a change in the pattern of living. More people tend to eat away from home; more housewives go out to work and have less time for preparing and cooking food; problems of storage and distribution increase. In spite of all these difficulties, there is no doubt that the range and quality of food that is generally available today to the great majority of town- dwellers is enormously superior to anything that was possible in the ‘good old days’ of 100 years However, changes are still occurring.Thus, there is an increased tendency for foods to be separately packaged and held on display either in the supermarket or in deep-freeze cabinets. Because of the lack of time for food preparation and the greater number of old people trying to fend for themselves, there has been a steady increase in so-called ‘convenience foods’. These foods are prepared and possibly cooked, or partly cooked, so that they can be brought to the table much more easily and speedily. A further extension of this approach is the introduction of precooked frozen foods. These foods are cooked centrally by experts and then held in store or distributed frozen.The reheating process can be relatively unskilled, so that such a system opens up possibilities of improving various forms of institutional catering, such as food supplies in large hospitals or schools or social services, such as ‘meals-on-wheels’ for old people. This system may also help to resolve difficulties that arise from a shortage of cooks and the need to cook food throughout the whole day seven days a week. It is important that the pattern of food production and supply should be properly adapted to community needs; these trends must, therefore, be taken into account when considering problems of food and nutrition. The raw materials used for food production may be either of animal or vegetable origin.Materials are chosen that contain useful components, such as carbohydrates, proteins, fats, electrolytes, trace elements and vitamins. R.I.C. Reviews 146 They also contain other constituents that are of more limited value, such as fibrous tissue, bone, lignin, or cellulose. Some of these, however, may have a significant nutritional role in giving acceptable texture to the food. Raw materials may also contain other groups of substances, such as natural toxicants, residues of chemical aids used in agriculture, or natural contami- nants derived from micro-organisms or moulds. All these components have to be given due consideration if the raw material is used for food production. Main nutrient constituents The amount of useful nutrients present in the raw material will vary, due to genetic and environmental factors. Thus, different varieties or strains of cereals will contain different amounts of protein or other nutrients.The same is true of all other plant and animal raw materials. A considerable amount of work is being done on the breeding of new varieties or strains of plant raw materials and of developing better animal stocks. The composition of any particular variety or strain may also vary according to the conditions under which it is grown. Thus, climatic conditions materially affect the protein content of wheat. Since man consumes a mixed diet, variation in the composi- tion of any particular food is not in itself of paramount importance. It is essential to know what the composition is, so that this can be taken into account in the construction of the whole diet.In general, the utilization of the main nutrients by the consumer is well understood; the chemistry of some of them was discussed in the second and third Frankland Memorial lecture^.^^^ I only propose to emphasize that the chemical nature of these nutrients is a matter of major importance. This can be illustrated by reference to two main food components, proteins and fats. In the case of proteins, the nature and the amounts of various con- stituent amino acids have great nutritional significance. Certain amino acids cannot be formed in the body by transamination. These are essential compo- nents of the diet; if one essential amino acid is deficient in a protein its use- fulness is seriously limited.Therefore, in discussing the nutritional significance of any protein it is necessary to consider the amounts of these essential amino acids present. The biological value of a protein can be assessed chemically on the basis of its essential amino-acid content. There are also certain fatty acids that cannot be synthesized in the body. These are polyunsaturated fatty acids with certain characteristics. The most important features of these essential fatty acids are: the presence of two or more double bonds, the separation of the double bonds by a methylene bridge, the spatial arrangement of the double bonds, which should start from the ~6 position, and the presence of cis configuration about the double bonds.7 The presence of polyunsaturated fatty acids of this type in a dietary fat has a significant effect on the blood cholesterol and P-lipoprotein levels of the consumer.* In some animals a definite essential fatty-acid deficiency syndrome can be demonstrated ; it is possible that deficiency may also occur in the young baby,gJO but it has not been adequately defined in adult man.Nevertheless, no-one doubts that these fatty acids play an important part in cell biochemistry in man and that they are and should be present in adequate quantities in the human diet. Frazer 147 All too frequently one still hears reference made to animal and vegetable proteins or fats as an indication of their chemical composition. This differen- tiation is totally inadequate.Some vegetable proteins are the equal of animal proteins in nutritional value; many vegetable fats contain plenty of poly- unsaturated fatty acids, but there are some that consist mainly of saturated fats; similarly, animal fats from ruminant animals contain a high proportion of saturated and little polyunsaturated fatty acid, but other animal fats, for example, pig fat, may contain substantial amounts of polyunsaturated fatty acids. Nutrients should be adequately defined in chemical terms. Table 2. Some possible pharmacological or toxic effects of foods Occurrence Tea, coffee Bananas and some other fruits Some cheeses Soya beans Almonds, cassava and other plants Quail Mussels Cycad nuts Some fish, meat o r cheese Sassifras Legumes Some beans Ackee fruit Brassica seeds and some other vegetables Rhubarb Green potatoes Many fish Many fungi Active agent Caffeine 5-hyd roxytryptam i ne; ad renal i n e ; no rad renal in e Tyramine Oestrogens Cyanide Due t o consumption of hemlock Due t o consumption of dinoflagellate, Gon yaulax Met hylazoxymet hanol N it rosami nes Safrole (p-ally1 methylene dioxybenzene) Haemogl u ti ni ns Vicine P-aminopropionitrile a, y-diaminobutyric acid cyano- I -alanine a-ami no-8-methylene cyclopropane propionic acid Th iooxyazol id ine, t h iocyan ate Oxalate Solanine; possibly other sapotoxins Various, often confined to certain organs o r seasonal Various non-edible fungi Possible erects Cerebral stimulant; diuretic Effects on central and peripheral nervous system Raises blood pressure; enhanced by monoamine oxidase inhibitors Similar to female sex hormones Interferes with tissue respiration Hemlock poisoning Tingling, numbness, muscle weakness, respiratory paralysis Liver damage; cancer Liver damage; cancer Cancer Red cell and intestinal cell damage Haemolytic anaemia Interferes with collagen formation Toxic effects on nervous system H ypogl ycaem ia Enlargement of thyroid gland (goitre) Oxalu ria Gastro-intestinal upset Mainly toxic effects on nervous system Mainly toxic effects on nervous system 148 R.I . C. Reviews Natural toxicants Plants are a well-known source of pharmacologically active substances.It would be surprising, therefore, if some of the plants chosen for food produc- tion did not also contain some pharmacologically active or even toxic principles. Some of the possible effects of a number of such agents are indi- cated in Table 2. It should be emphasized that most of the possible ill effects are avoided by proper choice and preparation of the raw materials. Some of these effects may be wanted; for example, the effect of caffeine in tea and coffee, which helps to allay fatigue.-Some of the more serious effects, such as poisoning by unsuitable fungi, fish, or shellfish, may be avoided by correct choice of edible varieties or restriction of consumption to a safe season.Many of these natural toxicants are of considerable interest chemically. I have chosen three for further comment. First, ackee fruit poisoning: The ackee fruit (Blighia sapida) grows in Jamaica and it is made up into a dish rather like scrambled egg, which is traditionally served with salted cod. It is an excellent dish, but, unfortu- nately, now and again it causes untoward effects in the consumer, such as vomiting, convulsions, and coma, which may prove fatal. These effects are due to a sudden fall in the blood sugar and the signs and symptoms can be dramatically relieved by intravenous injection of glucose. The action of the ackee fruit is due to the presence of a-amino-/l-methylenecyclopropane propionic acid (hypoglycin) ~ H,C=C’I NH3 ‘CH-CH~-CH-COO- I which is a metabolic blocking agent interfering with the storage of liver glycogen and the maintenance of the normal blood sugar level.11-13 Hypo- glycin is found particularly in unripe ackee fruit.A high protein diet and riboflavin afford some protection and the effect most commonly occurs in undernourished children. Coming nearer home, there is green potato poisoning. The normal potato tuber (Solanum tuberosis) contains less than 0.01 per cent of solanine, an alkaloid which has a sapotoxin-like effect, and causes severe gastro-intestinal disturbance. Under certain circumstances the solanine content can increase up to 0.05 per cent or more; this increase is commonly associated with the development of chlorophyll. The chlorophyll is, of course, harmless, but the accompanying solanine is toxic to man at a dosage level of about 200mg.There have been reports of epidemic outbreaks of green potato poisoning from time to time.14J5 A third example of a natural toxicant is a form of musselpoisoning, which occurred in NE England last May.16 Some 80 people were seriously affected, but fortunately none died, although fatalities have occurred in other out- breaks. The poisoning was considered to be due to the consumption of mussels that had been feeding on a dinoflagellate marine organism of the genus Gonyaulax, which gives rise to a neurotoxic agent, saxitoxin. Dino- flagellate organisms may suddenly increase in great numbers in late spring Frazer 149 11 or early summer and when this occurs the sea may appear red or brown- termed by fishermen ‘the red tide’.Such brown masses were reported by fishermen in early May off the east coast. Consumption of poisonous mussels results in paraesthesis and numbness, resembling the effect of a local anaesthetic, and this is followed by inter- ference with muscle movements. In severe cases the respiratory muscles may be paralysed. Cooking the mussels removes about 50 per cent of the toxic agent into the cooking water. This outbreak was limited partly by the fact that most of the mussels were well cooked before eating and partly by very prompt action on the part of the physician who saw the first case and the Medical Officer of Health. This episode illustrates the importance of seasonal restriction of some foods.The old adage ‘Do not eat shellfish if there is no R in the month’ may have a sounder and broader basis than might be expected. Residues of chemical aids used in agriculture In order to increase yields and prevent wastage, various chemical aids are used, some of which were discussed in detail by Professor R. L. Wain in the sixth Frankland Memorial Lecture.17 Their significance in relation to food production can be illustrated by reference to some of the problems concerned with nitrate fertilizers, pesticides, oestrogens and antibiotics. Nitrate fertilizers. It is often helpful to treat the soil with nitrates. These nitrates are converted into protein by plants. Under certain circumstances, however, some of the nitrate added to the soil is washed away in surface water into reservoirs and shallow wells, where it may stimulate the growth of blooming algae, some of which are toxic.The nitrate-containing water itself may also be used for human consumption. The nitrate taken up by the plant may not be completely converted into protein, so that an increased nitrate content may result. The increased use of nitrates as fertilizer may, therefore, lead to an increased intake of nitrate by the consumer either in water or in food. Nitrate is, however, a substance which is formed in the body in the course of metabolism in small amounts and it has a low toxic potential; there seems little likelihood that this increase in nitrate intake would in itself cause any harm. A different situation arises if the nitrate is reduced to nitrite, which is about 10 times more toxic than nitrate and causes quite different effects on the body.Conversion of nitrate to nitrite may occur if food containing an excess of nitrate is allowed to undergo bacterial spoilage, or if the nitrate is exposed to the reducing flora of the intestine. This is avoided by careful choice of raw materials for baby foods, by proper food hygiene, and by restricting nitrate intake in food or water in babies or in others with an active flora in the upper part of the intestinal tract, including ruminant animals. The maximum permitted level of nitrate nitrogen in drinking water for human use varies from one country to another-it is commonly 10-20 ppm.However, this does not allow a great margin of safety.18t19 Pesticide residues. Many pesticides leave no detectable residue, but others, notably the organochlorine pesticides, are more persistent. When these are R. I.C. Reviews 150 DDTIDDE Range 0.00-0.0 I 0.00 I5 Range ( p m ) headlday) (PPm) 0.00 I8 0.00- I .2 0.04-0.20 0.00-0.005 0.00 I 3 0.00-1 0.0 0.0 I 5 0.10- 0.8 0.00067 Corned beef 0.00425 0.00026 Potatoes 0.00-0.09 0.0033 - - - Table 3. Some pesticide residues in food (derived from Cook Report, 1964. More recent studies show no increase. Consumption calculated from National Food Survey.) BHC Dieldrin Consump- tion (mg/ Range (Wm) Consump- tion (mg/ headlday) 0.0065 Butter Milk Mutton fat Beef fat Consump- tion (mg/ headlday) 0.00 -0.24 0.0023 000 -001 0.0013 0.00 4 .7 0.0033 0.00 -0.15 0.00054 0.00 -0.25 0.00039 0.00 1-0.0 I9 0.00 I7 - - - 0.00-0.062 0.00054 used, residues may remain in the soil for years and they are also likely to find their way into the food chain. Consequently, the residues of these persistent pesticides can be found in many animal and plant tissues, including human adipose tissue. Minute amounts have also been detected in rain and in animals or birds in places remote from pesticide use.2o Fortunately, sensitive methods are available for the detection and measurement of these pesticide residues and it has, therefore, been possible to monitor them in raw materials and in foods.The amounts present in foods in this country at the present time are extremely small (Table 3).21 There is no evidence to suggest that the traces found in food have any significant effects on the consumer and it is also doubtful