Fd Chem. Toxic. Vol. 24. No. 10/11, pp. 1021-1030, 1986 0278-6915/86 $3.00 +0.

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Printed in Great Britain Pergamon Journals Ltd
OCCURRENCE OF LIPID OXI DATI ON PRODUCTS
IN FOODS
P. B. ADDIS
Department of Food Science and Nutrition,
University of Minnesota, St Paul, MN 55108, USA
Abst ract - - Li pi d oxidation products are ubiquitous in foods, although much variation exists in the levels
present. Although these levels are generally low, the problem of lipid oxidation severely compromises the
quality of some foods and limits the shelf-life of others. Lipid oxidation represents a key barrier in the
development of new food products and processes, especially convenience items and processes required to
manufacture them. Deleterious changes in foods caused by lipid oxidation include loss of flavour,
development of off-flavours, loss of colour, nutrient value and functionality, and the accumulation of
compounds which may be detrimental to the health of consumers. All foods that contain lipids are
susceptible to oxidation but especially affected are foods which are dehydrated, subjected to high
temperatures or cooked and subsequently stored, e.g. dehydrated eggs, cheeses and meats, foods fried in
frying oils, and cooked (uncured) meats. Specific examples of compounds which are of health concern
include lipid peroxides and the free radicals involved in their formation and propagation, malonaldehyde,
and several cholesterol oxidation products. Coronary artery disease (CAD) may be in part caused by the
consumption of lipid oxidation products.
I n t r o d u c t i o n
The st udy o f l i pi d oxi dat i on has been cl earl y est ab-
l i s hed as a ma t ur e field o f scientific i nvest i gat i on. The
l i t er at ur e is vast. The chemi cal r eact i ons i nvol ved and
t he pr oduct s f or med ar e compl ex. The effects o f l i pi d
oxi dat i on pr oduct s in t he di et on h u ma n heal t h, whi l e
needi ng f ur t her eval uat i on, ar e known t o be adver se
in s ome cases. The effects on f ood t echnol ogy, i ncl ud-
i ng pr oduct i on, pr ocessi ng and di st r i but i on, ar e enor -
mous. Ta ke n as a whol e, it is difficult t o over es t i mat e
t he mone t a r y i mpact o f l i pi d oxi dat i on.
Because of t he vast scope o f l i pi d oxi dat i on, no
a t t e mpt has been ma de t o cover t he l i t er at ur e com-
pl et el y and, i ndeed, s ome i mpor t a nt st udi es have
necessari l y been l eft out . Thi s r evi ew will first di scuss
t he heal t h i mpl i cat i ons o f t he pr oduct s of l i pi d ox-
i dat i on. Next , t he anal ysi s and occur r ence o f l i pi d
oxi dat i on pr oduct s will be revi ewed. Fi nal l y, t he need
f or f ut ur e r esear ch and r egul at or y act i on on l i pi d
oxi dat i on will be di scussed.
Sever al revi ews and cl assi cal r esear ch art i cl es ar e
al r eady avai l abl e and shoul d be consul t ed by any
seri ous s t udent o f l i pi d oxi dat i on. These i ncl ude
Published as Paper No. 14900 of the scientific journal series
of the Minnesota Agricultural Experiment Station on
research conducted under Minnesota Agricultural Ex-
periment Station Project No. 18-23.
Abbr e v i at i ons : CAD = coronary artery disease; GC = gas
chromatography; HDL = high density lipoprotein;
HMG CoA reductase = 3-hydroxy-3-methylglutaryl co-
enzyme A reductase; HPLC = high performance liquid
chromatography; IR = infra red; LDL = low-density li-
poprotein; MS = mass spectrometry, NMR = nuclear
magnetic resonance; TBA = thiobarbituric acid;
TBARS = thiobarbituric acid reactive substances;
TLC = thin-layer chromatography; VLDL = very-low-
density lipoproteins.
Addi s, Csal l any & Ki n d o m (1983), Fi noc c hi a r o &
Ri c ha r ds on (1983), Ku mme r o w (1979), Logani &
Davi es (1980), McBr i en & Sl at er (1982), Mc Ca y
(1985), Pari sh, Nandur i , Kohl & Ta yl or (1986), Pear -
son, Gr ay, Wol z a k & Hor ens t ei n (1983), Sevani an &
Hochs t ei n (1985), Si mi c & Kar el (1980), Smi t h (1981)
and Tayl or , Peng, Wer t hes s en e t al . (1979).
H e a l t h i mp l i c a t i o n s o f l i pi d o x i d a t i o n p r o d u c t s
St udi es on t he possi bl e pat hol ogi cal si gni fi cance o f
l i pi d oxi dat i on pr oduct s have devel oped in t hr ee
ar eas of i nvest i gat i on: l i pi d per oxi des (usual l y o f f at t y
acids), mal onal dehyde and chol est er ol oxi dat i on
pr oduct s. St udi es ar e not usual l y conduct ed on di-
et ar y c ombi na t i ons of t hese t hree t ypes of l i pi d
oxi dat i on pr oduct s, but it mus t be real i zed t hat such
mi xt ur es occur in our di et s dai l y.
L i p i d p e r o x i d e s
A series o f st udi es by Yagi and cowor ker s has
est abl i shed r el at i onshi ps bet ween ser um l i pi d per-
oxi de level and f act or s t hat appear t o be r el at ed t o
at her oscl er osi s. Suemat su, Ka ma da , Abe e t al . (1977)
st udi ed age- dependent changes in l i pi d per oxi de
levels in ser um l i popr ot ei ns o f humans and r epor t ed
l ow levels in chi l dr en and i ncreases wi t h age t o 70,
f ol l owed by decl i ni ng levels. Hagi har a, Ni shi gaki ,
Mas eki & Yagi (1984) conf i r med t hese fi ndi ngs by
demons t r at i ng hi gher l i pi d per oxi de levels in t he
l ow- densi t y l i popr ot ei n ( LDL) f r act i on of ser um
f r om ol d peopl e t han f r om young peopl e. Thi o-
bar bi t ur i c aci d ( TBA) was used t o quant i f y lipid
per oxi des (Yagi , 1976), and sampl es were pr epar ed t o
avoi d i nt er f er ence by bi l i r ubi n, sialic aci d and cer t ai n
wat er - sol ubl e subst ances, all of whi ch r eact wi t h
TBA. I n addi t i on, levels o f l i pi d per oxi des were
adj ust ed f or var i at i ons in t ot al l i pi ds in ser um so t hat
t he di fferences seen r epr esent ed t rue di fferences in
1021
1022 P. B. ADDIS
Tabl e 1. Serum l i poprot ei n lipid peroxi de levels in nor mal a nd di abet i c
subjects correct ed for t ot al l i pi dst
Lipid peroxi des/ t ot al lipids$
No. of
subjects VLDL LDL HDL
Nor mal 32 0.71 _+0.30 0.41 _+ 0.11 0. 42_+0. 18
Di abet i c 31 0.72 _ 0.32 0.53 -+ 0. 21" 0.74 _ 0.35**
t Ada pt e d f r om Ni shi gaki e t al . (1981).
