JOURNAL OF ANTI-AGING MEDICINEVolume 2, Number 3, 1999Mary Ann Liebert, Inc.

Free Radical Theory of Aging: Increasing the AverageLife Expectancy at Birth and the Maximum Life Span

DENHAM HARMAN, M.D., Ph.D.

ABSTRACT

Continued improvements in general living conditions—e.g., better nutrition, medical care,and housing—during the past two millennia have increased average life expectancies at birthfrom about 30 years in ancient Rome to almost 80 years in the developed countries with no

change in the maximum life span. Current average life expectancies at birth will be increasedlittle by further improvements. The rate of accumulation of damage inflicted on us by our in-herent aging process limits average life expectancy at birth under optimal living conditionsto around 85 years and the maximum life span to about 122 years. The inherent aging processis caused by chemical reactions that arise in the course of normal metabolism. Attempts tosignificantly increase average life expectancies at birth and the maximum life span in the fu-ture, unlike in the past, will require an understanding of aging. The free radical theory of ag-ing postulates that this process is caused by free radical reactions, largely initiated by super-oxide radicals arising from the mitochondria at an increasing rate with age. Some measures

based on the free radical theory of aging may further increase the life span without interfer-ing with the activities of normal life include: (a) caloric restriction, (b) compounds that de-crease O2 access to "electron-rich areas" of the mitochondria, and (c) substances that help tominimize mitochondrial damage. The foregoing are discussed briefly along with the amelio-ration of damaging reactions in early life that predispose to life-shortening diseases. The fea-sibility of the measures suggested above needs to be evaluated. This task should be both in-teresting and rewarding.

IINTRODUCTION improvements in living conditions. These are

now approaching optimum levels in the de-NTEREST in increasing the maximum life span veloped countries, while average life ex-

is a recent concern. During the past two mil- pectancies at birth are nearing plateau values,lennia average life expectancy at birth in- Past efforts to increase average life ex-

creased from about 30 years in ancient Rome pectancies at birth did not require an under-to almost 80 years today in the developed coun- standing of aging. Such knowledge will be nec-

tries; apparently maximum life expectancy has essary if average life expectancies at birth andremained unchanged. The increase in average the maximum life expectancy are to be signifi-life expectancy at birth is the result of gradual cantly increased in the future.

Department of Medicine, University of Nebraska College of Medicine, Omaha, Nebraska.

199

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200 HARMAN

AGING

Phenomena

Aging is the accumulation of diverse adversechanges, i.e., aging changes,1,2 in the cells andtissues that collectively increase the chance ofdisease and death.3 The chance of death of anindividual of a given age in a population—readily available from vital statistics data—serves as a measure of (a) the average numberof aging changes accumulated by persons ofthat age and (b) physiologic age. In the past,3improvements in general living conditions in a

population have been associated with de-creases in the chances for death and, in turn,increases in average life expectancies at birth.These conventional measures, e.g., better nu-trition, housing, and medical care, publichealth facilities, and accident prevention, are

becoming increasingly futile.3As living conditions in a population ap-

proach the optimum, and premature deaths a

minimum, the curve of the logarithm of thechance of death versus age shifts toward a

limit4-7 determined by the sum of (a) the irre-ducible contributions to the chance of death ofchanges that can be prevented to varying de-grees by conventional measures, e.g., those dueto the environment and disease, and (b) con-tributions that can be influenced little, if at all,by conventional measures, i.e., those due to an

inherent process(es), termed the "agingprocess." The now near-limiting chances fordeath rise almost exponentially after age 28:only 1-2% of a cohort die before this age.As chances for death approach limiting val-

ues in the developed countries, the associatedaverage life expectancies at birth rise towardplateau values4"7 of around 76 years for malesand 82 years for females. Average life ex-

pectancies at birth for the total population inthe developed countries are now around 6-9years less than the potential maximum valueof about 85 years8-10 that may be achieved byconventional measures. Optimistically, withcontinued application of improving conven-tional measures, the average life expectanciesat birth will eventually reach plateau values ofaround 80-82 years in the next 50 years or