whether the small residues found in adipose tissue of man and ani- mals have any toxicological significance. Much higher amounts have been found in the adipose tissue of people occupationally exposed to these sub- stances, but these higher residues apparently have no ill effect.20 Oestrogens. Oestrogenic substances occur in many plantsZ2 and they are normal body constituents. Many oestrogens have an anabolic effect; that is to say, they may stimulate protein synthesis and assist in the more economic use of food.They have been administered to cattle and to sheep as implanted pellets or in animal feed. In these animals no significant amount of the administered oestrogen can be demonstrated in the carcass meat. Oestrogens are also used in poultry. In addition to an anabolic effect, they alter the composition of the carcass and the distribution of fat. Thus, their use pro- duces a more acceptable table-bird, which closely resembles the caponized bird. These birds do, however, have a small residue of oestrogen demon- strable in the carcass meat. For this reason, the use of oestrogens in poultry is not permitted in a number of countries. The amount present is extremely small and causes no biological effects in the consumer. Large doses of certain oestrogens may affect the incidence of mammary cancer in certain strains of mouse.In United States law, the Delaney Amend- ment forbids the use of any substance, as a food additive, if it causes an increase in cancer incidence in any animal when administered at any dosage level by any route. This precludes the use of oestrogens in poultry. There is, however, evidence from human as well as animal studies that the minute Frazer 151 amounts of oestrogens present in poultry meat after chemical caponization do not give rise to any cancer risk in the consumer.23 Care must, of course, be taken to ensure that unexpended oestrogen pellets are not included in food products. Natural contaminants: mycotoxins. In 1960, 100 000 young turkeys suddenly died in Britain due to the mysterious ‘turkey X disease’.In due course it was shown that death was due to the eating of groundnuts infested with Aspergillus flaws. This mould, which grows readily in damaged groundnuts, especially if they are kept under hot humid conditions, results in the formation of a lactone, aflatoxin : Oral LD,, for 50 g duckling: 18.2 p g (5 per cent fiducial limits 14.0 -23.8 pg ) Aflatoxin is highly toxic to ducklings and young turkeys. A dose of 20 pg can kill a one-day old duckling. Other animals, such as rats, guinea-pigs, cattle and monkeys, develop liver damage when fed aflatoxin. In the rat this liver damage is followed by the development of malignant tumours.14-28 Thus, contamination with this mould results in the presence in the raw material, and possibly in the food made from it, of a highly toxic and perhaps cancer -inducing substance.Aspergillusflaws can produce aflatoxin when it grows in other cereals, or even in cheese. There are potent mycotoxins produced by other moulds. The significance of aflatoxin in the human diet is still uncertain. Extensive epi- demiological studies are in progress. Fortunately, mould growth is not acceptable on prime materials destined for human consumption and the processes used for refining oil for margarine manufacture remove any aflatoxin that might be present. Methods are available for detection of aflatoxin29 so that quality control of raw materials can be carried out. Efforts are being made to establish acceptable methods at an international level by the Food Section of IUPAC.The role ofthe chemist in relation to raw materials The chemist plays a vital part in the choice of raw materials for food produc- tion, since he can establish the presence and the amount of useful substances they contain. Chemical analytical methods for nutrients are usually quicker and more accurate than biological methods of assay. It is obviously necessary to make certain that the chemical assay method gives a true picture of bio- potency and this may require cross-checking from time to time. The chemist also plays an important part in controlling the presence of undesirable com- ponents, whether they are natural toxicants, residues of chemical aids to R .I . C. Reviews 152 agriculture, or natural contaminants. Thus, suitable specifications and tolerable levels of each of these groups of substances can be established. The chemist also plays a part in the development of new chemical aids and, from the study of biodegradation and other properties, he may be able to devise effective pesticides or other agents that are equally effective, but do not give rise to problems in food. THE DEVELOPMENT OF FOODS FROM RAW MATERIALS As already mentioned, many processes may be applied to raw materials to facilitate food production. Some of these aim to reduce the time interval required to develop acceptable food, others help to produce the measure of uniformity needed for large-scale manufacture, others improve shelf-life, while others again modify the appearance, texture, or other properties of the food to make it more acceptable to the consumer.The use of food additives and processes should be based on certain principles.30 1. A food additive or process should be technologically effective. 2. The amount of the additive used, or the extent to which the process is applied, should not be more than is required to secure the technological objective. 3. A food additive or process should never be used in a manner that might mislead the consumer as to the nature or quality of the food treated. 4. A food additive or process should be safe and give rise to no deleterious effect in the consumer, when properly used. 5. Non-nutrient substances should be kept to the practicable minimum in foods for general use.It is clearly necessary to study the possible effects of any new food additive or food process and this is required in Britain under the terms of the Food and Drugs Act, 1955.3l What are the sort of problems that require study in this connection ? They may be considered under four heads : modification of nutritional value, formation of toxic substances, safety evaluation, and control of contaminants. ModiJication of nutritional value of a food There is nothing sacrosanct about the nutritional properties of a particular food. It is important that the true nutritional value at the time of assimilation should be known. The amount of nutritional modification that the use of an additive or process might bring about can be assessed by chemical analysis, by biological assay, or by both methods.If modification has occurred, the action that is appropriate depends upon the importance of the nutrient in question and the significance of the particular food as a source of that nutrient. Thus, milling to about 70 per cent extraction causes a marked reduction in the thiamine content of flour. Thiamine is an important nutrient for man and flour is a significant source of this vitamin in the human diet. In Britain, flour milled to 70-72 per cent extraction must be supplemented with thiamine and certain other n~trients.3~ This is also required in several other countries. Milling, storage, aeration, treatment with maturing agents and baking all reduce the tocopherol content of flour. However, the nutritional value of Frazer 153 tocopherols in man remains obscure. It has not been thought necessary, therefore, to supplement flour with tocopherols.Variation in tocopherol levels in flour and bread does not appear to have any significant effect on the nutritional status of the consumer.33 In any case, if there was a shortage of tocopherols in the diet of the community, it would be more sensible to supple- ment margarine, since the tocopherols in flour are so labile. Formation of potentially deleterious toxic substunces in food In 1947, Mellanby34 showed that heavy loading of the diet of dogs with flour that had been over-treated with nitrogen trichloride (agene) caused the animals to have ‘running fits’.It was subsequently shown that this effect was due to the formation of an antimetabolite, methionine sulphoximine35 (Fig. 1). This substance causes neurological effects in a number of other animals. Such large doses are required to bring about minimal electro- encephalographic changes in man that it seems unlikely that bread made from agene-treated flour would ever have caused deleterious effects in the consumer. Nevertheless, on the basis of the animal studies, agene was withdrawn from use as a flour maturing agent and its place was taken by chlorine dioxide, which has been shown to give rise to no deleterious effe~ts.3~ The formation of antimetabolites by food additives or food processing is always a possible hazard.Small changes, such as the introduction of a hydroxyl or an amino group, may alter an essential nutrient, such as a vitamin, into a Treatment with I (CH2 )2 Not reversible C H j I HN=S=O I (CH2 12 I CH.NH2 I I COOH Methionine sulphoximine (Toxic anti metabol i te; ’running fits’ in dogs) Fig. I . Comparison of effects of two flour-maturing agents, nitrogen trichloride (agene) and chlorine dioxide, on methionine. 154 I Treatment with s=o I I COOH I CH.NH2 ((32 )z Methionine sulphoxide (Non-toxic m eta bo I i t e) R.Z.C. Reviews Structure R CH3 Pyridoxin HobcH2oH NH2 p -Amino- benzoic acid - - - - - - - - Fig. 2. Some antimetabolites that illustrate the effect of small modifications in chemical structure on biological effects.lethal blocking agent. Some well-known examples of these antimetabolites are shown in Fig. 2. Ever since the demonstration of the action of agene, it has been the practice to test treated or over-treated food as well as the food additive itself when assessing safety-in-use. This gives some assurance that a serious toxic agent has not been formed, especially if the study is combined with an intelligent appraisal of the possible modifications that might occur. The effects of antivitamins or other similar blocking agents may be prevented by the presence of an excess of the normal nutrient; thus, modification of only a small proportion of a nutrient may not be serious. Recently,37 attention has been drawn to another possible problem of this nature.Nitrates and nitrites have been used for many years as preservative agents for meat; they have considerable advantages and the nitrate or nitrite residues involved are known to be harmless. However, when fish is treated in this way, it may have toxic effects on the liver and this has been shown to be due to the formation of nitrosamines; these troublesome effects have been demonstrated in animals fed nitrite-treated fish-meal. The nitrate and nitrite treatment of fish for human consumption is not permitted in Britain. Certain nitrosamines, especially small molecules such as dimethylnitrosamine, Frazer Name of effective vitamin y 3 Thiamin OH NH2 CHZOH I CH3 CH3 \ N=NO / CH3 155 can give rise to cancerous changes in cells.Because of this, a considerable amount of work is now being done to find out whether nitrites can produce nitrosamines in other foods that contain secondary and tertiary amines in much smaller amounts than occur in fish. If nitrosamines are formed, it is important to discover their nature and the amounts present. Evidence from extensive feeding studies that have already been carried out on various meats seems to support the view that the nitrite treatment of meat is safe. Neverthe- less, further analytical and biological studies are now being undertaken to provide definitive evidence on the nature and effects of any substance formed in food as a result of treatment with nitrites. 6. CHECK SPECIFIC EFFECTS IN MAN (if necessary) 7.CONTROL BY ‘PERMITTED LISTS’ and ZONED DISTRIBUTION (if possible) Safety evaluation of food additives There are many food additives used and all new ones are thoroughly studied before they are introduced. Final control is commonly based nowadays on a permitted list system. This is a great improvement on the older approach, which required evidence to show that a substance was harmful before control could be exercised. The main steps in safety evaluation are given in Table 4. Since much has been written on this subject,38,39 I shall only comment on certain aspects. First, the establishment of adequate specifications for a new additive is of major importance. If this is not done, it is impossible to identify the substance or to validate tests alleged to have been made on it.Second, metabolic and biochemical studies are important and often neglected. They should form the basis for the choice of animals for study; in practice, many other factors determine the selection of animals for investigation. Third, the most important studies are long-term studies in animals. Human studies may be useful to indicate the pattern of metabolism in man, or as a means of checking whether some particular effect occurs in the human subject. Otherwise, studies in man have only limited value, especially in the food addi- tive field.40 Experience with drugs suggests that important toxic effects may only be seen when several hundreds of thousands of people are treated and they may well not occur in a clinical trial that only involves a few hundred Table 4.Scheme for control of the safety-in-use of a food additive I. SPECIFICATIONS: Identification: control impurities 2. BIOCHEMICAL STUDY (a) Effects on treated food (b) Digestion, absorption, metabolism, distribution, excretion, half-life 3 . BIOLOGICAL EFFECTS: Short, medium, long-term studies Multi-generation test (Test treated food as well as additive; choose animals on basis of metabolic pattern) 4. ASSESS ‘NO EFFECT’ LEVEL OF IN TAKE: From animal studies Usual safety margin x 100 5. CALCULATE ACCEPTABLE DAILY INTAKE (ADI) FOR MAN: 156 R.I.C. Reviews patients. There is no machinery at the present time for the effective screening of the effects of a new food additive on a large population of consumers.Fourth, this approach to the food additive problem is based on evaluation of scientific evidence. When policy is so based, allowance must be made for new scientific evidence to be taken into account when it becomes available. The system of control needs to be flexible so that appropriate action can be taken quickly. Some of the present administrative machinery used for the control of food additives in Britain is not satisfactory in these respects. Contaminants introduced into foods Certain polycyclic hydrocarbons, such as benzralpyrene, can induce cancerous changes in cells and many aspects of this problem were discussed in detail by Professor Haddow in the fourth Frankland Memorial Lecture.41 Benzpyrene seems to be ubiquitous and it often contaminates our daily food.This contamination may arise in several ways. Benzpyrene could get into food as a result of atmospheric pollution, or because of its presence in smoke used for smoking food. If smoke is generated from old ships’ timbers or road blocks, the tars in them may give rise to considerable amounts of benzpyrene and other related substances in the smoke; other unsuitable fuels may do the same. If such smoke is used for smoking fish or meat, the products may be significantly contaminated with this polycyclic hydrocarbon. A high incidence of cancer was shown to occur in a small community that consumed large amounts of smoked foods, which had been prepared by unsatisfactory methods.42 Contamination can also occur due to faulty cooking stoves.Another possible source of benz[a]pyrene is from paraffin wax containing this hydrocarbon. Changes in the cracking of petroleum may yield wax contaminated in this way; it is important to ensure that such contami- nated waxes are not used for making waxed cartons for f00ds.