:~Assayed by met hod of Yagi (1976).
Values are means + SD and those mar ked wi t h asterisks differ significantly
f r om the nor mal value; *P < 0.05; **P <0. 01.
absolute quantities. Increases in lipid peroxides in the
blood of diabetic patients may be of possible pat ho-
logical significance (Sato, Hot t a, Sakamot o e t al.
1979) because plasma lipid peroxide levels in di-
abetics with angi opat hy exceeded significantly the
levels in diabetics without angiopathy. Nishigaki,
Hagihara, Tsunekawa et al . (1981) noted similar
differences between diabetics and controls, even after
adjusting for total lipid variations and fractionating
the lipoproteins (Table 1).
The pri mary source of serum lipid peroxides,
whether in vi vo oxidation or diet, is unknown, but
bot h are probabl y involved. It is clear t hat bot h
linoleic acid hydroperoxides and secondary product s
of linoleic acid oxidation are absorbed into the
circulatory system and i ncorporat ed into the liver of
the rat with detrimental effects, including liver hyper-
trophy, increased serum transaminase activities and
elevated hepatic lipid peroxide levels (Kanazawa,
Kanazawa & Nat ake, 1985).
The research demonst rat i ng age- and diabetic-
association with lipid peroxide levels offers some
circumstantial evidence t hat lipid peroxides in serum
could play a role in coronary artery disease (CAD).
Absorpt i on studies establish t hat lipid oxidation
product s are absorbed from dietary sources into the
blood and internal organs and tissues. Recent reports
have provided further evidence for the pri mary in-
volvement of lipid peroxides in the pathogenesis of
atherosclerosis. Yagi, Ohkawa, Ohishi e t al . (1981)
showed t hat intravenous administration of linoleic
acid hydroperoxide in the rabbit caused aort a intimal
lesions which closely resembled the initial event in
atherosclerosis and included the adherence of aggre-
gated platelets. Nishigaki, Hagihara, Maseki e t al .
(1984) noted t hat LDL uptake by cultured smoot h
muscle cells was increased by 345 nmol/ml linoleic
acid hydroperoxide. Furt her research on cultured
endothelial cells from the human umbilical vein dem-
onstrated t hat incubation with 1.0 nmol/ml linoleic
acid hydroperoxide for 3 hr caused cellular damage,
including enlargement and dilatation of the rough-
surfaced endoplasmic reticulum and vacuolization
(Sasaguri, Nakashi ma, Mori mat su & Yagi, 1984). A
subsequent investigation compared differences in sus-
ceptibility to injury by linoleic acid hydroperoxide
between endothelial and smoot h muscle cells (Sasa-
guri, Morimatsu, Kinoshita et al . 1985). The findings
demonstrated that smoot h muscle cells were more
resistant t han endothelial cells to attack by linoleic
hydroperoxide. Therefore, the i ncorporat i on of LDL
by smoot h muscle cells and their subsequent trans-
format i on t o foam cel l s--a putative step in
at herogenesi s--coul d be favoured by lipid peroxides.
The foregoing discussion provides interesting and
perhaps highly significant dat a on the origins of
CAD. I n addition, such findings are consistent with
research on the relationship of dietary cholesterol
oxides to CAD (to be discussed later). Numerous
other systemic toxic effects of lipid peroxides (e.g.
hepatotoxicity and growth suppression) are reviewed
in many of the excellent reviews cited earlier. The
effects of lipid peroxides on cancer are complicated
by the effects of antioxidant level and certain para-
doxical effects as yet not totally underst ood (Corn-
wall & Morisaki, 1984). It is known t hat free-radical
generators, benzoyl peroxide and hydroperoxy fatty
acids, are participants in tumorigenesis. Yet, while
antioxidants generally act as inhibitors of tumori-
genesis, the number of t umours has increased after
antioxidant administration in some cases of chemical
carcinogenesis. Cornwall & Morisaki (1984) suggest
t hat these paradoxical effects may be explained thus:
cell proliferation is inhibited by lipid peroxidation,
and antioxidants would therefore favour t umour
growth by reducing the levels of peroxides. Yet
t umour initiation is inhibited by antioxidants. An
alternative explanation is t hat low concentrations of
antioxidants favour prostaglandin product i on, pro-
mot i ng cell proliferation, while high levels of anti-
oxidants suppress prostaglandin biosynthesis (Corn-
wall & Morisaki, 1984).
Therefore, al t hough much research remains to be
done, evidence is increasing t hat dietary lipid per-
oxides and antioxidants participate in the devel-
opment of cancer in humans. Indeed, the recent study
of Fujimoto, Neff & Frankel (1984), which demon-
strated a strong reaction between deoxyribonucleic
acid ( DNA) and several primary lipid peroxides, may
have provided an insight into a possible mechanism.
Ma l o n a l d e h y d e
There is little doubt t hat malonaldehyde, a second-
ary product of lipid oxidation, is toxic to living cells
(Addis et al . 1983; Pearson et al . 1983). Malon-
aldehyde can be formed in vi vo or pre-formed in
food products. As a dialdehyde, it can crosslink
proteins (Kwon & Brown, 1965), inactivate ribo-
nuclease (Manzel, 1967) and react with DNA, al-
t hough less intensely t han the pri mary lipid oxidation
product s (Fuj i mot a et al . 1984). Bird & Draper (1980)
noted several unt oward effects in skin fibroblast cells
incubated in culture with malonaldehyde, including
cytoplasmic vacuolization, karyorrhexis, micro- and
multi-nucleation, and a marked reduction in protein-
synthesizing capacity. Bird, Draper & Basrur (1982)
found t hat malonaldehyde was ten times more active
Lipid oxides: occurrence and toxicology 1023
t han acet al dehyde at i nduci ng chr omosomal aber r a-
tions.
Studies suggesting t hat mal onal dehyde may be
carci nogeni c (Shamberger, Andr eone & Willis, 1974)
and mut ageni c ( Mukai & Gol dst ei n, 1976) have been
quest i oned by Mar net t & Tut t l e (1980), who demon-
st rat ed t hat the mut ageni c pr oper t i es at t r i but ed to
mal onal dehyde in the earl i er research may have been
caused by an i nt ermedi at e f or med in the react i on
used to pr epar e mal onal dehyde. Moreover, several
i nvest i gat ors have criticized t he use of TBA reagent
to anal yse mal onal dehyde in food product s ( Addi s et
al. 1983). TBA overest i mat es the amount of mal -
onal dehyde present (Csal l any, Guan, Manwar i ng &
Addi s, 1984). Therefore, it is difficult at this time to
make a realistic assessment of the possi bl e heal t h risk
associ at ed with the consumpt i on of foods cont ai ni ng
mal onal dehyde.
Chol e s t e r ol ox i dat i on pr oduc t s
A comprehensi ve review of chol est erol aut -
oxi dat i on has been publ i shed by Smi t h (1981). Mor e
recent reviews coveri ng heal t h i mpl i cat i ons of choles-
t erol oxi dat i on in foods include t hose by Addi s et al.
