Antioxidants: Their health benefits

Full text

© CAB International 2014. Phytochemicals of Nutraceutical Importance
248 (eds D. Prakash and G. Sharma)
16.1 Introduction
An important field of research today is the
control of ‘free radicals’ generation or redox’
status with the properties of food and food
components. Reactive oxygen species (ROS)
may interact with cellular macromolecules
and modify several cellular proteins, lipids
and DNA, which results in altered target
cell functions. Oxidative stress occurs in
a cell or tissue when the ROS generation
level exceeds the antioxidant capability
of that cell (Kumar et al., 2011). ROS can be
produced both endogenously and exoge-
nously. Endogenous oxidative stress can
be the result of normal cellular metabolism
and oxidative phosphorylation. Exogenous
sources of ROS can also impact on the over-
all oxidative status of a cell. Drugs, hor-
mones and other xenobiotic chemicals can
produce ROS by either direct or indirect
mechanisms (Kakkar and Singh, 2007).
Several human chronic disease states,
including cancer, have been associated with
oxidative stress produced through either an
increased free radical generation and/or a
decreased antioxidant level in the target
cells and tissues (Rice-Evans and Burdon,
1993). Natural antioxidants present in the
diet increase the resistance toward oxidative
damages and they may have a substantial
impact on human health. It has been
reported that a diet rich in antioxidant
phytochemicals, such as polyphenolics,
carotenoids, terpenoids and flavonoids, pro-
tects against cellular damage due to ability
to quench oxygen-derived free radicals
(Dhakarey et al., 2005; Singh, P., 2008; Singh,
B.N., 2009a). If antioxidant defence systems
are not sufficiently present in critical situa-
tions like oxidative stress, contamination,
UV exposure etc., the production of free
radicals increases significantly (Singh, U.
et al., 2008). Non-enzymatic (vitamin E,
vitamin C, glutathione (GSH), etc.) and
enzymatic (superoxide dismutase, GSH per-
oxides, glutathione-S-transferase and cata-
lase) antioxidant levels in the cell can be
decreased through modification in gene
expression, decreased antioxidant uptake
in the diet, or can be overloaded in ROS pro-
duction, which creates a net increase in the
amount of oxygen free radicals present in
the cell. It has been reported that with the
administration of antioxidants, cells are
protected against carcinogen-induced dam-
age (Kumar et al., 2011). Mechanisms of
protection could be effective against a wide
16 Antioxidants: Their Health
Benefits and Plant Sources
R.L. Singh,1* Sapna Sharma2 and Pankaj Singh1
1Department of Biochemistry, Dr RML Avadh University,
Faizabad, India; 2Division of Nephrology, Department of Medicine,
University of Chicago Medical Center, Chicago, USA
* Email: drrlsingh@rediffmail.com
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Antioxidants: Their Health Benefits and Plant Sources 249
range of dietary carcinogens possibly influ-
encing several cancer sites. Antioxidant
enzymes are detoxi fication/biotransforma-
tion enzymes that are involved in the
detoxification of toxic substances such as
xenobiotics, carcinogens, free radicals and
peroxides by conjugating these sub-
stances with GSH (Tripathi et al., 2010).
Traditional medicine all over the
world is nowadays being revalued by an
extensive amount of research on different
plant species and their therapeutic princi-
ples. Experimental evidence suggests that
free radicals (FR) and ROS can be involved
in a high number of diseases (Richards and
Sharma, 1991). As plants produce a lot of
antioxidants to control the oxidative stress
caused by sunlight and oxygen, they can
represent a source of new compounds
with antioxidant activity. One of the clini-
cal specialities of Ayurveda is Rasayana.
Rasayana is not only a drug therapy but
is a specialized procedure practised in the
form of rejuvenating recipes and dietary
regimen promoting good habit. The pur-
pose of Rasayana is two-fold: prevention
of disease and counteraction of ageing
processes which result from optimization
of homeostasis. The meaning of the word
Rasayana (rasa: essence, water; ayana:
going) essentially refers to nutrition and
its acquisition, movement, circulation and
perfusion in the body tissues (Singh, 1992).
With regard to Rasayana drug therapy,
Sharma et al. (1992) reported the strong anti-
oxidant activity of any Rasayana: these com-
pounds were found to be 1000 times more
potent than ascorbic acid, a-tocopherol
and probucol.
16.2 Antioxidants
In living cell, two antioxidant defence sys-
tem are present against free radical damage.
The first line of defence includes antioxidant
enzymes (such as superoxide dismutase,
catalase, GSH peroxidase), whereas the sec-
ond defence system includes low molecular
non-enzymatic antioxidants (thioredoxin,
GSH, vitamins A, C, E, lycopene, lutein,
quercetin etc.). These antioxidants inhibit
the formation of FRs by breaking the chain
reaction or can reduce the concentration
of FR by donating hydrogen and an elec-
tron. They also act as peroxide decomposer
(vitamin E), enzyme inhibitor, singlet oxygen
quencher (vitamin E), synergist and metal-
chelating agents (tranferritin). To provide
maximum intracellular protection, antioxi-
dants are strategically compartmentalized
throughout the cell. So that FR is produced
intracellular and extracellular during metab-
olism, both enzymatic and non-enzymatic
antioxidants are able to detoxify FRs.