~3 Milk readily extracts benzopyrene from a contaminated waxed carton. Fortunately, there are sensitive methods available that make it possible to screen foods or waxes for benz[a]pyrene, and a generally acceptable method of analysis is now being elaborated by the Food Section of IUPAC. This problem illustrates the need for careful scrutiny of food preparation method, taking into account all the factors that may be involved.Packaging materials Canning was introduced rather more than 150 years ago and since that time many other forms of food packaging have been developed. No-one would deny that food packaging has made major contributions to the more effective preservation and distribution of food, to the development of cleaner food and to many other aspects of food supply. However, it is necessary to set off against all these advantages some possible disadvantages. The most important Frazer 157 one in this context is contamination of the food with chemical substances leached from the package. Many packaging materials contain substances in addition to the main ingredients, which help to give the package the proper- ties required. These include such substances as curing agents, antioxidants and plasticizers. Some of these substances might have significant toxic effects.Proposed packaging materials are subjected to extraction tests. Clearly, the most satisfactory packaging materials for food are those that give relatively little residue on extraction into a range of solvents simulating food materials. Furthermore, the substances that are extracted in anything more than trace amounts require careful scrutiny and should, if possible, be tested for biological effects. This has been done for many packaging materials, but some substances are not used in sufficient quantities to cover the cost of full toxicological investigation. Clearly, particular attention should be given to packaging materials that are used for staple foods, or for those foods or beverages likely to be consumed in quantity, especially by children.If packag- ing formulations are changed often enough, it is possible that the consumer will not be exposed to any of the ingredients for long periods. It is possible that some use might be made of clearance for relatively short periods of exposure, so that a rotation in the use of some packaging ingredients could be established. Such an approach is worthy of further study. The role of the chemist in relation to food production The chemist is involved at every step that the food technologist may take. He must provide the specifications for food additives used. He must also define in precise chemical terms the nature of the modifications brought about by the use of the food additive or process.He can assist in the identification and control of antimetabolites or other toxic agents that might be formed. He can provide suitable analytical methods which will ensure the control of contami- nation. In these analytical activities, his attention should also be directed to fuel used for food preparation and packaging materials that may be used for wrapping food. The chemist also has an important part to play in the study of the chemical basis of taste and flavour, so that the important organoleptic properties of food can be preserved, enhanced or replaced. EFFECTS ON THE CONSUMER There are a vast number of possible effects of food on the consumer. Some of the more important issues at the present time seem to be concerned with the availability of nutrients, the metabolism of foods, genetic factors and the relationship of food to the pathology and drug therapy of the consumer.Availability of nutrients The food manufacturer is required by law to ensure that food has its correct properties at the moment of sale. For the nutritionist, however, it is more important to know the state of the food at the moment of assimilation. Many things can happen to food between sale and assimilation; it is often stored, prepared, cooked or subjected to other procedures under very variable conditions. The food material may be consumed with other food materials and these might interfere with the absorption of nutrients in the food.For R.I.C. Reviews 158 example, it is well known that phytic acid, present in flour, may interfere with the absorption of calcium. Other factors that may modify the availability of a nutrient include faulty digestion or preparation for absorption, or changes due to the overgrowth of micro-organisms in the intestinal lumen. It should never be assumed that an individual is assimilating the nutrients that are eaten; even less should it be thought that the nutrients indicated from tables of food composition as being present in the foods purchased are necessarily available to the consumer. In the case of some nutrients, such as folic acid, the methods of assay may only indicate what is available to micro-organisms and not what is available to man.A great deal more research is needed on methods of assessing the availability of nutrients to the individual consumer. Metabolism in foods Substances may be modified during the course of digestion and absorption, and they may undergo further changes as a result of enzyme action in the liver or elsewhere. The pattern of enzymes concerned in all these processes differs from one individual to another, since it is genetically determined. It follows that each person deals with nutrients in a different manner. No doubt there are close similarities between a great many people, but each person is, in fact, unique. The pattern is not entirely dependent on genetic factors ; metabolizing enzymes may also be affected by the previous experience of the individual. Such changes have been extensively studied in the case of drugs and it is now well recognized that drug metabolizing enzymes can be increased in activity by induction.There is no reason to suppose that food materials differ funda- mentally from drugs so far as metabolism is concerned. Thus, the previous dietary experience of the individual might be expected to have some bearing on the way in which food is dealt with metabolically. The products of meta- bolism may differ in activity from the parent substance. The development of particular metabolizing enzymes may vary with species, sex, age, or other constitutional characteristics, and this may largely explain differences ob- served in the effects of drugs or of foods on different species of animals or different individuals.An aspect of metabolism that should never be forgotten is the possible effect of the intestinal flora. These micro-organisms may modify food materials and profoundly influence their effects on the body. This may become especially important if there is an overgrowth of intestinal organisms which invade the small intestinal lumen, since the products of floral meta- bolism are more likely to be absorbed from this part of the alimentary tract than from the colon. Genetic factors The influence of genetic factors can be dramatically illustrated by considering some of the so-called inborn errors of metabolism.44 For example, in a child suffering from phenylketonuria, there is an inability to convert phenylalanine into tyrosine due to a defect of the enzyme, phenylalanine hydroxylase (see Fig.3). As a result, phenylalanine accumulates in the tissues and cells, and this causes damage to important tissues, including the brain. Some 1 per cent of mental deficiency in Britain was found to be attributable to phenylketon- uria. Thus, this genetic fault converts a substance which is a dietary essential Frazer 159 Normal individual Phenylalani ne hydroxy lase CH2 CH2 I CH.NH2 CH.NH2 I COOH COOH Phenylketonuric individual Phenylalan i ne h droxylase agsentor deficient I Phen ylalan i ne - - - - - - - - - - - - - - - - - - - - - - - - - - - Phenylalanine accumulation I\‘...-------, Converted to: Damage to cells of central nervous I Ty rosi ne Tyrosine deficient Reduced melanin formation (pigment) Reduced adrenaline formation t phenylpyruvic, phenyllactic, p hen y lacetic acids Excreted in Mental retardation urine 1 ‘ X u 1s ions Fig.3. Enzyme defect i n phenylketonuria and its main consequential effects. for the great majority of people into a disastrous cell-damaging agent in a few. Fortunately, if this genetic fault is detected early enough in life it is possible to exclude phenylalanine from the diet sufficiently to permit normal develop- ment. This is but one example of this type of problem. It is also interesting that the heterozygote parents of these children, who show no apparent ill effect from eating a normal phenylalanine-containing diet, can be shown to deal with phenylalanine less effectively than normal individuals.Metabolic deviations of this sort due to heterozygote traits require much more detailed study. While such abnormalities may only produce the overt metabolic disease in the homozygote, they might contribute to other types of pathology over a much broader field. Detailed study of genetic factors in the nutritional field is much overdue. Pathology and drug therapy Food is consumed by sick people as well as by those that are healthy. For this reason, possible relationships between pathological changes in the consumer and the effects of food require careful consideration. It is well known, for example, that much larger amounts of sodium chloride can be ingested safely by a normal person than by one with hypertension.Again, patients with disease of the small intestine may not be able to tolerate disaccharides in the 160 R. I . C. Reviews diet due to lack of disaccharidases. There is undoubtedly a great deal more to be learnt about the aggravating and damaging effects of food in certain individuals, One of the more dramatic observations of recent years has been the complete recovery of many patients crippled by intestinal malabsorption due to their inability to deal effectively with wheat gluten. If such patients are placed on a gluten-free diet, they return to normal, but will relapse if wheat gluten is reintrod~ced.~5 Drug therapy also calls for consideration. This was well shown a few years ago in patients who were being treated with monoamine oxidase inhibitors.If such patients consumed foods, such as cheese, that are rich in amines serious toxic effects were manife~t.~6 The more that drugs which effect meta- bolizing enzymes are developed, the more careful one will need to be to adjust the diet appropriately to avoid unwanted effects. The role of the chemist in the study of the consumer Although the effects on the consumer may be primarilyamatter of clinical observation, the chemist also plays an important part. The chemist can help to unravel problems of availability of nutrients and he is an essential member of any team concerned with the study of metabolites. The sort of metabolitis formed, the enzymes that are concerned and the factors that affect the nature and amount of metabolites require the attention of chemists and biochemists.Chemical analysis is necessary to establish the biological half-life of any substance. Identification of abnormal levels of nutrients or metabolites in the blood or tissues also calls for skilled analysis. Thus, the whole problem of demonstrating the effects of food on the consumer and of controlling effects to those that are wanted, requires assistance at every turn from chemists and biochemists, working in close collaboration with nutritionists, microbiologists, pathologists and clinicians. CONCLUSION I hope that I have said enough to illustrate my theme-chemistry and nutrition. Nutrition is a broad subject; it is concerned with the chemistry and physics of food and the effect of food on life and health.It has to span the whole field from the production of raw materials through the complex processes of food development into the intricacies of the effects of all the components of food on the cells and tissues of the consumer. It is essential that these problems should be studied at the molecular, cellular and subcellular levels, and it is in this area that advances are likely to be made in the next 25 years. Chemists and biochemists of the highest quality are needed to unravel these problems. Skilled biologists and medical scientists are also needed to elucidate the effects on the consumer, for I hope that I have convinced you that genetic factors and previous experience of the individual must also be taken into account.It is no easy matter to bring about co-operation and collaboration over so wide a field. However, the newly-formed British Nutrition Foundation aims to do just this. It will do everything possible to promote and encourage research and education over the whole field of food and nutrition. I hope that we may count on the help and support of research workers in all relevant fields in this great undertaking. Frazer 161 REFERENCES 1 L. H. Lampitt, Lect. Monogr. Rep. R. Inst. Chem., 1949. 2 Oliver Goldsmith (1730-1774), The deserted village. 3 E. G. Cox, Lecture Ser. R. InJt. Chem., 1963, no. 3. 1963. 4 J. C. Drummond and A. Wilbraham, The Englishman’s food. London: Cape, 1964. 5 R.L. M. Synge, Lect. Monogr. Rep. R. Inst. Chem., 1952, no. 1. 6 M. Stacey, Lect. Monogr. Rep. R. Inst. Chem., 1956, no. 2. 7 K. Bloch, Lipid metabolism. New York: Wiley, 1960. 8 B. Bronte-Stewart, Br. med. Bull., 1958, 14, 243. 9 A. E. Hansen, D. J. D. Adam, H. F. Wiese, R. N. Boelsche and M. E. Haggard, Proc. fourth Int. Con5 biochem. ProbE. Lipids. London : Butterworth, 1958. 10 N. A. Pikaar and J. Fernandes, Am. J . clin. Nutr., 1966, 19, 194. 11 D. B. Jelliffe and K. Stuart, Br. rned. J., 1954, i, 75. 12 C. H. Hassal, K. Reyle and P. Feng, Nature, Lond., 1954, 173, 156. 13 E. C. De Renzo K. W. McKerns, H. H. Bird, W. P. Cekleniak, B. Coulomb and E. Kaleita, Biochem. Pharmac., 1959, 1, 236. 14 F. W. Harris et al., Am. J. Pharm., 1918, 90, 722. 15 J. C. Rothe, 2. Hyg. InfektKrankh., 1919, 88, 1. 16 J. P. K. McCollum, R. C . M. Pearson, H. G. Ingham, P. C. Wood and H. A. Dewar 17 R. L. Wain, Lecture Ser., R. Inst. Chem., 1965, no. 3. Lancet, 1968, ii, 767. 18 G. Walton, Am. J. publ. Hlth, 1951, 41, 986. 19 E. H. W. J. Burden, Analyst, Lond., 1961, 86,429. 20 A. C. Frazer, A. Rev. Pharmac., 1967, 7, 319. 21 Min. Agric., Fisheries & Food, Review of the persistent organochlorine pesticides. London: HMSO, 1964. 22 R. B. Bradbury and D. E. White, Vitams Horm., 1954, 12, 207. 23 S. Wallach and P. H. Henneman, J. Am. med. Ass., 1959, 171, 1637. 24 M. C. Lancaster, F. P. Jenkins and J. McL. Philp, Nature, Lond., 1961, 192, 1095. 25 B. F. Nesbitt, J. O’Kelly, K. Sergeant and A. Sheridan, Nature, Lond., 1962, 195, 1062. 26 H. DeIongh, R. K. Beerthuis, R. 0. Vles, C. B. Barrett and W. 0. Ord, Biochim. biophys. Acta, 1962, 65, 548. 27 R. Allcroft and G. Lewis, Vet. Rec., 1963, 75, 487. 28 W. H. Butler and J. M. Barnes, Br. J. Cancer, 1964, 17, 699. 29 H. DeIongh, R. 0. Vles and P. de Vogel, Mycotoxins in .foodstgfs. Cambridge, Mass. : MIT Press. 30 Tech. Rep. Ser. Wld. Hlth Org., 1957, 129; FA0 Nutr. Mtg Rep. Ser., 1957, 15. 31 Food & Drugs Act, 1955, 4. Eliz. 2. 16. 32 Decontrol of cereals and feeding stufs, UK, Cmd 8745. London: HMSO, 1953. 33 A. C. Frazer add J. G. Lines, J. Sci. Fd Agric., 1967, 18, 203. 35 H. R. Bentley, E. E. McDermott, T. Moran, J. Pace and J. K. Whitehead, Proc. R. Soc., 34 E. Mellanby, Brit. med. J., 1947, ii, 288. 1950, B137,402. 36 A. C. Frazer, J. R. Hickman, P. Meredith and H. G. Sammons, J. Sci. Fd Agric., 1956, 7, 361, 371, 375,464. 37 Editorial, Lancet, 1968, i, 1071. 38 A. C. Frazer, J. Sci. Fd Agric., 1951, 2, 1. 39 Tech. Rep. Ser. Wld Hlth Org. 1958, 144; FA0 Nutr. Mtg. Rep. Ser., 1958, 17. 40 A. C. Frazer, Lex et Scientia, 1967, 4, 63. 41 A. Haddow, Lect. Monogr. Rep. R. Inst. Chem., 1959, no. 4. 42 B. D. Kaufman, A. I. Mironova and L. M. Shabad, Vop. Onkol., 1959, 5, 314. 43 P. Shubik, G. Della Porta and K. Spencer, Acta Un. int. Cuncr., 1959, 15, 232. 44 J. B. Stanbury, J. B. Wyngaarden and D. C. Frederickson, The metabolic basis of inherited disease. New York : McGraw-Hill, 1966. 45 A. C. Frazer, Malabsorption syndromes. London : Heinemann, 1968. 46 A. M. Asatoor, A. J. Levi and M. D. Milne, Lancet, 1963, ii, 733. R.I.C. Reviews 162