(1983), Fi nocchi ar o & Ri char dson (1983) and Pear-
son et al. (1983). A di scussi on of chol est erol and its
oxi dat i on pr oduct s and human heal t h logically begins
with CAD, and indeed numer ous studies have been
publ i shed t hat i mpl i cat e aut oxi dat i on pr oduct s of
chol est erol in this disease. Dat a on cancer are ex-
t remel y cont r adi ct or y ( Addi s et al. 1983) with the
except i on of the mut ageni ci t y of chol est erol ~- and
fl -epoxi de, discussed by Sevani an (1986). Therefore,
t he present discussion will be limited t o CAD.
At he r oge ni c i t y
Ani t schkow (1913) is cited as one of t he first to
succeed in the i nduct i on of at heroscl erosi s as a
specific consequence of feeding chol est erol di ssol ved
in veget abl e oil to r abbi t s ( Tayl or et al. 1979). I nsof ar
as research i nt o at heroscl erosi s is concerned, Ani t s-
chkow' s met hod has been frequent l y repeat ed as a
rout i ne means of pr ovoki ng experi ment al at her o-
sclerosis in rabbi t s. Ost ensi bl y, this resembles human
at heroscl erosi s. Schwenk, Stevens & Allschul (1959)
recogni zed t hat chol est erol exposed to heat in ai r was
rendered mor e pot ent , but consi dered t hat purified
chol est erol by itself was capabl e of i nduci ng at hero-
sclerosis. However, i nvest i gat ors conduct i ng experi-
ment s on the effects of di et ary chol est erol on the
ci r cul at or y system have often not consi dered the
t endency of chol est erol spont aneousl y to undergo
oxi dat i on in ai r upon storage. This phenomenon
clearly i ndi cat es t hat chol est erol feeding experi ment s
shoul d be eval uat ed with caut i on when the pur i t y of
di et ar y chol est erol has not been det ermi ned and
carefully preserved. Unf or t unat el y, as a rule these
pr ecaut i ons are not t aken, even nowadays when good
anal yt i cal pr ocedur es are avai l abl e.
Smith, Mat t hews, Price et al. (1967) used i mpr oved
anal yt i cal met hods such as TLC to find some 30
oxi dat i on pr oduct s in USP- gr ade chol est erol , pr ovi d-
ing decisive evidence for the i nst abi l i t y of chol est erol .
Suggestions were made t hat the spont aneous ox-
i dat i on pr oduct s of chol est erol , i nst ead of pure cho-
lesterol, mi ght be the act ual i ni t i at i ng fact or in the
t r ansf or mat i on of nor mal cells i nt o those of the
lesion. This scepticism about the role of purified
chol est erol in at heroscl erosi s was st rongl y suppor t ed
by earl i er studies t hat demonst r at ed t hat endo-
genousl y i nduced hyperchol est erol aemi a (t hrough a
hor monal mechani sm) was onl y mi ni mal l y at hero-
genic, whereas a compar abl e level of hyper-
chol est erol aemi a i nduced by feeding the avai l abl e
commerci al chol est erol pr oduced severe at her o-
sclerosis (Chaikoff, Li ndsay, Lorenz & Ent enman,
1948; Seifter & Baeder, 1956). One woul d expect t hat
endogenousl y synthesized chol est erol is prot ect ed
from aut oxi dat i on, pr obabl y by in vi vo ant i oxi dant s
in the organi sm.
Some direct evidence t hat at heroscl erosi s coul d be
i nduced by feeding a chol est erol aut oxi dat i on pr od-
uct was report ed by MacDougal l , Biswas & Cook
(1965), who screened a large number of st eroi ds for
t hei r cyt ot oxi ci t y to r abbi t aort i c cell cultures. In
addi t i on, Cook & MacDougal l (1968) fed rabbi t s
30mg chol est anet r i ol / kg/ day for 27-310 days and
not ed superficial l i pi d deposi t i on in the aor t a, medi al
fibrosis and calcification.
Imai , Wert hessen, Tayl or & Lee (1976) were the
first to demonst r at e angi ot oxi c effects from con-
t ami nant s of USP- gr ade chol est erol . They admi ni s-
t ered chol est erol oxi de concent rat es, made from
USP- gr ade cholesterol, to rabbi t s by gavage. Wi t hi n
24 hr of admi ni st r at i on of 250 mg/ kg body weight, a
great er frequency of degenerat ed aor t i c cells was
observed in t he concent r at e- t r eat ed groups t han in
t hose given purified cholesterol. The admi ni st r at i on
of the concent rat e over a 7-wk per i od at a t ot al of
1 g/ kg i nduced aort i c lesions. However, these lesions
were di st i nct from t hose i nduced by convent i onal
chol est erol feeding, being charact eri zed by diffuse
i nt i mal pr ol i f er at i on of smoot h muscle cells and
fi brous st r oma wi t hout foam cells. In cont r ast to the
lesion i nduct i on by the concent rat e, the rabbi t s given
the purified chol est erol had no i nduced lesions.
Peng, Tayl or, Tham et al. (1978) used a cell
cul t ured t echni que i nst ead of feeding. Si mi l ar results
were obt ai ned, in t hat the concent rat e showed re-
mar kabl e in vi t ro aort i c smoot h muscle cell toxicity,
while the same dose of purified chol est erol had no
effect. The concent rat e was separ at ed by TLC i nt o six
fractions, i ncl udi ng cholesta-3, 5-dien-7-one, C-7 chol -
esterol oxides, 25-hydroxychol est erol and choles-
t anet ri ol . The most t oxi c deri vat i ves included
25-hydroxychol est erol and chol est anet ri ol , on the
basis of the frequency of dyi ng and dead cells from
the aort i c smoot h muscle cell culture. Al l these
observat i ons demonst r at ed t hat a compound or com-
pounds present in the current commerci al l y avai l abl e
USP- gr ade chol est erol , not chol est erol pe r se, mi ght
be responsi bl e for i ni t i at i ng atherosclerosis. Cyt o-
t oxi ci t y of chol est erol oxi dat i on product s was
confi rmed agai n, t oget her with the absence of cyt o-
toxic effects of purified chol est erol , in a fol l ow-up
st udy of 12 commerci al l y avai l abl e oxi dat i on pr od-
ucts (Peng, Tham, Tayl or & Mi kkel son, 1979). The
degrees of cyt ot oxi ci t y were gr aded and measured as
the percent age of dyi ng and dead ceils in cell cul t ures
within 24hr . 25-Hydroxychol est erol and choles-
t anet r i ol were the most toxic compounds among the
oxi di zed sterols tested, and they significantly de-
1024 P. B. ADDIS
pressed the act i vi t y of 3-hydroxy-3-met hyl gl ut aryl
coenzyme A reduct ase ( HMG CoA reductase), a
regul at ory enzyme of chol est erol biosynthesis.