more.3

Inherent aging processThe inherent aging process is the major risk

factor for disease and death in the developedcountries after age 28. It limits average life ex-

pectancies at birth of the populations, livingunder optimal conditions, to about 85 years8"10and the maximum life expectancy to around122 years.11 The contributions of this process tothe chances for death are small early in life butrapidly increase with age because of its expo-nential nature. Hence, few reach 100 years, andnone exceed about 122 years.The aging process is caused by chemical re-

actions that arise in the course of normal me-tabolism, which collectively, produce agingchanges that exponentially increase the chanceof death with advancing age even under opti-mal living conditions. To achieve future signif-icant increases of both average life expectanciesat birth and the maximum life span will requireknowledge of these reactions.Many theories12-19 have been advanced to

account for aging. No single theory is generallyaccepted.20-22 The importance attached to in-creasing the healthy, useful human life span be-yond the 3-5 years that may still be achievedby conventional measure dictates that agingprocess hypotheses be explored for practicalmeans of achieving this goal, while continuingto work toward a consensus.

FREE RADICAL THEORY OFAGING: ORIGIN

The free radical theory of aging3'17'23'24 is a

promising hypothesis. It, and the simultaneousdiscovery of the important, ubiquitous in-volvement of endogenous free radical reactionin the metabolism of biological systems, arosein 195423 from a consideration of aging phe-nomena and from the premise that a singlecommon process, modifiable by genetic and en-vironmental factors, was responsible for the ag-ing and death of aflliving things. The free rad-ical theory of aging postulates that the commonprocess is the initiation of free radical reactions.These reactions, however initiated, could be re-

sponsible for the progressive deterioration ofbiological systems with time because of their

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INCREASING THE MAXIMUM LIFE SPAN 201

inherent ability to produce random change.The theory was extended in 197225'26 with thesuggestion thatmost free radical reactions wereinitiated by the mitochondria at an increasingrate with age, while the life span is determinedby the rate of free radical damage to the mito-chondria. Collectively, the free radical reac-

tions initiated by the mitochondria constitutethe inherent aging process.The free radical theory of aging suggests that

conventional measures increase average life ex-

pectancies at birth by decreasing the rate of ac-cumulation of aging changes. For example, (a)better nutrition providesmore compounds, e.g,vitamins C and E, and ¿8-carotene, to decreasefree radical damage, (b) in the case of diseases,free radical reactions are widely involved17'23'24so that measures to improve prevention andtreatment would be expected to have beneficialeffects on life span, (c) housing improvementsmay decrease the amount of food metabolized,and hence free radical production, needed tomaintain body temperature, and (d) tissue in-jury causes free radical formation, thus betteraccident prevention measures should reducethe accumulation of aging changes.The free radical theory of aging also suggests

that measures to decrease the rates of initiationand/or chain lengths of free radical reactionsmay decrease the rate of aging and of diseasepathogenesis. Many studies now support thispossibility.3'23'24Most animal studies have useddietary antioxidant supplements to decreasechain lengths; these augment the natural de-fenses against free radical reaction damage. Forexample, addition of 1% by weight of 2-mer-captoethylamine hydrochloride to the diet ofmale LAFi mice, started shortly after weaning,increased average life expectancy at birth by30%;27 the maximum life span was increasedlittle, if at all. When 0.5% of ethoxyquin was

added to the diet ofmale and female C3H mice,the increase in the average life expectancy atbirth was 20% for both groups with no changein the maximum life spans.28 Thus, antioxidantsupplementation in mice decreases both pre-mature deaths and senescence periods, butdoes not increase the maximum life span.The above results in mice have been mim-

icked since 1960 in the U.S. population.29 Dur-ing this period, the percentage of the popula-