ISSN:0035-8940    

DOI:10.1039/RR9690200143

出版商:RSC    

年代:1969

数据来源: RSC

摘要:

NUCLEAR FUELS B. R. T. Frost, B.Sc., Ph.D., F.I.M. Metallurgy Division, Atomic Energy Research Establishment, Harwell, Didcot, Berks.* Fuel manufacture . . The fuel cycle, 168 Ore to metal or ceramic fuel, 168 Metal fuels, 169 Uranium dioxide, 17 1 Carbides, 175 Other ceramic fuels, 176 Reprocessing, 177 . . . . . . . . . . . . . . 190 - Conclusion . . . . . . . . .. .. .. .. . . 204 References . . . . . . . . . . * . .. . . . . . . .. . . . . 168 Physical and chemical properties. . . . .. . . . . . . 181 . Physical properties, 18 1 Chemical properties, 185 Irradiation behaviour 2ggTh + In0 -+ 2iJ;Th + y 2$iTh -+ 0/3- + ";Pa ";Pa -+ 0/3- + 233U 92 It has taken only 27 years since the discovery of nuclear fission to establish the nuclear reactor as a cheaper source of electrical power than coal or oil (see Fig.I ) . 1 The unique feature of a reactor is its core in which a self-sustaining fission process is used as a convenient heat source. In the great majority of reactors the fissioning species is 235U which occurs in natural uranium at a concentration of 0.7 per cent, the remainder being 238U. The latter will undergo fission when bombarded with neutrons of high energy but more generally it absorbs neutrons to produce 239Pu by the reactions: 239Pu is in many respects a better fuel than 235U so that this breeding process enlarges our energy resources considerably by utilizing the otherwise useless 238U. The energy resources may be further enlarged by producing fissionable 233U from thorium: * Present address: Metallurgy Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, Illinois 60439.163 .. .. . . 205 Frost 1.5 I I I I I I I I I I I I I Nuclear (plagnox) Coal-fired (remote from central coal fields) 2 Y 0, L n aJ aJ a C 0.5 / Oil-fired (including oil tax) I ! I975 0 0 0 Coal-fired (near central coal fields) I I I I I I I I I I I I970 - w I 1 Year of commissioning I965 Fig. I. Nuclear power stations now produce electricity at a lower cost than coal-fired plants serving t h e same parts of the country. The Magnox reactors, on which the first 5000 MW nuclear programme of the CEGB is based, use uranium metal fuel of natural enrichment and will produce considerable quantities of 239Pu as a by-product.This will have to be separated by chemical means from the uranium and fission products and will almost certainly be used as the fuel in fast breeder reactors, the prototype of which is being constructed at Dounreay. The Advanced Gas-Cooled Reactor (AGR), large-scale commercial versions of which are under construction at Dungeness and Hinkley Point, uses slightly enriched uranium in the form of UOZ pellets. The enrichment of the 235U isotope is performed in a process which is dependent on the slightly different rates of gaseous diffusion of 235U + nl + 95X + 139Y + 2nl I .o 0 238UFg and 235UFg. A fissionable atom, e.g. z35U, located on a crystal lattice in, say, a pellet made of sintered UOZ crystals undergoes fission when a neutron of suitable energy is absorbed.This occurs because the resulting nucleus is very unstable and splits into two parts of roughly equal mass, e.g. There is a discrepancy in mass between the two sides of this equation which corresponds to an energy release of about 200 MeV.* Most of this energy is imparted to the two fission fragments, which leave the site of fission very rapidly, travelling in straight lines of length about 10 pm before coming to rest. In doing so they impart their energy to the parent lattice, essentially as thermal vibrations, and incidentally causing considerable damage to the ~ ~~ ~~~ _ _ _ _ _ - ~ - _ _ _ ~ ~ _ _ _ _ _ J. * 1 MeV is approximately equal to 1.602 x R.I.C.Reviews 164 lattice. It is this energy, plus a smaller amount arising from other sources, which represents the heat source in the nuclear fuel which must be converted to a more useful form. Until now this has been done by using the heat to make high-temperature steam (typically 560 "C and 2400 lb in-2 pressure) which drives a turbine connected to an alternator (Fig. 2). In this method, nuclear fuel is being used as a direct alternative to coal or oil and there is a similar incentive to derive the maximum Carnot efficiency by working at high temperatures. The nuclear plant usually costs more than the fossil fuel plant to build (although there is recent evidence to suggest that this is no longer the case)2 and it is the cheapness of the nuclear fuel compared with coal or oil which makes its electricity generating costs competitive.It is possible to use nuclear heat in other ways. Two important examples are : 1. As a direct producer of dc electricity. If the fuel is a suitable electron emitting material, e.g. UC, it can be made to form the cathode of a diode and so produce electricity directly. The fuel surface must be at a temperature in excess of 1500 "C to produce an efficient diode. Experimental prototypes have operated successfully for many hours.3 2. As a source of process heat. An outstanding and fast-developing example of this application is in the desalination of sea water by flash distillation. A large nuclear plant might produce both fresh water and electricity on an economic basis in arid regions.4 The economic use of nuclear fuel depends on its ability to remain dimension- ally and chemically stable for a long period of time in a very unusual environ- ment. Firstly, the fuel limits its own life by the production of fission products which occupy a greater volume than the original uranium atoms and cause damage to the fuel material.Ten per cent of the fission product atoms are feed pump 165 Fig. 2. A schematic diagram t o show the conversion of nuclear-generated heat into energy via high-pressure st earn. Frost 12 noble gases (Xe and Kr) and if these congregate together to form bubbles they will cause an even greater volume change. At the same time the fuel must operate at a high surface temperature (-600 "C) to give a high thermodynamic efficiency and it must be reasonably inert chemically to the coolant, which may be a gas such as COz or a liquid such as sodium.Thus, the qualities required of a nuclear fuel are: (i) high melting point, (ii) high thermal conductivity, (iii) high density of uranium atoms, (iv) absence of highly absorbing species, e.g. boron or gadolinium, and (v) chemical inertness to oxidizing liquids and gases and to metal cladding materials. These conditions are most closely met by ceramic fuels, the properties of which are compared in Table 1. However, the first developments were con- cerned with metallic fuels. The earliest type of reactor to be developed in the UK on a large scale was gas-cooled and graphite moderated.To achieve criticality on a natural uranium fuel it was necessary to use the metallic form since this has the Table I. Comparison of ceramic fuel materials Fuel Metal uo2 PUOZ Cubic (f I uo r i t e) UOZ-~OPUOZ Solid uc PUC UC-2OPuC UN Pu N UN-20PuN PUP a At fuel operating temperatures us PUS UP 166 f 30 I660 2480b 2850 i 30 2600 (I atm) > 2500 2450 i 30 2330 2610 f 30 2600 (2 atm) Density density Melting expansion conductivity Structure (g cm-3) (g cm-3) point ("C) ( x 106 "C-1) (cal cm-1 s-1 "C-1) 9.66 10.96 Cubic (fluorite) 10.12 2750 It 40 2280 11.46 2740 9.76 I I .06 solution FCC 13.63 2350b 12.97 12.87 13.53 13.6 12.9 FCC Solid solution FCC 14.32 13.53 14.25 FCC 13.47 14.3 10.86 FCC 13.5 9.57 10.57 10.23 FCC FCC 9.35 9.07 9.89 FCC 8.77 10.8 0.018 (100 "C) (to 800 "C) 0.008 (700 "C) 9.3 10.3 9.7 11.9 9.2 (to 800 "C) 13 8.6 (to 980 oc) 9.7 b 2000 ppm oxygen.Coefficient of thermal Thermal - 0.005-0.007a 11.5 0.05 (to 870 "C) (I 00-700 "C) - 0.046" 0.04 (300 "C) 0.05 (800 "C) - 9.8 0.043" 11.8 0.028 (100 "C) (to 1000 "C) 0.041 (800 "C) 18.5 - 0.032 (100 "C) 0.039 (800 ocj - R. I.C. Reviews maximum density of uranium atoms. Uranium metal does not have a very high melting point (1 133 “C) and it exists in three different crystallographic forms in the solid; temperature excursions from one form to another cause a change in shape of the crystals.This phenomenon is known as ‘growth’ and has imposed limitations on the use of uranium metal. The metal is also chemically reactive, oxidizing readily in air, C02 and water. Finally, its low melting point together with the internal stresses set up by growth lead to a high fission-gas swelling rate. Notwithstanding these limitations the Magnox reactors have provided a reliable and increasingly competitive form of power, as Fig. I illustrates. The development of water-cooled reactors, initially as submarine propulsion units in the USA, led to the realization that U02 was, in many respects, a better fuel than uranium metal. Although the low fissile-atom density in U02 generally requires the use of enrichment, this is offset by the chemical inertness, very high melting point (2800°C) and lack of phase transformations in the solid state.5 Thus, it is possible to burn several times the number of 235U atoms in oxide to produce the same swelling as in metal.A further advantage of UOZ is that it is isomorphous with Pu02 and Tho2 and solid solutions of UO? Pu02 can be used as a fast reactor fuel and of UO2-ThO2 for thermal breeders. However, a major disadvantage of U02 is its low thermal conductivity which leads to high fuel operating temperatures-a temperature in the centre of the fuel pellets of 2000°C is not uncommon. Under these conditions a high proportion of the gaseous fission products move out of the fuel and pressurize the metal can which separates the fuel from the coolant, thereby complicating fuel element design.There are two answers to this problem. One is to disperse the oxide in a metallic matrix, e.g. stainless steel, which improves the thermal conductivity and restrains the swelling of individual oxide granules.6 These ‘cermet’ fuels are used in special applications, e.g. research and military reactors, where reliability is more important than economic operation. A second solution is to use uranium carbide which has nearly as high a melting point as U02 (2350°C) but a thermal conductivity five to 10 times higher. The carbide is more chemically reactive (comparable with uranium metal) but has a higher density than UOZ. Its use, therefore, has been confined to reactors using relatively inert coolants : sodium and terphenyls.A special application of carbide is as a dispersion of uranium-thorium dicarbide (U,Th)C2 in graphite. In the Dragon type of high temperature gas-cooled reactor the fuel element contains the fuel and moderator, thus minimizing graphite radiation- damage problems and avoiding the use of neutron-absorbing metal cladding. A unique feature of this type of fuel element is the coating of each dicarbide particle ( 4 0 0 pm diameter) with pyrolytic carbon to retain the fission products and minimize fission product release.’ From this brief discussion it can be seen that different reactor types demand the development of different types of fuel. The development of a fuel can be a lengthy and complex process, starting with the preparation of pure material from the ore and its fabrication into suitable shapes, followed by the evaluation of physical and chemical properties and a study of irradiation behaviour.Finally, when a fuel element design has been produced it must be tested Frost 167 rigorously on a statistical basis, generally in a prototype power-producing reactor such as the Calder Hall Magnox reactors and the Windscale AGR. These steps in the evolution of a fuel element will be traced in some detail in the remainder of this review. FUEL MANUFACTURE The fuel cycle Nuclear fuels have a high specific value, that is they are expensive to produce in terms of cost per gram but each gram is a potential source of much energy. Thus, a kilogram of 235U costs about E5000 but can theoretically produce 2.4 x 1010 kilowatts of heat when burned in a reactor.The conservation of fuel and its strategic use are important considerations in power reactor technology and the fuel cycle must be seen as one of the three important factors affecting the economics of nuclear power, the other two being the capital cost of the reactor plant and the operating costs. Figure 3 shows a schematic diagram of a fuel cycle. Ore to metal or ceramic fuel Uranium is most commonly found as uraninite, a form of pitchblende, which is basically a mixture of uranium oxides associated with lead, lanthanons and thorium.8 Commercial ores, as mined at present, contain at least 0.1 per cent u308. World reserves of uranium from this type of ore are estimated as several million tons.Lower grade ores are known to exist in large quantities and the dissolved salts of uranium in sea water, which are at the ppm level, represent a source of lo9 tonnes of uranium.9 However, extraction of uranium from these more dilute sources will involve more expensive extraction processes and will only be exploited when the supply of more concentrated ores is exhausted. Initially, the ores are crushed and concentrated physically or by roasting and then the uranium is dissolved by acid or sodium carbonate leaching. The crude liquor, containing about 5 g 1-1 of uranium, is purified by an ion exchange process, and then extracted into tributyl phosphate, the impurities being discarded in the aqueous phase.The pure uranyl nitrate solution which results is converted to uranium dioxide. In the UKAEA process,lO ammonia is added to Fig. 3. Schematic diagram of a fuel cycle. R.I. C. Reviews 168 Reactor Storage hoppers / ng t Spra To reduction reactor jacket heater ija; i pipe + Internal ' I1 heaters Lift pot 1 -% To scrubber From co m pressor Heat exchanger Fig. 4. Schematic diagram of the process for the denitration of uranyl nitrate. the nitrate to precipitate ammonium diuranate (ADU), (NH4)2U4013. This may be decomposed, by heating, to U03 or US08 which is reduced in hydro- gen to U02. The route chosen depends on the end product required. For the production of metal, ADU is decomposed to U03 below 350°C and reduced with hydrogen at about 700 "C.For ceramic oxide, ADU is decomposed above 450 "C to u308 which is reduced in hydrogen