25-Hydroxychol est erol was the most pot ent i nhi bi t or,
exerting up to 83% i nhi bi t i on at a 3-/zg/ml con-
cent rat i on in cul t ure medi um; chol est anet ri ol was
moder at el y pot ent . Thus, the sequence of degree of
i nhi bi t i on was not correl at ed with t hat of cyt o-
toxicity. Purified chol est erol showed a very mi ni mal
effect on the act i vi t y of HMG CoA reduct ase (Bres-
low, Lot hr op, Spaul di ng & Kandut sch, 1975; Brown
& Gol dst ei n, 1974; Peng e t a l . 1979). The di screpancy
between the cyt ot oxi ci t y and enzyme i nhi bi t i on po-
tency of chol est anet ri ol was discussed by Peng e t a l .
(1979), who specul at ed t hat chol est anet ri ol may also
affect the cell membrane. The pot ent cyt ot oxi ci t y of
chol est anet ri ol may be due to the st ronger pol ar
groups on one end of t he mol ecul e and t he hydr o-
phobi c gr oup on the ot her end, which makes it
possi bl e for this compound to get i nt o the membr ane
easily and t hereby cause membr ane dysfunct i on.
Imai , Wert hessen, Subr amanyam e t a l . (1980) re-
peat ed t hei r earl i er experi ment on the cyt ot oxi ci t y of
the concent rat e (Imai e t a l . 1976) by injecting it i nt o
rabbi t s, i nst ead of using gavage, because the l at t er
rout e mi ght have led to uncert ai nt i es associ at ed with
bi ot r ansf or mat i on, ret ent i on, excret i on and absor p-
tion. The aor t ae of the r abbi t s were exami ned 24 hr
aft er i nt ravenous i nj ect i on of the same concent rat e.
The increase in smoot h muscle cell deat h caused by
the concent rat e confi rmed the angi ot oxi ci t y of the
aut oxi dat i on cont ami nant s. Maj or branches of the
pul monar y ar t er y r esponded to i nt ravenousl y injec-
ted sterols by undergoi ng grossly visible sequent i al
t hi ckeni ng due to a series of changes rangi ng from the
initial i nfl ammat i on to subsequent r epai r by
fi bromuscul ar thickening. The concent rat e, syn-
thesized chol est anet ri ol and 25-hydroxychol est erol
were equal l y pot ent . Nei t her 7-ket ochol est erol nor
the epoxi de affected the maj or branches. Nei t her
freshly purified chol est erol nor t he vehicle i nduced
changes in the maj or or mi nor branches of t he
pul monar y arteries.
Baranowski , Adams, Bayliss Hi gh & Bowyer
(1982) i nvest i gat ed t he act i vi t y of oxi di zed sterols in
pr omot i ng tissue i nf l ammat i on and necrosis and in
causing cell deat h in tissue culture. In mouse
fi brobl ast s and pi g vascul ar smoot h muscle, the most
mar ked i nhi bi t i on of growt h was observed with cho-
lestanetriol. The epoxi de was found to be mor e toxic
t han 25-hydroxychol est erol . When tested sterols were
injected subcut aneousl y i nt o the abdomen of rat s, the
hi st ol ogi cal results showed t hat oxyst erol s in general
pr omot ed a great er i nf l ammat or y react i on t han di d
purified cholesterol. Chol est anet ri ol i ni t i al l y pr o-
duced the largest lesion but, at a l at er stage, the
25-hydroxychol est erol i mpl ant became the largest.
Al l of these studies were conduct ed with unreal -
istically high dosages not likely to occur under the
nor mal human di et ary practices. Nevertheless, it can-
not be denied t hat these results bear the very
significant evidence t hat i t was chol est erol oxides t hat
exerted cyt ot oxi ci t y and at heroscl erot i c lesions and
not pure cholesterol.
The i mpl i cat i on of chol est erol oxides in at her o-
sclerosis is furt her st rengt hened by the finding of
compounds identified as 26-hydroxychol est erol and
7-ket ochol est erol (Brooks, Har l and & Steel, 1966),
the esters of 7~t-hydroxy-, 7fl -hydroxy-, 24-hydroxy-
and 26-hydroxychol est erol (Brooks, Steel, Gi l ber t &
Har l and, 1971), the diesters of 26-hydroxychol est erol
(Brooks, Gi l ber t & Har l and, 1972) and 25-hydroxy-
chol est erol (Smith, Teng, Lin & Seitz, 1981) in at h-
er oma from grossly diseased human aor t al tissue.
Chol est erol ct-epoxide was found in sera of pat i ent s
with varyi ng degrees of hyperchol est erol aemi a and
at heroscl erosi s but not in nor mal vol unt eers ( Gr ay,
Lawri e & Brooks, 1971). Accor di ng to the obser-
vat i on of McDougal l e t a l . (1965), 26-hydroxy-
chol est erol was one of the four st eroi ds found to be
highly toxic t owar d organ cultures of r abbi t aort a.
The quant i t y of this sterol per unit weight of dry
aor t a i ncreased with the severity of at heroscl erosi s in
paral l el with the increase of chol est erol and t ot al
lipids (Brooks e t a l . 1972). Even t hough these results
may suggest a rel at i onshi p between some of the
chol est erol oxides and at heroscl erosi s, no definitive
expl anat i on for the presence of these pr oduct s in
at heroscl erot i c tissues or in sera from hyper-
chol est erol aemi cs has yet been fort hcomi ng.
Recently, mor e convi nci ng i nf or mat i on was ob-
t ai ned in suppor t of the hypot hesi s t hat some ox-
i dat i on deri vat i ves of chol est erol are pr obabl y the
pri me cause of at heroscl erot i c lesions and t hat
chol est erol and its esters have a role t hat is merely
secondary. 25-Hydroxychol est erol was shown to be
preferent i al l y i ncor por at ed i nt o the at herogeni c l i po-
prot ei ns LDL and VLDL (very-l ow-densi t y l i po-
prot ei n) but t o be bound onl y to a small degree by
hi gh-densi t y l i popr ot ei n ( HDL) , which is ant i -
at herogeni c (Peng, Tayl or, Mosbach e t a l . 1982).
Count i ng of the r adi oact i vi t y in each l i popr ot ei n
fract i on 24hr aft er oral admi ni st r at i on of 25-hy-
droxychol est erol and [~4C]cholesterol to ten squirrel
monkeys showed t hat the maj or i t y of labelled
25-hydroxychol est erol was l ocat ed in the LDL and
VLDL (55.1 and 34.7%, respectively) with onl y
10.2% in the HDL. The di st r i but i on of labelled
chol est erol in VLDL, LDL and HDL was al most
i dent i cal to t hat of unl abel l ed cholesterol. These
results suggest t hat most 25-hydroxychol est erol is
t r anspor t ed to and i ncor por at ed in the peri pheral
tissue, i ncl udi ng vascul ar tissue, by VLDL and LDL
in squirrel monkeys. The preferent i al i ncor por at i on
of 25-hydroxychol est erol i nt o at herogeni c lipo-
prot ei ns led the aut hor s to speculate t hat the art ery
coul d be i nj ured by the angi ot oxi ci t y of 25-hydroxy-
chol est erol deri ved from di et ary sources.