tion that take antioxidant supplements hasgrown from probably a fraction of 1% to40-50%. The average life expectancy at birthrose from 69.7 years in 1960 to 75.4 years in1990 and to 75.7 years in 1974. Increases in theaverage life expectancy at birth were associatedwith relative increases in the size of the olderpopulation. Thus, between 1960 and 1990 thenumber of individuals aged 65 years or older(the elderly) grew by 89%, those age 85 andolder by 232%, and the total population in-crease by 39%. Chronic disability among the el-derly has been declining since 1982, cancer in-cidence started to decline for the first time in1991, and the continuing decline in cardiovas-cular disease became greater after 1965. Theforegoing changes are due to the joint action ofimprovements in conventional measures andthe increasing use of antioxidant supple-ments/improved nutrition. It is reasonable toexpect on the basis of extensive animal and epi-demiologic studies that direct efforts to de-crease free radical reaction damage over thepast 40 years (sales of antioxidant supplementsare now $4—5 billion per year) have contributedsignificantly to the above changes in the U.S.population.

MAXIMUM LIFE SPAN

Three antioxidants have been reported to in-crease the maximum life spans of mice: 2-mer-captoethanol (2-ME)30 and two pyridine com-

pounds.31,32 Apparently, these studies have notbeen repeated. The study with 2-ME is the onlyantioxidant experiment reported in which foodconsumption and body weights of both con-

trols and treated mice were maintained thesame; addition of 0.25% by weight of 2-ME tothe diet of BC3Fi mice increased average lifeexpectancy by 13% and maximum life span by12%. Thus, 2-ME increased life span in the ab-sence of the confounding influence of the com-

monly lower body weights of mice receivingantioxidants.The general failure of antioxidants to in-

crease maximum life span is attributed to3 (a)depression of mitochondrial function by thecompounds at concentrations below thoseneeded to slow free radical damage to the mi-

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202 HARMAN

tochondria (i.e., as the dietary concentration ofan antioxidant is increased it significantly im-pairs mitochondrial function before it slowsmitochondrial aging—2-ME and the two pyri-dine compounds discussed above may be ex-

ceptions), and/or (b) progressive increases inthe initiation of free radical reactions with ageby the mitochondria eventually nullify the ben-eficial effects of the added antioxidant. Failureof antioxidants to increase the maximum lifespan led to the suggestion in 1972 that mito-chondria determined the life span.25,26Cause

The innate life span is determined by the rateof the increase in production of Superoxide rad-icals with age by the mitochondria. Support forthis statement includes the following: (a) Themajor source of endogenous free radical reac-tions are Superoxide radicals arising from themitochondrial respiratory chain in the course

of normal metabolism.33 (b) The rate of mito-chondrial Superoxide radical formation in-creases with age.24,25,33^0 This makes it pro-gressively more difficult, and eventuallyimpossible, to prevent free radical damage tothe body by dietary means, including the useof antioxidant supplements, (c) Comparingbirds and mammals of similar metabolic rates,the much longer life spans of the birds is re-

lated to a lower rate of formation of Superox-ide radicals by the mitochondria.41,42 Likewise,in two closely related rodent species.43 (d) Lifespans of different mammalian species35 are re-lated to the rate of mitochondrial Superoxideformation, (e) The frequency of the mitochon-drial genotype, Mt5178A, in Japan is higher incentenarians than in healthy blood donors.44As expected, since mitochondria are of mater-nal origin, siblings of centenarians live longerthan normal.45 The high frequency of Mt5178Ain the Japanese population (45%) may be re-

lated to the fact that the Japanese are thelongest lived of the world populations. In ac-

cord with the foregoing, oxidative stress hasbeen reported to be lower in healthy centenar-ians than in younger individuals.46Decreasing mitochondrial Superoxide radi-

cal formation would increase3 both the maxi-mum average life expectancy at birth, now

about 85 years,8-10 as well as the maximum lifespan, now 122 years.11 Decreasing the adversechanges in the early period of life47 would alsoserve to increase both average life expectancyat birth and maximum life span.