at -700 "C to give a powder with a lower reactivity or surface area per unit weight. In the USA, uranyl nitrate is evaporated to form molten uranyl nitrate hexahydrate which decomposes to U03 at 300 "C (Fig. 4). uo3 is then reduced with hydrogen at 600-800°C. These processes may be operated in a fluidized bed to give a continuous throughput. To produce uranium metal, U02 is converted to UF4 by treatment with HF: U02 + 4HF + UF4 + 2H20 -AH298 = 265.6 kJ mol-1 The UF4 is then reacted with high-purity calcium or with magnesium in a 'bomb', the efficiency of conversion being about 97 per cent. This is carried out by UKAEA at its Springfield Works on a 200 kg batch scale.The processes just described refer to the use of uranium at its natural isotopic composition. In many reactors it is necessary to enrich the z35U isotope con- centration to achieve criticality. This is done by gaseous diffusion of UF6 through a cascade of membranes. The UF6 is made by fluorination of UF4 between 450 and 550 "C (Fig. 5 ) and, after the diffusion process, is reconverted to UF4 by reaction with hydrogen in a fluidizing column or is hydrolysed to UO2F2 and reacted with aqueous ammonia to form ADU, which is then converted to U02 as described above. Metal fuels Enriched or natural uranium metal billets are melted by high-frequency induction heating under vacuum in magnesia-coated graphite crucibles and are Frost 169 I I L Fig.6. Cutaway view of a Mk 5 fuel element showing i t s method of construction. R. I.C. Reviews 170 Fig. 5. Flow diagram for uranium hexafluoride production. t F6 Stack cast into rods close in size to the final fuel rod. These are then machined to the final dimensions. For certain applications uranium is alloyed with other metals to produce fuels with improved performance or resistance to corrosion. For example, the ‘driver’ charge in the Dounreay fast reactor consists of highly enriched uranium alloyed with molybdenum. This allows it to operate to higher burn-ups than would otherwise be possible. The fuel elements in water- moderated research reactors of the DIDO type in this country and the MTR and ETR types in the USA are made from uranium-aluminium alloys.The two metals are melted together and cast into rectangular blocks which are then clad in aluminium and rolled into sheets which can be formed into desired shapes (Fig. 6). The uranium is in the form of small particles of UAl4 or UAl3 dispersed in a matrix of aluminium. In this way the swelling effects of the fission products are minimized and the uranium cannot react with the water coolant (vide infra). Uranium dioxide UOZ is mainly used in the form of dense, sintered cylindrical pellets, typically 1-2 cm in diameter. These are generally made by cold pressing and sintering UOZ powder, although other methods such as hot pressing are occasionally used. The ideal characteristics of UOz powder are:5 ( i ) a small particle size, (ii) regular shape, (iii) a ‘clean’ surface, i.e.a minimum of adsorbed gases, and (iv) a large surface area per unit mass. To some extent i and iv are incompatible with iii. The characteristics of the powders obviously depend upon the nature of the material from which they are formed.12 When U03 and u308 are reduced to U02 there is a large decrease in specific volume which leads to particle fracture and an increase in surface area. Careful control of the reduction temperature is needed to ensure that this effect is not counteracted by coalescence. Generally, UOZ powder from the reduction process is reduced in size by grinding in a rod or ball mill for several hours. Care must be taken not to pick up ceramic particles in this process.This can be done by proper choice of mill materials; rubber-lined mills are often used. In Fig. 7 the effects are shown of the powder preparation route, the precipitant and the milling procedure on UOz particle size. After milling, the powders are not generally free flowing and therefore will not fill dies with the same quantity of material each time. The powders are treated by the addition of 1-2 per cent of binder, generally an organic fluid such as polyethylene glycol, paraffin wax or polymethyl methacrylate ; this may be added dry or in solution. The powders are then granulated through sieves to produce a free flowing powder which is poured into tungsten carbide lined dies and compacted under a pressure of up to 150 ton in-2.To ensure uniform densification of the powder and for ease of loading into stainless- steel tubes, UOz is generally used in the form of right cylinders. The ‘green’ pellets are debonded (i.e. the binder is removed) by heating in an inert or 171 Frost 90 - A-High-pressure steam oxidation B-MCW PWR core-I UO, 8o - C-MCW Precipitated U02 - 60 50 70 - D-Fluidized-bed denitration, UO, reduction - .- 100. M P 2 9 0 - W 2 80- 7 0 - C Q) 60 - - 50 a 40- 30 - - 20 - 10 0 Fig. 7. Uranium dioxide particle size distribution: (a) the effect of powder preparation, and (b) the effect of milling procedure. slightly oxidizing atmosphere at about 800°C. They are placed on molyb- denum trays and heated in a hydrogen atmosphere in molybdenum wound furnaces to promote sintering; several hours at 1500-1700°C are required to produce high-density pellets.13 Generally the aim is to produce pellets with about 5 per cent residual porosity ( i e .95 per cent of theoretical density). The porosity is then mostly within the grains and is ‘closed’, i.e. not accessible from the outside of the pellet. The reasons for choosing this density is discussed later when properties are considered. Figure 8 shows the effects of various process variables on the final sinter density of UOz. The most common form in which ceramic fuels are used is as right cylindrical pellets inside metal tubes. However, the quest for improved performance and 172 (a) - - - - - - - - - (b) - - - - - - - - 6-U rea-preci pitated diuranate UO, C-’B’ milled D-’D’ milled - I 1 I R.I.C. Reviews 74 72 90 'i 70 - t 88 86 68 Condition: as received - 66 U U f- + 1% PVA Ram diam., 64 E 9 a 0.41 I in Sintered for IOh 82 62 200 I Si ntered: 50 I 1800 a t 1725OC in H, I00 Compacting pressure (ton in-,) Compacted a t 100 ton in-, Sintered in H, I I 1700 1600 Si nteri ng temperature ("C) 5 s 25 86 I I t 1400 1500 Fig.8. Effects of processing variables on the sintered density of MCW UOz compacts. (a) As pressed and sintered density as a function of compacting pressure. (b) Sintered density as a function of compacting pressure and sintering time.(c) Effect of time and temperature on sintered density. (d) Sintering time as a function of compacting pressure and sintered density. economics has led to the development of other fuel forms, the most important being vibrocompacted fuel and coated particles. If fuel swelling considerations (see below) dictate the need for a low fuel density it may be uneconomic to make carefully sized pellets. It may be cheaper to make sintered spheres of about 500-1000 pm diameter and pack these into metal tubes;14 a single size of sphere gives a maximum packing density of about 74 per cent. By adding smaller particles, ideally reducing in diameter by multiples of seven, densities up to 90 per cent of theoretical can be made.Packing is achieved by vibrating the metal tube mechanically. 200 300 Compacting pressure (ton in- 2 ) Frost Sintered at 1725 OC t '& 90% dens^^^^ 60 74 ' I 40 as received + Condition: I yo PVA in H2 I 1 I l l 100 Compacting pressure (ton in-2) 100 200 250 - 5 I- I 92% dens: \ ' \ \ \ \ \ \ 194% \ dense -1 I 60 30 i 173 As explained later, improved fuel performance can be obtained by dis- persing fuel particles in an inert, conducting matrix. Stainless steel-UO2 cermets have given outstanding performance in small military reactors where fuel cycle economics are not of primary importance. In the Dragon type of high-temperatureereactor the fuel particles-oxide or carbide-are dispersed in graphite moderator.To restrict fission product release the particles are coated with carbon which is deposited pyrolytically from hydrocarbons in a fluidized bed (Fig. 9). The fabrication of oxide fuels enriched with plutonium instead of 235U is similar to that of UOZ. Plutonium, unlike uranium, can exist in the trivalent state so that a wider range of oxygen : metal ratios is possible than in UO2 and care must be taken in selecting the sintering atmosphere. The mixed oxide powder may be prepared by physically mixing PUOZ and U02 powders; the PuOz is made from nitrate solution by precipitation as the hydroxide followed by calcining. Alternatively, plutonium hydroxide and ADU may be coprecipi- tated from a mixed nitrate solution, calcined and reduced in hydrogen.The resulting powders are then pressed and sintered in the same way as U02, the sintering atmosphere being argon or helium-hydrogen mixtures or wet hydrogen.15 During these operations, and indeed in all operations on enriched fuel, precautions must be taken to limit the concentration of fissile material in localized areas to well below that required to form a critical mass. Generally this involves careful control of fuel movement. All steps in the fabrication of plutonium fuels must be carried out inside glove boxes to isolate the toxic &-active plutonium. In the future, when plutonium is recycled after irradiation to high burn-ups, appreciable levels of y-activity will arise and operations will have to take place in ~$7 shielded cells.Fig. 9. Photomicrograph of fuel particles, showing pyrolytically deposited carbon coating. R. I , C. Reviews 174 uoz + 3 c + u c + 2 c o 1' u02 + 4 c +uc2 + 2 c o 1' Carbides Of the three compounds in the uranium-carbon phase diagram, the mono- carbide is of interest in fast reactors and the dicarbide in high temperature gas- cooled (Dragon) type reactors. These carbides may be prepared by at least three methods : (i) direct reaction between uranium metal and carbon-this is difficult to control ; (ii) reaction of uranium, made reactive by hydriding, with hydrocarbons16 -this works at 600-800°C but it involves the use of uranium metal, introduc- ing an undesirable extra step in the fuel cycle; and (iii) UOz-carbon reaction-this is the most commonly used route and can be employed to make monocarbide, and dicarbide, To drive the reaction forward the CO must be removed rapidly; this is done either by reduction i n vacua with a high pumping speed17 or in a fluidized bed with a rapid flow of argon to entrain the CO.18 The reaction is generally carried out at 1400-1 500 "C.The product contains between 0.2 and 0.4 wt per cent oxygen and less than 100 ppm nitrogen, both in solid solution, and is very reactive to air and particularly to water vapour. From this stage onwards the fabrication technique is generally similar to that for UOZ except that all operations, whether with plutonium or not, must be carried out in dry atmospheres in glove boxes. In experiments at Harwell, Russell and Harrisonlg crushed reacted (U,Pu)C to - 30 mesh and milled the powder in a tungsten carbide mill with tungsten carbide balls. The powder was then pressed into pellets at 40 ton in-2 and sintered at 1550 "C for four hours in flowing argon.The final density was -94-95 per cent of theoretical; this was only achieved by taking precautions to eliminate surface oxidation of the powder which had been milled to about 2 pm particle size. High sinter densities can be achieved more easily by adding a small amount of nickel to the powder prior to pressing. In experiments at the United Nuclear Corporation in the USA 0.1 to 0.5 per cent nickel was added and densities as high as 98 per cent of theoretical could be obtained without difficulty;20 the mechanism involved has not yet been elucidated.UC has a lower melting point than U02 (-2350 "C as compared to 2800"C), is a good conductor, and does not have very high vapour pressure at its melting point. It is possible, therefore, to make carbide fuel by arc melting uranium and carbon together and casting into moulds. A considerable amount of development of this route was carried out by Atomics International in the USA for a UC fuelled thermal reactor of the sodium-cooled graphite- moderated type.21 Rods up to 1.0 in diameter were cast on a production scale for incorporation into long fuel elements. Arc-cast carbide differs from the sintered product in having a much larger grain size and a lower oxygen and nitrogen content.With plutonium fuels the loss of plutonium by vaporization is high and although this route has been studied it has been dropped in favour of the sintering method which has the advantage that it can be carried out in 175 Frost Final 3 so I evaporator column M (L___JI 1 s z Thermal denitration vessel I I g& Aqueous waste tank uo2 Sol feed tank n Other ceramic fuels Fig. 10. Schematic flowsheet for the production of (U,Pu)O2 spheres. equipment built for oxides, with the simple addition of the carbothermic reduction step. The coated carbide fuel particles used in Dragon-type reactors are generally not UC2 but (U,Th)Cz since the original objective was to convert thorium to 233U. Three methods have been used to make fuel particles. 1.Powder agglomeration. UOZ, Tho2 and carbon flour are mixed together with a binder and granulated to form green granules of -500 pm diameter. These are reacted and sintered either under vacuum at about 2000°C or in a fluidized bed at -1 700-1 800 "C. The latter is preferred since, after the reaction, hydrocarbons can be introduced to coat the particles with carbon. 2. Melting in an arc or plasma, or in a bed of carbon. The granules so produced contain shrinkage cavities and high internal stresses. 