Ot her cyt ot oxi c effects of chol est erol oxi dat i on
deri vat i ves have been not ed which are not necessarily
rel at ed to CAD but are wort h ment i oni ng. Higley &
Tayl or (1984) demonst r at ed t hat several chol est erol
oxi dat i on pr oduct s were st eat ot i c and cyt ot oxi c to L
cells of mice.
Lipid oxi dat i on products in foods: anal ysi s and
occurrence
Numer ous studies have est abl i shed the fact t hat
l i pi d oxi dat i on product s occur in foodstuffs. Un-
fort unat el y, because of the compl ex nat ure of such
oxi dat i on pr oduct s and their i nst abi l i t y, the pr o-
Lipid oxides: occurrence and toxicology 1025
Tabl e 2. Compar i s on of HPLC a nd TBA met hods for mal onal dehyde
quant i fi cat i on*
Levels of mal onal dehyde ( #g/ g wet
t i ssue)t det ermi ned by:
No. of
Sampl e sampl es HPLC TBA TBA/ HPLC
Beef 9 0. 14 _+ 0.085 0.44 + 0.19 3.14
Por k 9 0.11 _+ 0.06 0.39 + 0.12 3.55
*Adapt ed f r om Csal l any e t al . (1984).
"t'Means + SD.
pensity of foods to contain large quantities of inter-
fering substances, and the lack of specificity of the
analytical met hods used, the quantification of specific
compounds has been difficult. Indeed, it is only
recently t hat reliable met hods have been developed
for free malonaldehyde and the cholesterol oxides.
Many met hods have been used to determine lipid
peroxides but TBA is the most widely used and is also
used, inappropriately, for quantifying mal-
onaldehyde. Therefore, this discussion begins with a
review of the lipid peroxide and malonaidehyde
content of foods and the met hodol ogy involved, and
stresses the need for caution in interpreting published
values. Cholesterol oxidation product s are discussed
separately and, al t hough the methods are quite
different, a similar set of cautions applies to them
concerning published dat a on foods.
TBA is the most frequently used met hod for
quantification of lipid peroxides in foods and biolog-
ical fluids and tissues and, indeed, Yagi (1980) sug-
gests t hat it is useful to apply the same test (TBA) for
bot h types of procedure to permit some degree of
standardization. The discovery t hat TBA was useful
as a measure of aut oxi dat i on product s was made by
Kohn & Liversedge (1944). Milk fat was the first food
to be analysed by TBA (Dunkley, 1951; Dunkl ey &
Jennings, 1951). Pat t on & Kurt z (1951) first sug-
gested t hat mal onai dehyde was the compound likely
to be reacting with TBA t o form the coloured com-
plex absorbing at 532 nm. The studies of Sinnhuber
& Yu (1985a, b) established the term ' TBA number' ,
defined as ppm mal onal dehyde (mg/kg or/ ~g/ g) in a
sample of food. Unfort unat el y, this excellent research
has led many to conclude t hat TBA is a specific
measure of mal onal dehyde and TBA results are
frequently expressed in terms of ppm malonaldehyde.
However, the fact t hat it is erroneous to use TBA as
a direct measure of malonaldehyde has been dis-
cussed by many authors, including Rethwill (1979),
Guan (1982), Csallany e t a l . (1984), Sevanian &
Hochstein (1985) and Fuj i mot o e t a l . (1984). Furt her
complicating matters is the wide variety of TBA tests
available (Rethwill, 1979). These problems have stim-
ulated the search for alternative methods for mon-
itoring lipid oxidation (Sevanian & Hochstein, 1985)
and for the direct quantification of free mal-
onaldehyde. Such a met hod has now been published
by Csallany e t a l . (1984). High-performance liquid
chr omat ogr aphy (HPLC) of free mal onal dehyde was
conduct ed on a TSK61000PW column. Rat liver,
pork and beef muscle were analysed by the HPLC
procedure and the TBA met hod of Tarladgis, Watts,
Younat han & Dugan (1960) and Siu & Draper
(1978). Compari son of the HPLC procedure with the
TBA met hod clearly demonst rat ed a large over-
estimation of malonaldehyde by TBA (Table 2).
Therefore, it must be concluded that whereas TBA
procedures are very useful for the measurement of
some of the numerous lipid oxidation products, it
should not be considered as a procedure for mal-
onaldehyde quantification, nor should the results be
expressed as ' ppm malonaldehyde' . The term ' thio-
barbituric acid reactive substances' or ' TBARS'
avoids the inaccuracies inherent in ' ppm mal-
onaldehyde' and should be used in its place.
With the foregoing information as i mport ant back-
ground material, we can now evaluate the existing
literature of lipid peroxides and malonaldehyde in
foods.
Virtually every type of food has been reported to
contain malonaldehyde (and lipid peroxides). Foods
t hat are of most concern are dehydrated products,
cooked (uncured) and stored meats, and animal and
vegetable fats used for industrial frying (broasting,
deep-fat frying). Shamberger, Shamberger & Willis
(1977) surveyed the "mal onal dehyde" content of
food using TBA and reported levels as high as
39. 0ppm (cooked chicken) and 27. 0ppm (cooked
beef).
On the basis of the work of Csallany e t a l . (1984)
it is extremley doubtful t hat free malonaldehyde
levels approached even 50% of the quot ed levels, but
the values do indicate the clear presence of pri mary
and secondary lipid oxidation products, some of
which are likely t o be more toxic t han malonaldehyde
(Fuj i mot o e t a l . 1984; Nishigaki e t a l . 1984). A similar
survey by Siu & Draper (1978) reported far less
"mal onal dehyde", presumably because of differences
in the met hodol ogy used by them and by Shamberger
e t a/.(1977). The highest value reported by Siu &
Draper (1978) was in good agreement with a sub-
sequent publication by Newberg & Concon (1980).
Yagi (1980) modified the TBA procedure and ana-
lysed foods, reporting the results not as malon-
aldehyde but as "lipid peroxides, nmol / g". His
reported values were as high as 238.3 (fried
cuttlefish).
There have been numerous reports of toxicity in
animals associated with consumpt i on of oxidized
frying oils (Yagi, 1980). Thompson & Aust (1983)
noted several deterioration reactions during the heat-
ing of soya-bean oil for 100 hr, including the devel-
opment of toxicologically significant levels of poly-
mers. Rhee & Stubbs (1978) noted more rapid
deterioration of oils available in health-food stores
(no antioxidants) t han of conventional oils (with
antioxidants).