Increasing maximum life spanMeasures that may decrease Superoxide rad-

ical formation without significantly loweringATP production include those listed in Table 1and are discussed here.

Caloric restriction. Decreases in caloric intakeare associated with proportionate decreases inO2 utilization. Over 90% of the O2 consumedby mammals is utilized by mitochondria; ofthis, a small fraction is diverted to form super-oxide radicals. Thus, food restriction may in-crease the life span by decreasing mitochon-drial free radical reaction initiation rates.3,17'24This provides the most parsimonious explana-tion of the numerous changes48 found in caloricrestriction studies.In accordance with the above possibility, de-

creasing the daily caloric intake of rats by 40%,while maintaining essential nutrients,49 de-creased body weight by 40%, increased aver-

age life span by 40%, and increased maximumlife span by 49%. The metabolic rate, i.e., thecaloric consumption per day per unit of bodyweight, was the same for the two dietarygroups—because the percentage decrease in

Table 1. Measures that May DecreaseMitochondrial Superoxide Radical Formation

Caloric restrictionCompounds that compete with O2 for electrons fromthe respiratory chainSpin-trapsNitroxidesHydroxylamines

Substances that block access to the electron-rich areasof the respiratory chainBuckminsterfullerene and its derivativesAmantadine

Compounds that help to decrease loss of mitochondrialfunction with ageCoenzyme Q10GlutathioneThiaxzolidine carboxylate derivativesGinkgo biloba

Aminoethylcysteine ketimine decarboxylated dimerGenetic change

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INCREASING THE MAXIMUM LIFE SPAN 203

the restricted group, in caloric intake per dayand in body weight, were equal. This studydemonstrated that caloric restriction slows pro-duction of aging changes by the inherent agingprocess: the slope of the curve of the log of thechance of death versus age for restricted ani-mals is less than that of the controls.3,17A similar study has been initiated with pri-

mates.50Caloric restriction can almost certainly in-

crease the maximum life span of humans. How-ever, the increase associated with a tolerablelevel of restriction would undoubtedly besmall. The goal for humans is to significantlyincrease our healthy life span while living a

normal life. Efforts to achieve this goal shouldalso include some acceptable degree of caloricrestriction.

Compounds that compete with O2 for access to"electron-rich areas" of the mitochondria.

(a) Spin-traps commonly used in biologicalsystems are nitrones ( —* N —» O) or nitroso(—N=0.) compounds.51 They react with freeradicals to form relatively stable nitroxides(>N—O.) that are readily reduced to hydroxyl-amines (>N—OH). Although spin-traps are an-tioxidants, in biological systems their antioxi-dant activity seems small in comparison to theapparent ability of the nitroxide derivatives, atleast that of N-ferf-butyl-a-phenylnitrone(PBN), to inhibit initiation of free radical reac-tions. PBN gradually enhanced performance ofold gerbils52,53 in a radial-arm maze test (a mea-sure of memory) to near that of young gerbilsover a 2-week period of twice a day intraperi-toneal (i.p.) injections of 32 mg/kg PBN (thethreshold dose for protection against ische-mia/reperfusion brain injury).52 This was ac-

companied by increases in the brain of the ra-

tio of unoxidized to oxidized protein and of theactivities of glutamine synthetase and neutralprotease. After injections were stopped, thesemeasures returned slowly after 2 weeks to theoriginal values. The above results are those tobe expected if PBN had disproportionatelylowered the free radical reaction level in oldgerbils to near that of the young. This wouldincrease activities of both neutral protease andglutamine synthetase. The protease would in-crease the turnover rate of oxidized pro-

teins,54,55 changing the ratio of normal proteinto oxidized protein towards that of the youngand improving maze performance. Assumingthe foregoing is correct, it was in part a conse-