3. Sol-gel process.22 This has been developed primarily for oxide fuel but can be used for carbides. Oxide is dispersed in an electrolyte to form a sol. Droplets of the sol are dispersed in an immiscible liquid and are hardened or gelled by reaction with a suitable reagent, e.g.2-ethylhexanol. The spheres are then dried and calcined. A flowsheet developed at Oak Ridge National Laboratory for the production of (U,Pu)Oz spheres is shown in Fig. 20. To make carbides, carbon black is added at the sol stage and the final calcining temperature is higher. The advantages of this process are that it produces spheres continuously in simple columns, the product being very regular in size and shape and possessing a very fine grain size which facilitates sintering. Laboratory studies have been made and are continuing on uranium nitride, phosphide and sulphide which are similar in structure to UC.23 Generally the G3-l I Reject 1 *duct Roundometer 176 '.LJ -9 Drier 1 1 1 Furnace i j Screens I R.I. C. Reviews starting point for their preparation is uranium, prepared in a reactive form by first forming UH3 at 250-270 "C. The temperature is raised to 400-600 "C and a suitable gas admitted (nitrogen, phosphine or HzS). The hydride decomposes to UN + N2 at 1400°C in argon. With phosphine, U3P4 is formed and is and a compound is formed. With nitrogen, UN1.7 is formed and is decomposed decomposed to UP at 1400°C. HzS forms ,%US2 which decomposes to a mixture of US2, u2s3 and U3S5 at 1400°C and to US at 1800°C. The powders may be pressed and sintered in a similar manner to UC. Reprocessing The life of a nuclear fuel in a reactor core is generally dictated by the nett burn-out of fissile material, the production of neutron-absorbing fission products, and dimensional changes in the fuel due to the conversion of heavy fissile atoms to less heavy fission products.An additional factor which may over-ride these considerations is the production of plutonium either for military purposes, in which case the need is to keep the higher isotopes of plutonium to a low level, or in the blankets of fast breeder reactors where the objective is to produce plutonium rather than power. With fuels irradiated in thermal reactors the objective of reprocessing is to separate the uranium, the plutonium (or z33U if thorium is presbnt) and the fission products. The separated uranium is either made up to the required enrichment and recycled through the thermal system or it may become the breeder of a fast reactor. The plutonium is probably used in a fast reactor although, until their technology is firmly established, plutonium may be used to enrich thermal-reactor fuel.This is certainly the intention in the USA; the UK, however, plans to introduce fast reactors as early as possible and will reserve its plutonium for this purpose. In fast reactor fuels the main requirement is to remove the fission products. Since the fuel contains both uranium and plutonium these need not be separated in reprocessing although the plutonium concentration will need adjustment. There are basically two types of reprocessing : hydrometallurgical-which operates below -100 "C-and pyrometallurgical or high-temperature pro- cessing.The first stage in hydrometallurgical reprocessing is to dissolve the fuel in nitric acid. Ideally the fuel should be separated from its cladding and in the Magnox reactors which use metal fuel this is relatively easy; with ceramic fuels, particularly those in dispersed forms, it is necessary to chop up or crush the whole assembly and to leach the fuel from the structural material. The separation of uranium and plutonium from the fission products is based on the solubility of the two actinides in their four-valent state in a number of organic compounds such as diethyl ether, tributyl phosphate and Butex (/3,/3'- dibutoxydiethyl ether). Plutonium may then be separated from uranium by reducing it to a lower valency, e.g. by ferrous sulphamate. The process is engineered by means of countercurrent contactors which may be packed columns, pulsed columns or mixer-settlers.24 The process is shown diagram- matically in Fig.11. The aqueous (nitrate) solution is fed into the middle of the columns and flows upwards. This phase, rich in U and Pu, flows to the plutonium stripping column and finally to a uranium stripping column; the Frost 171 Demin. Demin. Demin. Sodium water carbonate Demin. Water HNO, I Nitric a c i d 0 Plutonium Demin. water ~~~~~~ Solvent Solvent Uranium solution solution water plutonium reductant Metal fuel elements l r I r rei3 Scrub . I i Solvent: tributyl phosphate *ea Flow sensing device (orfice or remote indicating rotameter) Flow control motor solution To To To waste disposal uranium processing plutonium processing Fig.1 I. Processing flowsheet for separating Pu, U and fission products by solvent extraction. material to prevent accidental build-up of insolubles, the prevention of excessive radiation decomposition of the organic solvent, and the disposal of the radioactive wastes. A view of part of the Windscale plant is shown in Fig. 12. This account deals only with general principles and scarcely does justice to an important and complex subject; the interested reader should see references 24 and 25. Falls in the USA.26 The simplest variant of this type of processing is melt refining where a metallic fuel is melted under vacuum in oxide crucibles. The more volatile fission products vaporize and are trapped elsewhere while the rare earths and yttrium form a slag or dross with the crucible material.The remaining melt, which comprises uranium, plutonium and the noble fission products such as ruthenium and molybdenum, may be filtered or bottom- poured into moulds. There is generally a remelting and casting step where the 179 Fig. 12. Part of the Windscale, UK, recovery plant. important product in each case is an aqueous solution of the actinide and the organic phase is sent for recovery and recycle. The uranium and plutonium solutions are concentrated, after adequate decontamination, and converted to ADU and hence to U02 or to plutonium hydroxide and Pu02. Important considerations in the design and operation of such plants are the prevention of the formation of critical masses of solutions, accurate accounting of the fissile Aqueous reprocessing has become the standard method world-wide.However, a considerable amount of effort has been devoted to alternative processes, most of which are pyrometallurgical in character. Indeed, one such process has operated as an integral part of the EBR-I1 fast reactor at Idaho Frost plutonium content is adjusted. The product is a U,Pu-fissium alloy which is used as the fuel for the EBR-I1 reactor. This fuel does not perform very well from an irradiation standpoint : oxide and carbide give better performance, but there is a problem of whether they can be accommodated in a high- temperature process. In one process developed in the USA the oxide fuel is reduced to metal with magnesium at -750°C and dissolved in a liquid metal such as zinc.By a fractional crystallization process the uranium is separated (as U-Zn compounds) from the fission products and the zinc is finally removed by vaporization. Alternatively the molten U-Pu alloy may be con- tacted with molten salts to remove the fission products and eventually con- verted back to oxide or made into carbide. A notional scheme for such a process is shown in Fig. 13. The other type of process which has received considerable attention is the ‘fluoride volatility process’ in which oxide or carbide fuel is converted to hexafluoride in a fluidized bed.27 This separates the U,Pu and volatile fission- product fluorides from the non-volatile fluorides.UF6 and PUF6 are then separated from the fission-product fluorides by fractional distillation and finally converted to the oxide or to the tetrafluoride and hence to metal and carbide. A flow sheet for a carbide process is shown in Fig. 14. Economic assessments show that where aqueous processing is firmly established, as in the UK and the USA, these high-temperature processes have little to offer in the way of improved fuel cycle economics although they may offer an advantage in reducing fuel cooling times and hence reducing fissile inventories. The waste products from chemical reprocessing, i.e. the fission products, present a problem in their storage and disposal. At present the common Fig.13. Scheme for pyrochemical reprccessing of oxide and carbide fuels. R.I.C. Reviews 180 practice is to reduce their bulk by evaporation and store them indefinitely in large tanks. It has been shown that they can be adsorbed on clays which are then fired to produce a glassy product. The rate of leaching from this material is low and it can probably be dumped safely in ocean deeps. The fission products represent an increasingly important source of radio- active isotopes. 90Sr is extracted and converted to strontium titanate-a stable compound which is used as an energy source in thermoelectric generators, such as the Ripple 5 W generator which powers navigation lights in remote places.28 The design of fuel elements cannot proceed without a detailed knowledge of the physical and chemical properties of the fuel, the latter being important also in the development of fuel fabrication and reprocessing routes.The problems may be illustrated best by confining the discussion to those proper- ties of importance to the design of ceramic fuelled elements of the type used in the AGR and the SGHWR. Their design is a compromise between a number of factors. In particular the cladding must remain intact to burn-up levels at which the decrease in fissile atom concentration and the increase in fission product absorption cause a drop in power output. Neutron economy dictates that the cladding must be as thin as possible; therefore, it must be stressed as lightly as possible, i.e. fission gas release and swelling must be low, and corrosion losses should be small.These aspects are considered in more detail in the remainder of this review. Frost Fig. 14. Simplified flow diagram for fluid-bed fluoride-volatility processing of carbide core and blanket from a fast reactor. PHYSICAL AND CHEMICAL PROPERTIES Physical properties Gas release and swelling are temperature-dependent phenomena and, hence, a detailed knowledge of the thermal conductivity of fuel is essential in order to 181 13 h - I W M -0 '6 0.07 .- c, x .e 0.06 5 3 0 8 C.05 (d 0.04 - E E 182 calculate temperature distributions.29 U02 is an extrinsic semiconductor up to 1100-1200 "C, heat being conducted by phonon-phonon interactions. At high temperatures the electronic contribution becomes more important.The conductivity of U02 is low, typically 0.03 W cm-1OC-l at 1000°C, and this produces large temperature gradients between the surface and centre of fuel pellets which have a dominant effect on fuel performance. The conductivity varies with temperature (Fig. 25), and it is affected by irradiation below 500 "C. It has become an accepted practice, therefore, to relate fuel performance to the integrated thermal conductivity, rather than to attempt to calculate a temperature profile. The value of the integral is equal to the linear heat output of a fuel pin in W em-l divided by 4~ and its value where TI = 500°C and T2 = 2800°C (the melting point) is about 70 W cm-1 (see Fig. 26).30 The thermal conductivity of UC is between five and 10 times higher than that of U02 since it is an electronic conductor.31 The conductivity varies much less with temperature (Fig.17) and the Jk. d0 concept is therefore less import- ant. For similar heat outputs UC fuel rods experience much lower centre temperatures and thermal gradients and hence have different gas release and swelling characteristics. The effects of fabrication variables on the conductivity of UOz have been studied. The effect of the fractional porosityp is to decrease the value of k to k p according to the relation: Fig. 15. Thermal conductivity of irradiated stoicheiometric polycrystalline UOa. 0.09 Curve: 0.08 \ I. Saclay 2. Chalk river 3. AERE 4. AERE unirradiated 5.GE San JosC (based on f i t toORNL curve for unirradiate UOz at lower temperatures) 0.03 0.02 1600 1400 1200 1800 600 800 200 400 1000 Temperature ("C) R.I.C. Reviews I 80 70 60 I- 50 3 40 $ 5; In 30 20 10 0 500 I000 Fig. 16. Comparison-of integrated thermal conductivity out-of-reactor and in-reactor. where n has values between 1.5 and 4.0 depending on the shape and distribu- tion of the porosity. Oxygen in excess of an oxygen to metal ratio of two reduces the conductivity since the excess oxygen atoms act as impurity scattering centres. Single crystals are more conducting than polycrystalline material but they are usually metal rich and the effect of free uranium as a grain boundary phase is difficult to assess.The effect of irradiation on thermal conductivity has been studied by direct in-pile measurements in which thermocouples were placed in the fuel centre and along radii,32 by the comparison of pre- and post-irradiation measure- ments, and by the use of ‘markers’ in the fuel microstructure. Markers are grain growth and melting which occur at well-defined temperatures. Thus, from the irradiation of a pin with a known heat output in a known tempera- k, = k(l - np) Frost I I I T / Revised / ~ A E C L / / AProbzble range of ou t-of- dI I I I I :ztor E- San josC 3000 2500 2000 I500 Temperature (“C) 183 Sin tered (4.85% C 91% T.D.) 3 Cast 4.76% C (irradiated) 50.4/, 0.5-2 wt%O a) Arc cast 50.8% C (d) Oxycarbides b) Arc cast 51.7% C @) Arc cast 52% C @ A.C.cast 52% C @Cast UC (a) 52.5%C (bJ @ Cast 52.5% C + Absolute values 100 200 300 400 500 600 703 800 900 1000 1100 1200 1300 Temperature ("C) Fig, 17. Thermal conductivity of UC (close to stoicheiometric composition). ture environment (e.g. in a pressurized water loop at 300°C) the markers can be used to derive a thermal conductivity value. Below 500 "C the irradiation-induced decrease in conductivity is dependent on the temperature and burn-up; the damage becomes more stable as the burn-up increases. A major uncertainty in determining fuel temperatures is the thermal gradient through the filling gas in the gap between the fuel and the cladding. Often the gas is pure helium which becomes contaminated with fission gases during the irradiation, decreasing in conductivity and causing the fuel tem- perature to rise.Later (p. 198), it is seen that fission gases are retained within the fuel to a large extent at temperatures up to 1500"C, and form bubbles which cause the fuel to swell. In these lower temperature regions the fuel is fairly strong. Thus, although porosity may be present, the fuel will not deform by plastic flow into the porosity but will swell outwards and strain the cladding. The desire to remedy this situation has led to an intensive study of the mechanical properties of fuels out-of-pile and in-pile, with particular emphasis on creep processes.33 Both hypo- and hyperstoicheiometric U02 flow more readily than U02.000 and this has led to attempts to influence creep strength by the addition of 3- or 5-valent oxides to UO2-without marked success to date.The basis for this is the concept that vacancies play an important role in diffusion-type creep processes. This suggests that irradiation of fuel, which produces many vacancies, will produce an enhancement of the creep rate in those temperature regions where thermal effects are small, i.e. below 1000-1200 "C. In uranium carbide the variation of creep rate with composition under fixed load and temperature conditions is rather different.34 An increase in carbon R.I. C. Reviews 184 content leads to an increase in strength; since UC has a very narrow range of existence an increase in carbon content leads to the formation of a second phase of u2c3 or UC2 the particles of which may impede dislocation move- ment or lock grain boundaries.At low carbon concentrations uranium precipitates in the grain boundaries, improving grain boundary sliding, and within the grains. Quite small departures from stoicheiometry lead to changes in creep rate of one or two orders of magnitude. Chemical properties Lack of space prevents a full discussion of chemical aspects of fuel behaviour but some important aspects of the interaction of a fuel with its environment will be discussed. This is important first of all in fabrication processes. The flow and sintering characteristics of powders are affected by the nature of their surfaces and hence by the adsorption of water vapour and other gases; for example, oxygen is chemisorbed onto UOz at temperatures above -195°C with a maximum enthalpy of adsorption of 230 kJ These atoms are mobile at room temperature and above, and surface layers of higher oxides form readily. Uranium carbide oxidizes even more readily and, while it is possible to handle U02 powders at room temperature in fairly oxidizing atmospheres, UC must be handled in glove boxes containing an inert gas with a low water-vapour level.Under normal conditions the fuel inside a fuel pin is separated from coolants by the cladding. Commonly, this