It is apparent from the foregoing sample of litera-
ture on the subject, t hat levels of bot h lipid peroxides
and mal onal dehyde and other secondary oxidation
FoCT. 24/10-11~
1026 P. B. ADDIS
Tabl e 3. Some possible pitfalls in the quant i f i cat i on of cholesterol oxi dat i on pr oduct s in foods*
Type Exampl e
I. Loss of cholesterol oxides
2. Pr oduct i on of art efact s
3. Loss of 7-ket ochol est erol with
puri fi cat i on
4. Mi s-i dent i fi cat i on
5. Insol ubi l i t y of cholesterol oxide in
non- pol ar sol vent
6. Poor resol ut i on
7. Inst abi l i t y dur i ng l engt hy TLC
8. Inst abi l i t y dur i ng gas c hr oma t ogr a phy
9. Poor det ect i on by HPLC UV det ect ors
ct-epoxide, hot saponi fi cat i on
7-ket ochol est erol t o 3, 5-chol est adi ene-7-one
Same as (2)
No use of MS, I R or NMR
Chol est anet ri ol i nsol ubl e in pet r ol eum et her
TLC
TLC
Pr oduct i on of dehydr at i on pr oduct s of cholesterol
oxides at hi gh col umn t emper at ur es
- and fl -epoxi des of chol est erol and
t hei r triol hydrol ysi s pr oduct s
*Sources: Tsai (1984) a nd Par k (1985).
pr oduct s may reach levels high enough to concern
researchers.
These results st rongl y suggest t hat oxi di zed frying
fats coul d have del et eri ous effects on persons con-
suming deep-fat -fri ed foods and t hey i ndi cat e a need
for furt her wor k in this area.
In spite of the many est abl i shed toxic effects of
chol est erol oxides, t hei r occurrence in t he human diet
has been successfully st udi ed onl y recently. There
have been technical difficulties with the i sol at i on of
chol est erol oxides from food, a much mor e compl i -
cat ed mat r i x t han ai r-aged or model system choles-
terol. To dat e, onl y a limited number of f ood pr od-
ucts has been exami ned. In most of t hose studies,
some technical quest i ons are left unanswered.
The i dent i fi cat i on of chol est erol oxides has been
based upon the ret ent i on i ndex measured by chro-
mat ogr aphy, in spite of t he l i mi t at i ons of such met h-
odol ogy ( chr omat ogr aphy was not designed for qual -
i t at i ve analysis). Consi deri ng the compl ex nat ur e of
foods and the t race levels of sterols among l i pi d
component s, numer ous compounds can elute at or
ar ound the ret ent i on site of the sterol of interest,
easily l eadi ng to mi s-i dent i fi cat i on. Therefore, cross-
confi rmat i on by I R spect roscopy or MS is a necessity.
Most earl y studies di d not provi de quant i t at i ve dat a,
and some studies were conduct ed wi t hout pr oper
precaut i ons agai nst art efact f or mat i on duri ng mani p-
ul at i on of t he samples.
In t he light of these common shortfalls, t here seems
no rel i abl e or aut hor i t at i ve met hod, among t he earl y
report s, for the quant i t at i ve det er mi nat i on of chole-
sterol oxides in food. Met hods vary from the simple
det er mi nat i on of mel t i ng poi nt to st at e-of-t he-art
chr omat ogr aphy. Therefore, the studies publ i shed
shoul d be i nt erpret ed with great care. The
quant i fi cat i on of t race quant i t i es of sterol oxides
from foods t hat cont ai n much higher levels of choles-
terol, triglyceride, phosphol i pi ds and ot her inter-
fering chemi cal s is difficult, and pitfalls abound. A
t hor ough review of met hodol ogy has been present ed
by Par k (1985).
In spite of the compl ex nat ur e of the food mat ri x,
a number of f ood pr oduct s have been r epor t ed to
cont ai n some chol est erol oxides. Two recent reviews
listed t hose foodstuffs ( Fi nocchi ar o & Ri char dson,
1983; Smith, 1981). However, most of the chol est erol
oxides found in those foodstuffs appear to be art e-
facts of human i nt ervent i on due to i nt ent i onal ox-
i dat i ve stress, such as i r r adi at i on or ext ended heating.
The l ack of pr oper pr ecaut i ons duri ng the
saponi fi cat i on of the samples frequent l y resulted in
the f or mat i on of chol est a-3, 5-di en-7-one (Tsai, 1984).
Mor eover , earl y studies were conduct ed with less
sophi st i cat ed anal yt i cal tools. Accordi ngl y, earl y re-
por t s shoul d be regarded as the possi bi l i t y t hat the
oxi dat i on pr oduct s of chol est erol may occur in food
under cert ai n condi t i ons rat her t han as definitive
evidence for t hei r presence in the foodstuffs. An
except i on to this caut i on is the recent finding of
epi meri c epoxides. A br i ef sampl i ng of met h-
odol ogi cal pitfalls uncovered by Par k (1985) and Tsai
(1984) in t hei r reviews of t he l i t erat ure is summari zed
in Tabl e 3.
It t herefore appear s reasonabl e to restrict this
review to the mor e recent publ i cat i ons. Summari es of
earl y studies have been publ i shed (Park, 1985; Tsai,
1984).
Definitive studies on i sol at i on, i dent i fi cat i on and
quant i fi cat i on of chol est erol ct- and fl -epoxi des in
powder ed eggs were conduct ed by Tsai and co-
wor ker s (Tsai & Hudson, 1984 & 1985; Tsai, Ijichi,
Hudson & Meehan, 1980). Briefly, the met hod in-
volved chl or of or m- met hanol ext ract i on, silicic aci d
chr omat ogr aphy and GC and HPLC. Qual i t at i ve
confi rmat i on was accompl i shed by MS, NMR and I R
spectral det ermi nat i on. Tsai & Hudson (1985) pub-
lished quant i t at i ve results obt ai ned on commerci al
dry egg product s. A synopsis of these results appear s
in Tabl e 4. They demonst r at ed t hat a great var i at i on
exists among samples, t hat some very high i ndi vi dual
samples were found, t hat the average of the sampl es
was high and t hat st eam-i nj ect ed spray dryers cause
less chol est erol oxi dat i on t han gas-fired. These results
do not include dat a on the six or mor e ot her common
chol est erol oxides nor on the l i pi d peroxi des and
ot her secondar y l i pi d oxi dat i on pr oduct s t hat were
pr obabl y present. Therefore, the pot ent i al for t oxi ci t y
of powder ed eggs may be very significant. It must be
Tabl e 4. Chol est erol ~t- and fl -epoxi des in
powder ed scr ambl ed egg mix*
Mi x cont ent (ppm)
of cholesterol epoxides:
Egg mix
sampl e ~ + fl ct
A 30 6
B 33 7
C 7 1
*Adapt ed f r om Tsai & Huds on (1985).
Lipid oxides: occurrence and toxicology 1027
recogni zed t hat chol est erol oxidizes slowly compar ed
to pol yunsat ur at ed fat t y acids.