quence of the free radical scavenging effect ofPBN. However, most likely the major action ofPBN was to decrease the initiation rate of ad-verse free radical reactions. This could proba-bly be accomplished by the joint action of thefollowing: (a) addition of a free radical, e.g., Su-peroxide radical, to PBN followed by associa-tion of the resulting nitroxide with a mito-chondrial respiratory chain56 where, incompetition with O2 for electrons, a hydroxyl-amine is formed instead of a Superoxide radi-cal (the hydroxylamine could then diffuseaway from the mitochondria and be readilyconverted back to the nitroxide by reactingwith a free radical, e.g., a hydroxyl radical, thusresulting in cyclic oxidation of the respiratorychain; in essence, the spin-trap in a cyclic fash-ion causes the removal of a free radical whilepreventing formation of a new one); (b) de-crease formation of the hydroxyl radical byhelping to maintain cellular iron in the ferricstate57,58; (c) the nitroxide serving as a super-oxide dismutase mimic; and (d) depressing hy-droxyl radical formation inside Cu/Zn SOD(superoxide dismutase).57 In accordance withthe above discussion: 1) C57BL male mice at24.5 months of age were divided into two

groups of 50 each.59 The experimental groupreceived 0.25 mg/ml of PBN in their drinkingwater. The mean life spans for the control andtreated groups were 29.0 and 30.1 months, re-spectively, while the corresponding maximumlife spans (last survivors) were 31.7 and 33.3months. 2) Eleven 24-month-old male Sprague-Dawley rats were started on daily i.p. injectionsof 32 mg/kg of PBN for the 9.5-month studyperiod; the 12 controls of the same age were

similarly injected with 0.9% saline solution.60The average life span of the controls was 28months, while that of the experimental groupwas 30 months. At the end of the study period,36% (4 rats) of the PBN group were alive, butonly 8% (1 rat) of the controls. PBN improvedcognitive performance in several tasks; this wasassociated with decreased oxidative brain dam-age. 3) Daily intraperitoneal injection of senes-cence-accelerated mice (SAM-P8)—mice that

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204 HARMAN

seem to age at a higher than normal rate—with30 mg/kg of PBN increased both average andmaximum life spans.61 This study suggests thatthe increased free radical reaction levels inSAM-P8 mice62 are caused by higher than nor-mal rates of mitochondrial formation of super-oxide radical (faster aging).

(b) In view of the discussion above, twoclasses of compounds, nitroxides and hydrox-ylamines, should act like nitrones and nitrosocompounds.Blocking agents. Compounds that can associ-

ate with the electron-rich areas of the mito-chondrial respiratory chain, but not react sig-nificantly with it or elsewhere, may blockaccess of O2 to these areas to some extent andthus decrease Superoxide radical formation. Asearch for such substances may be productive.

(a) The "free radical sponge"63-66—bucky-ball, e.g., fullerene (Cóo)—or some of its deriv-atives may have the foregoing properties.These empty ball-like compounds showpromise as neuroprotective agents.67

(b) The antiviral agent, amantadme,68,69 mayalso be useful. This soluble, stable 10-carbonamine prevents cellular entry of a virus,70 pos-sibly by coating the virus. This compound isexcreted unchanged in the urine.

Compounds that may decrease loss of mtDNAfunction with age.

(a) Coenzyme Q10 is an essential componentof the electron transport chain71 and servesalso as an important antioxidant in both mito-chondria and lipid membranes.71 Levels ofcoenzyme Q10 in both animals and humans,decrease with age.72-74 Dietary supplementa-tion of rats with coenzyme Q10 significantly in-creased mitochondrial content of the com-

pound.75 The decline in the level of thissubstance with age hinders the transfer of elec-trons from complexes I and II to complex III.76This increases the electron density of com-

plexes I and II, resulting in a higher rate offormation of Superoxide radical. Thus, coen-zyme Q supplementation should both de-crease the block and oxidative damage to themitochondria. Apparently, very few long-termstudies have been made with coenzyme Qio-73A lifelong study of mice and rats supple-

mented with coenzyme Qio found no increasein life span and no shortening.74 Further stud-ies are indicated.