is zirconium in water-cooled reactors and stainless steel in COZ- or sodium-cooled reactors. Only when the cladding fails is the fuel exposed to the coolant; it is important that this does not then result in large quantities of fuel becoming entrained in the coolant.Under these circumstances we can rule out UC for reactors with water or C02 as coolant since clad failure will result in rapid oxidation of the fuel to u308 with a large volume increase and progressive splitting of the can until the us08 powder falls from the can. UOZ, on the other hand oxidizes more slowly. In fact, in water reactors the reaction rate at -350OC is so low that reactor operation can often continue after a fuel element failure has been detected, although this is highly dependent on the oxygen level in the water.5 In sodium, U02 and U O Z - ~ are stable. With U02+$ the excess oxygen atoms dissolve in the sodium; this can often lead to the disintegration of UOefz pellets in a sodium environment.Similarly UC and UC1-% are stable in sodium with a low oxygen content (typically 10 ppm in fast reactor circuits) but in UCl+$ the UC2 needles are decomposed by sodium to give UC and carbon. If a carbon ‘sink’ is available (e.g. stainless steel or zirconium) the carbon is transferred via the sodium to that sink (Fig. 18).36 The compatibility of fuel with its cladding is, of course, a question of thermo- dynamics and kinetics. Thus UOZ is thermodynamically less stable than zirconium but this combination is regularly used in water reactors, including the UKAEA’S Steam Generating Heavy Water Reactor at Winfrith, because the kinetics at normal operating temperatures are very slow.Experience shows that it is difficult to design fuel-cladding combinations from first principles and it is usually necessary to study reaction rates with a number of fuel-metal Frost 185 I300 h U 0, 1200 2 c U a I- 6 I loo I000 Fig. 18. Carbon contents of Type 304 and Type 410 stainless steels in equilibrium with carbon- saturated sodium. combinations. Generally a fuel specimen is sandwiched under pressure between two pieces of cladding material, held at a constant temperature and examined by sectioning followed by optical microscopy and by an electron microprobe, to determine reaction rates and the nature of the reaction products. In this field and in the development of improved fuels phase diagram studies have some importance.Since we are concerned with ceramic fuels with two species of atom, the ternary diagram is usually the simplest one studied-four- and five-component systems often require investigation. The melting points of U02 and UC are so high that the preparation of homogeneous alloys and the determination of liquidus and solidus temperatures demands the use of special high temperature techniques including refractory metal resistance furnaces, arc-image and electron-beam heating devices, tungsten/tungsten-rhenium thermocouples and optical pyrometry. X-ray diffraction studies, both of lattice parameters to determine phase boundaries and of crystal structures, play a vital role in this work. Finally, we are interested in the stability of the situation within an operating fuel; will the large thermal gradient cause changes in the distribution of the components? Markin and Rand37 have recently examined this situation for U02 and (U,Pu)Oz where a thermal gradient of 103°C cm-1 is not unusual.They showed that the presence of small quantities of carbon within the fuel (as little as 10 ppm, which is below the normal level) leads to the formation of a CO2/CO gas which is then able to transport material along the thermal gradient. Thus it is assumed that a constant CO&O ratio exists across the 186 Carbon content of steel (yo) R.I.C. Reviews 0 200 z h 3 4 0 0 IU- P 600 800 I .90 To vacuum system Constriction sealed before equil i brat ton I .95 Furnace Reference metal metal oxide Cooling water Heat shield Uranium oxide in crucible R.F.coil Drain Optical flat 117L-__ Pyrometer “Black body” hole 0:U ratio Fig. 19. Variation of ACoa with 0 : U ratio at 2200 K. Fig. 20. (a) Gas equilibration apparatus. (b) Oxygen distribution across a U0.85Puo.1502fy fuel pin for various COz : CO ratios. 2.05 O 2.00 I .95 I .92 Centre 2.00 1930 1765 1575 2.10 Frost I I ? .- U 0 e 1 I 1 I 2.05 Temperature (K) 1345 I I r2x 1,02(crn)2 cpo2 - I ratio I .982 I 1 2 3 4 5 6 Sukace 187 fuel. The oxide composition in equilibrium with a given CO2/CO ratio at any given temperature can be obtained from an Ellingham diagram; for a particular temperature distribution, the oxygen to metal (0 : M) ratio throughout the fuel can then be plotted.Results for a ‘mixed’ oxide fuel containing 15 per cent Pu and 85 per cent U are plotted in Fig. 19. An oxide with O:M fi 2.00 shows a negligible oxygen gradient, whereas deviations from stoicheiometry produce large oxygen gradients. In turn this alters the U:Pu ratio across the fuel; material with a high initial 0: M ratio will build up a high oxygen level towards its centre, vapour of composition uo3 will transfer uranium down the gradient leaving the plutonium enriched at the centre, possibly resulting in still higher temperatures. Autoradiography of irradiated fuel cross sections qualitatively confirms these predictions. To carry out this type of calculation it is necessary to know the value of the oxygen potential ACh2 and its tempera- ture coefficient.These have been measured by a galvanic cell technique using a solid electrolyte, reversible to oxygen ions only, e.g. Tho2 + Y203, or by a gas equilibration technique (H20/H 2 or C02/CO) using a reference metal/ metal oxide mixture to fix the oxygen potential (Fig. 188 2 Secondary knock-on path 3 Tertiary knock-on path H)I Intense ionization X Interstitial 0 Thermal or displacement Fig. 22. Five mechanisms of radiation effects. Represented are intense ionization vacancies, interstitials, impurity atoms and thermal or displacement spikes. Grid-line intersections are equilibrium positions for atoms. Fig. 23. Fission fragment tracks in vacuum-deposited UOz film 23 nm thick irradiated with 5x 1015 slow neutrons cm-2. The mean grain size is about 3 nm diameter.Frost 189 I o3 I o2 IRRADIATION BEHAVIOUR The irradiation behaviour of a fuel determines its performance in the reactor and is therefore of paramount importance. When a uranium or plutonium atom undergoes fission it splits into two fission fragments having atomic weights between 72 and 161, each atom having an initial energy of nearly 100 MeV. In addition, between two and three neutrons are produced, each with an initial energy of several MeV (Fig. 21). Both fission fragments and neutrons move rapidly through the lattice, exchanging their energy with the lattice atoms until they come to rest (Fig. 22). The energy carried by the fission fragments predominates and within the fuel we can probably neglect the neutrons.Each fission fragment is highly ionized (about 20+) and has a range of about 7-10 pm in fuel materials (Fig. 23). Over this range it excites atoms up to 100 A radius from the track centre, equivalent to raising their temperature to thousands of degrees locally; this is known as a 'fission spike'. Within each spike a number of atoms are displaced from their lattice positions to produce single vacancies and interstitials. If the temperature is high these recombine. If it is low they remain as single defects and have a large effect on transport processes such as thermal and electrical conductivities. At intermediate temperatures they may cluster and collapse into loops.Such effects are ob- servable in a number of ways, e.g. by x-ray lattice parameter measurements, by transmission electron microscopy and by resistivity measurements. However, they only have intrinsic practical importance in relation to their effect on thermal conductivity. They are also significant in relation to gaseous fission M h . 10' . E" v I .; a c L 0' n 2 to-' lo-; to-' 190 Zr Fig. 24. Mass distribution of fission products from thermal fission of 235U. R. I. C. Re views I00 I 80 60 40 30 .- : z 20 g - aJ x c, : 10 ? 3 m al .? 6 - !! .- aJ x $ 4 .- C 3 U L (d w .- E TI $ 2 U In I 0.8 0.6 0.4 1 I I 80 90 Fig. 25. Relative fission yields for 2351) and z39Pu.0.3 0.2 70 product behaviour and to diffusion processes. In the latter context, diffusion is controlled by vacancies and fission produces an abundant supply of these. It is possible therefore that such processes as creep and sintering may be enhanced while fission is proceeding. Once the fission fragments have come to rest. thev are more usuallv described in Fig. 24. Furthermore, they are radioactive and undergo decay processes, and Frost I I I I I _----- 1 Pu-239 thermal neutron' spectrum U-235 thermal neutron spectrum 150 --- I 100 130 160 110 140 5 I20 Mass number 191 they may capture neutrons so that the prediction of their concentration at some arbitrary point during irradiation is difficult.There are some significant differences between the yields from uranium and plutonium fission (Fig. 25). Studies of fission products have tended to concentrate on the stable species, that is those present in fuels some weeks after fission has ceased, since it is generally inconvenient to study fuel while it is in a reactor. An exception is the ‘swept capsule’ type of experiment in which an inert gas is used to entrain the volatile fission products from a fuel sample in pile to suitable detection equipment. The abundant fission products may be grouped conveniently according to their physical and chemical characteristics. Noble metals Mo, Tc, Rh, Ru, Pd. At high oxygen potentials Mo may exist as Moo2 or Moo3 Cs, I, Br, Te. Some of these may be Readily oxidized but insoluble in U02 BaO, SrO Readily oxidized but soluble in UOZ Zr, Ce, Nd Volatiles present as caesium halides Xe, Kr Stable gases The detailed study of fission products is important for several reasons: (a) they must be removed and stored or else used as tracers or heat sources, (b) they cause the fuel to swell, (c) the gases if released create a pressure, and ( d ) some may react with cladding materials.In terms of fuel element behaviour, swelling and gas release are the most important effects. The ‘basic’ swelling of a fuel is the nett volume increase incurred when all the fission products are atomically dispersed but are able to attain their equili- brium oxidation states. In an oxide fuel, solution of zirconium and the rare earths in the lattice causes a volume decrease while major increases arise from caesium, molybdenum, the noble metals, barium and strontium.Davies39 and Wait40 have derived a basic swelling rate of 0.5 per cent volume increase per 1 per cent burn-up while Anselin and Baily4l using different assumptions derive a value of 0.30 per cent AV/V. Davies’ corresponding figure for carbide is 0.9 per cent due to the large contributions from carbide-forming Mo and Ru and from Cs. To derive these figures it was obviously necessary to determine the oxidation states of the fission products and this was done by referring to the published thermodynamic data. The results are sensitive to the oxygen potential of the fuel, i.e. they will differ as the O:M ratio varies.The general conclusion is that the O:M of the fuel will rise as burn-up proceeds. In irradiated fuel elements the distribution of fission products is not random; the volatile fp’s (particularly Cs) migrate down the temperature gradient, the noble fp’s agglomerate to form a precipitate of second phase visible in the optical microscope (Fig. 26) and the gases, being insoluble and fairly mobile, agglomerate to form bubbles. A great deal of work has been done and continues to be done to study the distribution of fp’s in irradiated fuel elements and a number of techniques is available : 1. Autoradiographs of fuel sections. 2. Gamma spectrometry; the fuel is scanned by a counter enclosed in a R. I . C. Reviews 192 Fig.26. Agglomeration of noble fission products as seen through a microscope. Fig. 27. Caesium movement in (U, Pu)Oz. Frost 193 heavy-metal collimator and connected to a multi-channel analyser. It is possible to scan the cross-section of fuel pin in -1.0 mm steps for various isotopes, the results being printed out as a contour map or a coded colour pattern (Fig. 27). 3. Small diameter corings may be drilled from across the radius of a fuel pellet and subjected to radiochemical analysis. 4. The electron microprobe analyser has been used to an increasing extent to study segregated fission products. Either a shielded instrument is used or, as in the pioneering work of Bradbury et ~ 1 . ~ 4 2 the specimen activity is minimized by using a thin polished section.The majority of the inclusions in oxide fuel contain the ‘noble’ metals Mo, Ru, Rh, Tc and Pd (Fig. 28). A minority contain Ba, Zr, Sr and Ce; the presence of zirconium is unexpected since it dissolves in the oxide lattice. It is possible that it is present as barium zirconate. High burn-up oxide fuels in which the centre has been molten during irradiation usually contain ingots of the noble fission products at the bottom 250 I x .- n - v) v v) k‘ c U - C aJ 0 16’02’ Fig. 28. Electron probe microanalysis shows the inclusion of ‘noble’ metals in oxide fuel elements. 194 Ru I2O50’ Bragg angle R.I. C. Reviews of the shrinkage void which forms on freezing. The melting point of these ingots has been found to be about 1850°C.However, it is the gaseous fission products which dominate fuel element per- formance. Ten per cent by weight of the stable fission products are accounted for by xenon and krypton. Stated in another form, 24 cm3 of gas at NTP are formed in each gram of UOZ after 1 per cent burn-up. It can easily be seen that if all of this is released into the fuel element can a high pressure will build up unless precautions, such as adding a large plenum to the end of the element, are taken. The importance of predicting fission gas release has led to an intensive study of the kinetics and mechanisms of gas release as a function of temperature, composition and burn-up. In rather oversimplified terms, the presently understood position is that at temperatures below 1000 "C gas atom migration rates are very slow and any gas release occurs by 'recoil' (a fission event near to the surface produces a gas atom or its daughter and this escapes from the surface) or by 'knock-out' when a fission product collides with a static gas atom and knocks it out of the fuel.Between 1000 "C and 1600 "C gas atom mobility increases to significant values and some gas is able to diffuse to surfaces during the lifetime of the fuel element. Gas diffusion is controlled by the usual diffusion equation: D = DO exp (- Q/kT) During irradiation the fractional release of gas, F, is given by9 where t is the irradiation time and S/V the surface area:volume ratio of a particle of radius a, known as the equivalent sphere, and usually found by BET surface area measurement.A common method of studying gas release has been to measure the gas evolved from an irradiated fuel sample when heated in the laboratory. Under these conditions44 F = 6 d $ - ; ; i 3 Dt - 3D't where D' = - D a2 D' is the apparent diffusion coefficient. For U02 of 95 per cent theoretical density D has a value at 1400°C of -5 x lO-I5 cm2 s-1 and the activation energy Q, derived from measurements over a range of temperature, has a generally accepted value of 293 21 kJ mol-l. For UC D is about an order of magnitude smaller (-5 x 10-16 cmz s-l) and the activation energy is about the same (Fig. 29). This type of analysis has had some success but anomalies have appeared which, in general, indicate some hindrance to diffusion.Often called 'trap- ping',46 this is probably due to two processes acting in addition to atmoic diffusion. Firstly, atoms will agglomerate to form stable bubbles (Fig. 30), the motion of which will differ from that of atoms and, secondly, gases become trapped at grain boundaries and when cracking occurs-as at a reactor shut- 195 Frost - \ 3 ( - 14 - 15 - 1 6 - N n E - ? 