Par k & Addi s (1985a) devel oped a relatively mi l d,
r api d and accurat e HPLC met hod for chol est erol
oxides but were limited by technical difficulties to the
C-7 derivatives. Briefly, the met hod uses a
chl or of or m- met hanol ext ract i on, silica-gel chro-
mat ogr aphy puri fi cat i on and HPLC on a 10pr o
#- Por asi l column. UV det ect i on was used. Al l posi-
tive peaks from food sampl es were collected, evapo-
r at ed under ni t rogen, dissolved in ethyl acetate and
subjected to MS. The results showed high levels of the
t hree chol est erol oxi dat i on pr oduct s (13.8-70.1 ppm)
in brai n and liver pr epar at i ons sol d at heal t h- f ood
stores, significant levels in french fries obt ai ned at
fast -food r est aur ant s (4.1-58.8 ppm) and 1.1 ppm in
a pancake mix which cont ai ned powder ed egg. Raw
beef (muscle, liver and brai n), fried chicken, cooked
hamburger, beef j er ky and liver sausage were not
found to cont ai n C-7 derivatives.
Because of the l i mi t at i ons of the r epor t by Tsai &
Hudson (1985) to the chol est erol epoxi des and of
t hei r own st udy t o C-7 derivatives, Par k & Addi s
(1985b) devel oped a met hod t hat permi t s quantifi-
cat i on of all chol est erol oxides believed to be of
t oxi col ogi cal significance by a single GC injection.
Fused-si l i ca capi l l ar y col umns were eval uat ed but
t hermal i nst abi l i t y of t he di ol deri vat i ves of choles-
t erol necessi t at ed der i vat i zat i on of the chol est erol
oxides to the t ri met hyl si l yl ethers. The deri vat i zed
chol est erol oxides were compl et el y resolved on a
DB- l col umn using t emper at ur e pr ogr ammi ng.
Fl ame i oni zat on was used as the met hod of det ect i on
(Park & Addi s, 1985b). Food was pr epar ed for
anal ysi s by addi ng an i nt ernal st andar d (40/~g
5~t-cholestane), ext ract i ng with chl or of or m- met hanol
(2:1, v/v), evapor at i ng the solvent and col d-
saponi fyi ng t he dr i ed lipids for 20 hr (23C). Af t er
addi t i on of 10ml distilled water, the non-
saponi fi abl es were ext ract ed t hree times with 10-ml
por t i ons of di et hyl et her and washed once with 5 ml
0.5 N- KOH and t hree times with 5 ml distilled water.
Af t er dryi ng, t he washed non-saponi fi abl es were dis-
solved in 100pl pyri di ne, deri vat i zed and injected
i nt o the gas chr omat ogr aph.
Ryan, Gr ay & Mor t on (1981) demonst r at ed by
TLC the devel opment of chol est erol oxides in t al l ow
heat ed t o frying t emperat ures. Par k & Addi s (1985a)
not ed t he existence of C-7 oxi di zed chol est erol in
Fr ench fries cooked in tallow. Therefore, studies were
done by Par k & Addi s (1986a, b) to appl y GC (Park
& Addi s, 1985b) t o quant i fi cat i on of chol est erol
oxi dat i on pr oduct s in t al l ow heat ed t o t emperat ures
used in fast -food rest aurant s. Al l positive chr omat o-
graphi c findings were confi rmed by GC- MS. Heat i ng
t al l ow at either 155 or 190C resulted in the loss of
hal f of the initial cont ent of chol est erol at 250 hr, with
sampl es heat ed at 190 being affected slightly mor e
severely t han t hose at 155 . Af t er 250 hr at 155 , 7%
of the chol est erol had been convert ed to 7-ket o-
chol est erol with lesser quant i t i es of 7ct- and 7fl-hy-
dr oxy (1.5 and 2. 5%, respectively) and ~- epoxi de
(4%) deri vat i ves of chol est erol (Park & Addi s,
1986a), with onl y 50% of the ori gi nal chol est erol
remai ni ng. A subsequent st udy confi rmed and ex-
t ended these findings by demonst r at i ng f or mat i on of
i . 2 and 1.1% of the chol est erol i nt o 7-ket o and
-epoxi de derivatives, respectively, aft er 70 hr at 135
and the del ayi ng of chol est erol degr adat i on by
~t-tocopherol ( 100ppm) plus ascorbyl pal mi t at e
(500 ppm).
Par k (1985) also demonst r at ed t hat pr ecooked
roast beef was resi st ant t o chol est erol oxi dat i on in
spite of the wel l -est abl i shed pr opensi t y of precooked,
uncured meat s to di spl ay muscle membr ane phos-
phol i pi d oxi dat i on (warmed-over flavour).
Dr y eggnog mix, subjected to fluorescent light
exposure, devel oped i ncreasi ng levels of 7~- and
7fl -hydroxychol est erol up to 80 days of st orage
(Heri an & Lee, 1985). TLC was used to anal yse
but t er oil and gr at ed cheese for cholesterol oxides
( Fi nocchi ar o, Lee & Ri char dson, 1984). Positive sam-
ples were anal ysed by HPLC and MS. Bot h bleached
but t er oils and gr at ed aged cheese di spl ayed levels of
chol est erol oxides t hat were pr obabl y significant.
However, dat a are not compl et e on this question.
Kou & Hol mes (1985) devel oped an HPLC pr o-
cedure specifically for 25-hydroxychol est erol , pr oba-
bl y the most at herogeni c chol est erol oxi de known.
None was det ect ed in l ard, cream, fresh egg yol k or
spr ay- dr i ed egg yol k powder. However, ot her re-
searchers have det ect ed 25-hydroxychol est erol in
powder ed scrambl ed egg mix (Missler, Wasi l chuk &
Merri t t , 1985). Pike & Peng (1985) st udi ed the sta-
bility of shell egg and liquid yol k lipids duri ng st orage
for 18 mont hs at 4C. They not ed little in the way of
oxi dat i ve det eri orat i on. Therefore, it is possi bl e t hat
onl y in spr ay- dr i ed pr oduct s do lipid oxi dat i on pr od-
ucts accumul at e to a significant degree. Because the
egg yol k is an ext remel y rich source of chol est erol ,
t hor ough i nvest i gat i ons shoul d be made on the po-
tential for egg yol k oxi dat i on.
It is obvi ous t hat the pot ent i al for chol est erol
oxides to form in foods is very real, al t hough numer-
ous foods are somewhat resi st ant to this most severe
t ype of aut oxi dat i on. Research on the occurrence of
chol est erol oxides in foods is in its infancy. Onl y
duri ng the past 4 y r have accurat e met hods been
devel oped. There has not been enough time for a
t hor ough analysis of all chol est erol -cont ai ni ng foods
to be conduct ed. A high pr i or i t y should be pl aced on
such i nvest i gat i ons al ong with sui t abl e t oxi col ogi cal
studies.
Lipid autoxidation, antioxidants and human health
It is appar ent from the foregoi ng di scussi on t hat
the human f ood suppl y includes generous quant i t i es
of lipid aut oxi dat i on pr oduct s as well as vari ous
ant i oxi dant s pr ovi ded to i nhi bi t them. At t empt i ng to
formul at e a ri sk/ benefi t rat i o for ant i oxi dant s is a
difficult t ask and will not be at t empt ed here. The
pur pose of this paper is to assess the risk of not using
ant i oxi dant s t hr ough a pr el i mi nar y eval uat i on of the
heal t h risk of oxi dat i on pr oduct s in our f ood supply.