(b) Increases in oxidative damage to mtDNAwith age are associated with decreases in mi-tochondrial glutathione (GSH) content.77,78 Thechanges are reversed with oral antioxidants,e.g., thiazolidine carboxylate (TC),77 and a

Ginkgo biloba extract (EGb 761).78 Other mea-sures directed to increasing mitochondrialGSH79 include providing GSH esters, and pre-cursors of substrates for GSH synthetase and-y-glutamylcysteine synthetase. The maximumlife span80 of Drosophila melanogaster was in-creased about 18% when maintained on a dietsupplemented with either sodium or magne-sium thiazolidine carboxylate.

(c) Aminoethylcysteine ketimine decarboxy-lated dimer81-83 may slow mitochondrial Su-peroxide radical formation by inhibiting oxi-dation of mitochondrial components atconcentrations that do not inhibit function ofcomplex I.

(d) 2-ME30 and two pyridine compounds31,32have been reported to increase the maximumlife span. If these studies can be reproduced,they should prompt evaluation of other an-

tioxidants.

Genetic change. Birds have high metabolicrates compared to comparable size mammalsand yet have relatively long life spans. This isattributed to a genetically determined diver-sion of a smaller fraction of the oxygen they uti-lize to Superoxide radicals than do mam-

mals.41,42 Likewise,43 and apparently for thesame reason, the white-footed mouse (Per-omyscus leucopus) lives longer than the commonmouse (Mus musculus).Efforts to determine the cause, such as more

detailed knowledge of the structure of the com-plexes,84 of these differences in O2 diversion to

Superoxide radicals may result in measures todecrease the diversion in humans. These effortsmay be helped by studies of neuroleptics as

they may alter mitochondrial gene expres-sion.85-88The short life span of SAM-P8 mice, dis-

cussed above, is likely also caused by a muta-tion that increased mitochondrial Superoxideformation.

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INCREASING THE MAXIMUM LIFE SPAN 205

Decreasing ill effects of early lifeAntioxidant supplementation of mouse diets

from before mating until the offspring areweaned increases offspring life spans.47 This ef-fect is attributed to reduction in life-shorteningmutations secondary to the high mitotic andmetabolic activity of early life. Similar mutationsmay take place in humans. Accumulating dataimplicates changes in early life to diseases ofadulthood that shorten life: e.g., breast cancer,89,90prostate cancer,91 coronary heart disease,92 hy-pertension,93 diabetes,93 and Alzheimer disease.94The life-shortening effects of early life may

be decreased by increasing intake of dietary an-tioxidants and /or antioxidants supplements.47The adverse effects of early life can be attrib-uted in part to steroid estrogens;89,90 serum lev-els are elevated in pregnancy.95 These com-

pounds are converted by estrogen 2- and4-hydroxylases to catechol estrogens,96,97 2-and 4-estradiol. The catechol estrogens are eas-

ily oxidized to their quiñones.98 These com-

pounds, via the quinone/hydroquinone redoxsystem, are a source of Superoxide free radi-cals.98 Thus, the elevated estrogen levels dur-ing pregnancy may increase the rate of forma-tion ofmutations that adversely affect life span.Efforts to minimize the rise in estrogen levelsin pregnancy,99 e.g., a low-fat diet with an in-creased ratio of n-3 to n-6 polyunsaturated fattyacids, may increase offspring life spans.

CONCLUSION

This paper has briefly discussed some mea-

sures based on the free radical theory of agingthat may further increase the life span withoutinterfering with the activities of normal life.The feasibility of these measures needs to beevaluated. This task should be both interestingand rewarding.

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HARMAN

Address reprint requests to:Denham Harman, M.D., Ph.D.

Department ofMedicineUniversity of Nebraska College ofMedicine

984635 Nebraska Medical CenterOmaha, NE 68198-4635

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