0 - 17 - t8 - 19 - 20 9 8 5 6 7 Temperature I / T ( K x lo4) Fig. 29. Diffusion of lssXe from uranium carbide. Fig. 30. Aggtomeration of atoms to form stable bubbles. R.I.C. Reviews 196 10 I00 h v - G 1.0 0. I c s 2 E vl W 0.0 I 900""l'OOO down or start-up-these are suddenly released.This latter phenomenon has been called the 'burst' effect in some experiments. Recent work by Whapham47 and others has shown that even this picture is too simple since the fission process can cause bubbles to break up and the gas disperse into the lattice. Under steady powder conditions a large release of grain boundary gas is delayed until the bubbles touch one another. Above 1600°C grain growth begins to be significant in U02; up to 1800- 2000°C grain boundary sweeping will enhance the rate of gas release. Above 2000°C the bubbles will grow rapidly and move up the temperature gradient by evaporation from the hot face and condensation on the cold. We can simplify prediction by assuming that all the gas is released above 1600°C (Fig. 31).Analogous information on carbide is lacking at present. The fact that the fission gases form bubbles is significant in relation to fuel swelling. Below 1000°C the contribution of bubbles to swelling is small and above 1600 "C most of the gas has been released. In the intermediate region bubbles become important. The swelling of a fuel sample after burn-up b is: Fig. 3 I . Gas release from U02 as a function of fuel-cell temperature. Frost 14 I 100 I200 I300 1400 1500 1600 1700 1800 1900 2000 2100 2200 Fuel centre temperature ("C) -- *'- Sb + 4 p r - r3 3 V 197 where S is the solid fp swelling and p the number of bubbles of radius r. We need to evaluate p and r. Greenwood et aL48 derived an expression for = ( 8n'u2r~Dg) 3p l I 2 where /3 is the gas generation rate, Dg the gas diffusion coefficient, ro the radius of bubble nucleus (a few lattice spacings) and a the lattice parameter.When a gas atom comes to rest in the fuel lattice, it strains the lattice. This strain can be relieved if vacancies flow to the gas atom. During irradiation vacancies are plentiful and the internal gas pressure p is balanced by the surface tension force y in a bubble of radius r such that 2Y P", p must also satisfy the modified gas law: p v = nc.- kT N where n is the number of gas atoms per bubble and N is Avogadro's number. Thus we should be able, for any given temperature and irradiation condition, to calculate p and r.49 In practice the temperature varies across the fuel section and this has an effect on both p and r ; more bubbles are nucleated near the centre and their growth rate there is greater.Bubbles can migrate by a variety of mechanisms depending on their size- small ones may exhibit a Brownian motion or they may move by surface or volume diffusion until trapped by a dislocation or a grain boundary. When the bubbles grow larger (>65 nm radius) they can break away from dislocations and move up the temperature gradient until they meet a grain boundary. Above 450 nm radius the temperature gradient driving force exceeds the grain boundary force and either the bubble escapes or it will drag the boundary with it, giving rise to long columnar grains. The situation in an operating fuel pin of UOz is illustrated in Figs 32a & b which show schematically a typical microstructure together with a plot of bubble velocity versus radius. Up to -1500°C the original grain structure remains.At the cooler rim of this region the gas bubbles are small and at the inner edge they are larger and more numerous but probably less than 65 nm diameter. Above -1500°C grain growth begins to be measurable and gas bubbles reach grain boundaries fairly rapidly, where they link up and form a path for rapid release. Above about 1750 "C the bubbles have exceeded 450 nm radius and they move up the gradient forming columnar grains and removing all volatile fission products rapidly, together with the initial sinter pores which form a cavity at the centre of the fuel. Below 1500°C the bubbles persist and contribute to the fuel swelling.The larger bubbles at the centre contribute more than those at the edge and a stress gradient is set up-compressive at the centre and tensile at the edge. The net swelling is determined by the bubbles at the region of zero stress which is about two-thirds of the distance from the centre of the region (Fig. 33).50 198 R.I.C. Reviews Expressed more quantitatively, the contribution to swelling in ceramic fuels of the fission gases is of the order 0.5-1.0 per cent AV/Vper 1 per cent burn-up (Fig. 34). This must be added to the solid fission product value of between 0.25 and 0.5 per cent A V/V to give a maximum swelling of 1.5 per cent A V/ Y per 1 per cent burn-up for oxide and about 2.0 per cent AV/V for carbide.Experimental values derived from diameter changes in fuel pins are generally Centre temperature Onset of columnar grain growth Onset of equiaxed grain growth Original sintered grain structure persists Gas bubble behaviour in pellet Schematic view of highly rated UO, pellet (b) Fig. 32. (a) Cross-section of irradiated UOz pellet (courtesy 1. nucl. Mater.30); (b) schematic view, showing gas bubble behaviour. Frost 199 14§ 0.0 I 0 - 0.01 - 0.02 - 0.03 a h - a I m - 0.04 t -0.05 -0.06 v v -0.07 - 0.08 I - 0.09 -0.1 -0.1 I t -0.12 Fig. 33. Non-uniform swelling stress in a fuel pellet. *: 6 1 . 1 0 1 . 1 2. I 3.2 r4 Com press ion 4.3 6 Z 5.3 .a 0, 6.4 x ul 7.5 ; 8.5 9.6 10.6 11.7 12.7 10 9 I 7 ! 8 1 lower than this, which is not surprising as the claddingand the external coolant pressure tend to exert a restraining influence on bubble growth.Fast reactor fuels should be capable of attaining a burn-up in excess of 5 per cent heavy atoms and preferably 10 per cent to give an acceptable fuel cycle cost. To achieve this it must be possible to contain fuel swelling of the order of 10 per cent or more within the cladding without failure. If voidage is provided within the fuel, whatever the form (central hole, sinter pores, cavities in packed powder) it is unable to accommodate the swelling efficiently because the fuel cannot deform sufficiently in the cold regions below 1000-1200 "C.Thecladding, which is generally stainless steel, is itself impaired by the fast neutron irradia- tion being unable to sustain more than about 1 per cent creep strain.51 There is, therefore, a considerable effort being applied to the problem of altering the fuel properties or its geometry relative to the can, in order to accommodate swelling more effectively. As an example, one approach is to leave a generous gap between the fuel and its can, and to fill the gap with sodium. This acts as a good conductor of heat, keeping the fuel cool, and may be displaced to a plenum or reservoir as the fuel swells. Since the fuel cycle cost is a major component of the power cost in fast reactors the potential cost benefit of such developments is very considerable.However, it should be recognized that fuel and fuel element irradiation studies are very expensive. There are many steps in the development of a fuel element to the stage where it is acceptable to reactor constructors and their customers. The first steps are Fig. 35. Very high burn-up fuel particle testing rig. Frost 20 1 concerned with evolving a fabrication method and measuring important properties out of pile, as discussed earlier. Then two types of in-pile test may be used; first, short lengths of fuel element, typically 5-15 cm long, are made with realistic radial dimensions to establish the correct temperature profiles and are irradiated in a materials testing reactor (MTR), such as DIDO and Fig. 36. Detailed examination of irradiated pins.R . I.C. Reviews 202 PLUTO at Harwell, under carefully controlled and monitored conditions (Fig. 35).52953 This establishes whether there are any important problem areas and gives preliminary data on swelling and gas release. Detailed examination of the irradiated pins is performed in shielded cells, usually with a cell devoted to each type of measurement (mensuration, fission gas release, optical and electron microscopy, fission product distribution etc.) (Fig. 36). Secondly, the kinetics of fission product release are measured in a swept capsule rig in an MTR.54 A small sample of fuel is held inside an electric heater at a fixed temperature while helium flows over it; the gases are entrained to a Fig. 37. In-pile assembly. Frost 203 counter and the volatiles to a metal surface which is subsequently removed for analysis (Fig.37). Having established the fuel performance on a small scale, full-size prototype fuel elements are made into clusters and, where possible, tested in prototype power reactors either as part of the main ‘driver’ charge or in special loops to test failure characteristics. The essential point is that statistical information must be amassed because in the first place there must always be some latitude in manufacturing specifications and in the second place conditions vary considerably from point to point in a reactor core in terms of temperature and neutron flux and spectrum. Consequently, small scale testing may give misleading information.The examination of irradiated fuel pins on a statistical scale plus the careful analysis of the results is a demanding and costly operation but nevertheless it is vital to the successful exploitation of any reactor type. CONCLUSION Much of the engineering and plant aspects of nuclear reactors is based on conventional engineering and chemical engineering practice. The fuel cycle is a unique but vital part of the system demanding intensive research and develop- ment at every stage from the ore to reprocessing of the irradiated fuel. The nature of the work makes it expensive, particularly in?erms of capital facilities such as test reactors, hot cells, glove boxes and remote operations. However, the rewards are potentially high and may be measured in terms of savings of hundreds of millions of pounds in power programmes.The approach in the UKAEA to these problems is broadly based with the preliminary exploratory work and the basic research on mechanisms being centred on the Research Group at Harwell. The development of fuel element designs and statistical testing is the responsibility of the Reactor Group in a number of laboratories, and the production of fuel elements and their re- processing is the responsibility of the Production Group. This review has been concerned primarily with expounding the general principles behind this work rather than with descriptions of detail. The interested reader is referred to other works for this detail, and particularly to the annual reports of the UKAEA.REFERENCES 1 Report of the Select Committee on Science and Technology, The UK nuclear reactor programme, xxxvii, London: HMSO, 1967. 2 G. F. Tape, 3rd International Conference on the Peaceful Uses of Atomic Energy (ICPUAE-3), Geneva 1964, 1, paper P192, 69. 3 P. D. Dunn et al., Nature, Lond., 1962, 195, 65. 4 J. T. Ramey et al. ICPUAE-3, Geneva, 6,428. 5. J. Belle (ed.) Uranium dioxide USAEC, 1961. 6 B. R. T. Frost et al. ICPUAE-3, Geneva 1964. Paper P153. 7 R. A. U. Huddle and L. R. Shepherd, IAEA Conference on New Nuclear Fuels, Prague 1963, 2,467. 8 W. D. Wilkinson, Uranium metallurgy. New York: Interscience, 1962. 9 UKAEA Ann. Rept No. 11, para. 202; No. 13, para. 252; and No. 14, para. 223. 10 ICPUAE-1, Geneva 1955. Vol. 8. 11 S. A. Cottrell et al., ICPUAE-3, Geneva 1964. Paper P150. 12 S. Naymark and C . N. Spalaris, ICPUAE-3, Geneva 1964. Paper P233. 13 ICPUAE-2, Geneva 1958, 6, 569-629. 14 J. E. Ayer and F. E. Soppet, J. am. ceram. Soc., 1965, 48, 180; 49, 207. R.Z. C. Reviews 204 15 L. E. Russell and J. D. L. Harrison, ENEA Symposium on Reactor Materials. Stockholm, 1959. 16 F. Brown et al., Carbides in nuclear energy (ed. L. E. Russell), vol. 10, 692. London: Macmillan, 1963. 17 R. Ainsley et al., ibid., 540. 18 J. D. L. Harrison and J. W. Isaacs, ibid., 556. 19 J. D. L. Harrison et al., ibid., 629. 20 K. M. Taylor et al., ibid., 668. 21 H. Pearlman and R. F. Dickerson, ICPUAE-3, Geneva 1964. Paper P234. 22 J. P. McBride et al. 0 ~ ~ ~ - 3 8 7 4 , 1966. 23 M. Allbutt and R. M. Dell, J. nucl. Muter., 1967, 24, 1. 24 G. R. Howells et al., ICPUAE-2, Geneva 1958, 17, paper P307, 3. 25 ICPUAE-3, Geneva 1964,lO. 26 L. Burris et al., ICPUAE-2, Geneva 1958, 17, paper P538, 401. 27 S. Lawroski, Chem. Engng. Prog., 1955’51,461. 28 F. W. Yeats, Atom, Lond., December 1966, no. 122,282. 29 IAEA Technical Reports Series No. 59, Thermal conductivity of uranium dioxide, Vienna 1966. 30 J. A. L. Robertson, A. M. Ross, M. J. F. Notley and J. R. MacEwan, J. nucl. Muter., 1962, 7, 225. 31 J. A. Leary, R. L. Thomas, A. E. Ogard and G. C. Wonn, Carbides in nuclear energy (ed. L. E. Russell), vol. 1, 365. London: Macmillan, 1963. 32 D. J. Clough and J. B. Sayers. ~~1~3-R.4690, 1964. 33 W. M. Armstrong and W. R. Irvine, J. nucl. Muter., 1963, 9, 121. 34 J. J. Noreys, Carbides in nuclear energy (ed. L. E. Russell),vol. 1,435. London : Macmillan, 1963. 35 J. D. M. McConnell and L. E. J. Roberts in Chemisorption (ed. W. E. Garner), 218. London: Butterworths, 1957. 36 B. A. Webb, North American Aviation Report NAA-SR-6246, 1962. 37 M. H. Rand and T. L. Markin, ‘Some thermodynamic aspects of (U,Pu)Oz solid solu- tions and their use as nuclear fuels’, A E R E - R . ~ ~ ~ ~ , 1967. 38 T. L. Markin, ‘Thermodynamic data for U02 and (U,Pu)Oz applied to fuel preparation problems’, mrn-R.5538, 1967. 39 J. H. Davies, unpublished work. 40 E. Wait and B. R. T. Frost, IAEA Conference ‘Plutonium as a reactor fuel’, Brussels 1967, paper SM-88/25, Proceedings, 469. 41 F. Anselin and W. E. Baily, Trans. Am. nucl. Soc., 1967, 10, 103. 42 B. T. Bradbury, J. T. Demant, P. M. Martin and D. M. Poole, J. nucl. Muter., 1965,17, 227. 43 J. I. Bramman, R. N. Sharpe, D. Thorn and G. Yates, J. nucl. Muter., 1968, 25, 201. 44 A. H. Booth, Canadian report CRDC-721, 1957. 45 D. Davies and G. Long, AERE-R.4347, 1963. 46 J. R. MacEwan and W. H. Stevens, J. nucl. Muter., 1964, 11, 77. 47 A. D. Whapham and B. E. Sheldon, 6th International Conference on Electron Micro- scopy, Kyoto, Japan, 1966. Proceedings, 375. 48 G. W. Greenwood, A. J. E. Foreman and D. E. Rimmer, J. nucl. Muter., 1959, 1, 305. 49 R. S. Barnes and R. S. Nelson, AErn-R.4952, 1965. 50 B. L. Eyre and R. Bullough, J. nucl. Muter., 1968,26, 249. 51 P. T. Nettley et al., British Nuclear Energy Society Conference on Fast Reactors, London 1966. 52 0. S. Plail, NucE. Pwr, December 1960. 53 N. H. Hancock, ~~m-R.4156, 1966. 54 G. Jackson, D. Davies and P. Biddle, A E R E - R . ~ ~ ~ ~ , 1966. Frost 205

ISSN:0035-8940    

DOI:10.1039/RR9690200163

出版商:RSC    

年代:1969

数据来源: RSC