The mai n focus has been CAD, al t hough numer ous
ot her human afflictions can be rel at ed to con-
sumpt i on of l i pi d oxi dat i on product s.
Wi t h respect to CAD, one must be i mpressed with
the copi ous dat a collected on t he levels of l i pi d oxides
in our foods, as well as t he acut e angi ot oxi ci t y caused
by l i pi d oxides in ani mal and cell cul t ure studies.
1028 P. B. ADDIS
Even mor e i mpr essi ve is t he close agr eement of ma ny
o f t hese i nvest i gat i ons ( pr i mar i l y f r om t he l abor a-
t ori es o f K. Yagi and C. B. Tayl or ) wi t h t he cur r ent
t hi nki ng of pat hol ogi st s in t he field of at her oscl er osi s
research.
An i nf or mat i ve r ecent revi ew o f cur r ent l y accept ed
model s f or at her oscl er osi s has been publ i shed by
Mo o r e (1985), who out l i ned t he t wo ma j or con-
t endi ng model s of at her oscl er osi s by suggest i ng t hat
el ement s of bot h t heor i es ma y be true. He states:
"One theory postulates that atherosclerosis is caused by
the deposition of lipid in the vessel wall during periods
of abnormal elevation of blood lipid levels or in
association with disordered metabolism of lipoproteins.
The other theory maintains that the disease is essen-
tially a response of the vessel wall to injury and
although it plays an important part in lesion
progression, the deposition of lipid is a secondary
phenomenon. Both theories are supported by experi-
mental findings in animals that indicate that lesions,
which are morphologically similar to atherosclerotic
lesions in humans, can be induced by either a supple-
mentation of dietary lipid or by injury. Regardless of
experimental setting, the design of the experiments has
raised questions regarding the relevance of the results
to human atherogenesis."
I nj ur y t o ar t er i al endot hel i al cells, aggr egat i on of
pl at el et s and rel ease of pl at el et - der i ved gr owt h f act or
( P DGF ) and subsequent pr ol i f er at i on of s moot h
muscl e cells and t hei r conver s i on t o l i pi d-fi l l ed f oam
cells event ual l y resul t in t he i nt i mal t hi ckeni ng seen
in at her oscl er osi s. Resear ch f r om Yagi ' s l a bor a t or y
on l i pi d per oxi des and f r om Tayl or ' s l a bor a t or y on
chol est er ol oxi des ma y account f or ma ny o f t he key
steps in at her ogenesi s.
Recent st udi es r evi ewed by Maj no, Jor i s & Za nd
(1985) st r ongl y suggest t hat a l i nk has been di scov-
ered bet ween at her oscl er osi s and i nf l ammat i on. I f
t rue, t he st udi es of Bar anows ki et al . (1982), showi ng
t hat oxyst er ol s pr oduce gr eat er i nf l ammat i on t han
pur e chol est er ol , become rel evant .
Pl at el et aggr egat i on is a key f act or in CAD. Recent
st udi es have de mons t r a t e d t hat c o - 3 f at t y aci ds
f r om fish oils are ver y useful in r educi ng t he t endency
of pl at el et s t o aggr egat e (Sanders, 1983) as well as in
l ower i ng ser um t ri gl yceri des, chol est er ol and even
possi bl y L DL ( I l l i ngwor t h, Har r i s & Connor , 1984).
Fi sh oils in t abl et f or m are bei ng used by t he medi cal
pr of essi on t o accompl i s h benefi ci al changes in ser um
lipids in pat i ent s at risk f r om CAD. However , by
t hei r very nat ur e, 0 9 - 3 f at t y aci ds ar e very sus-
cept i bl e t o aut oxi dat i on. Fi sh oils ar e also ri ch in
chol est er ol . The qual i t y of fish oil pr epar at i ons vari es
widely. Recent ( unpubl i shed) studies in our l abor a-
t or y suggest t hat some of t he oils avai l abl e ar e of
poor qual i t y and t hat t hese have si gni fi cant levels of
chol est er ol oxi dat i on pr oduct s. Such fish oils woul d
al so be c ont a mi na t e d wi t h fat t y aci d hydr oper oxi des
and mal onal dehyde and coul d concei vabl y i ncur
si gni fi cant losses of t he e9 - 3 f at t y aci ds so benefi ci al
t o hyper l i pi daemi c pat i ent s.
It is appar ent t hat much r esear ch r emai ns t o be
done t o pr ot ect t he publ i c f r om levels of lipid ox-
i dat i on pr oduct s del et er i ous t o heal t h. Per haps wi t h
mor e r esear ch we will ar r i ve at a level of concer n t hat
i nvokes r egul at or y act i on agai nst pr oduct s cont ai n-
i ng si gni fi cant levels of l i pi d oxi dat i on pr oduct s.
Gi ven t he power f ul ar r ay of ant i oxi dant s and pr o-
cessi ng t echni ques avai l abl e, t he appear ance of hi gh
levels of oxi dat i on pr oduct s in f oods shoul d be a
ma j or concer n t o t he f ood i ndust r y and gover nment al
r egul at or y bodi es.
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QUESTI ONS AND ANSWERS
V. S i n g h , Ho f f ma n - L a Ro c h e : Dr Addi s , do you ha ve a ny d a t a or ar e t he r e a ny d a t a a va i l a bl e t o i ndi cat e
t h a t t hese oxi da t i on p r o d u c t s of c hol e s t e r ol ar e f or me d in vi vo i n h u ma n s , or ar e t hese al ways a va i l a bl e f r o m
f ood s our ces ? Secondl y, is t he r e a ny r ol e f or a n t i o x i d a n t s l i ke vi t a mi n C or E t o pr ot e c t a ga i ns t t hi s?
P. Ad d i s : To a ns we r y o u r fi rst ques t i on, t he r e ar e p a p e r s ci t i ng t he oc c ur r e nc e o f t hes e oxi da t i on p r o d u c t s
i n bi ol ogi cal f l ui ds a n d s er um, a n d al so i n ma mma r y gl a nd as pi r at es , whi c h wer e d o n e by Pe t r a kus at t he
Uni ve r s i t y of Ca l i f or ni a , Sa n Fr anci s co, Howe ve r , i n ma n y o f t hese cases, one is not sur e wh e t h e r t hey wer e
di et ar y or in vi vo pr oduc t s . Ce r t a i nl y i n t he case of in vi vo p r o d u c t i o n , a n t i o x i d a n t s c oul d pl ay a r ol e i n
s uppr e s s i ng oxi da t i on pr oduc t s . The p r o b l e m wi t h in vi vo me t a b o l i s m is t h a t t he l evel s ar e so mu c h l ower
t h a n wh a t we see i n f oods, t h a t i t ' s di ffi cul t t o wor k at s uch l ow levels.