Certain antioxidant enzymes (super-
oxide dismutase, catalase and GSH) are pro-
duced within the body. Other antioxidant
agents are found in foods, such as green
leafy vegetables, and it is believed that
diets rich in antioxidant (such as b-carotene
and vitamins A, C and E) are beneficial to
human health (Halliwell and Gutteridge,
1989). Therefore, antioxidant naturally pre-
sent in body or supplied in the form of diet
(phytonutrients) plays an important role
to control various diseases resulting from
oxidative stress. Fresh fruits and vegetables
are of more importance than cooked, because
of the high concentration and maximum
absorption of antioxidants. In recent years,
researchers have been researching the
relationship between antioxidants and pre-
vention of some diseases, such as cardiovas-
cular disease and cancer (Kubola and
Siriamornpun, 2008).
As soon as these FRs are generated in
the body, they are trapped by antioxidant
present in extracellular and intracellular
defence system. If the generation of free
radicals is much more than the concentra-
tion of antioxidants then oxidative stress
arises. As a result of oxidative stress, arthri-
tis in joints, emphysema and bronchitis in
lungs, atherosclerosis or heart disease in the
blood vessels, peptic ulcer in the stomach,
ageing and wrinkling in the skin are caused.
In the nucleus, it also alters the sequence
of nucleotide base pair, strand break etc. in
the DNA resulting in transformed and
mutated DNA. Mutated DNA will produce
diseases like cancer, leukaemia and lym-
phoma (Prakash et al., 2012).
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250 R.L. Singh et al.
16.2.1 Antioxidant enzyme
Three groups of enzymes play significant
roles in protecting cells from oxidative stress.
Superoxide dismutase
Superoxide dismutase (SOD) has been recog-
nized to play an important role in the body
defence mechanism against the deleterious
effect of superoxide FR in the biological sys-
tem. It acts on two superoxide molecules and
converts it into hydrogen peroxide and oxy-
gen. The beneficial aspect of this reaction is
that it produces less toxic hydrogen peroxide.
The organisms that resist oxygen toxicity
must have the SOD enzyme. On the basis
of metal cofactor, the organism has three
distinct types of SOD. In eukaryotes, cytosol
has the copper- and zinc-containing form
of SOD while mitochondria and bacterial
cells have the manganese-containing form of
SOD (Table 16.1). Iron-containing SOD is
found in bacteria, cynobacteria and some
plants. Newly discovered forms of SOD, also
found in bacteria, contain nickel as a cofactor.
Interestingly, SODs are inducible enzymes,
i.e. with the increase in the concentration of
oxygen in the environment of the cell, the
concentration of SOD enzyme also increases.
The main source of naturally occurring SOD
enzyme is green vegetables such as in barley,
broccoli, Brussels sprouts, cabbage, wheat and
most green plants (Gassen and Youdim, 1999).
Catalase
The catalase activity of mammalian tissue
varies greatly. It is highest in liver and kidney
and low in connective tissue. In the cell, it is
mainly particle bound (in mitochondria and
peroxisomes) whereas in erythrocytes it exist in
soluble state. Catalase activity received much
attention for its role in oxidative metabolism as
well as protective function by acting as H2O2
scavenger. Catalase located in the organelles
acts as regulator of H2O2 levels and, on the other
hand, in erythrocytes, catalase and GSH peroxi-
dase jointly exert a protective function for hae-
moglobin and other SH-protein. It degrades
hydrogen peroxide to water and oxygen, and
hence finishes the detoxification reaction
started by SOD (Gassen and Youdim, 1999).
Glutathione peroxidase
GSH peroxidase is a member of family of
GPx enzymes, whose function is to detoxify
Table16.1. Important enzymatic and non-enzymatic physiological antioxidants.
Antioxidants Location Properties
Enzymatic
Superoxide dismutase Mitochondria, cytosol Dismutase superoxide radicals
Glutathione peroxidase Mitochondria and cytosol Removes hydrogen peroxide and organic
hydroperoxides
Catalase Mitochondria and cytosol Removes hydrogen peroxide
Non-enzymatic
Vitamin E Cell membrane Chain-breaking antioxidant in cell membrane
Vitamin C Aqueous phase of
cell Sap
Acts as free radical scavenger and
recycles vitamin E
a-Lipoic acid Endogenous thiol Effective in recycling vitamin C, may also
be an effective glutathione substitute
Carotenoids Membrane tissue Scavengers of reactive oxygen species,
singlet oxygen quencher
Bilirubin Blood Extracellular antioxidant
Ubiquinones Mitochondria Reduced forms are efficient antioxidants
Metals ions sequestration:
transferrin, ferritin,
lactoferrin
Chelating metals ions, responsible for
Fenton reactions
Nitric oxide Free radical scavenger, inhibitor of LP
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Antioxidants: Their Health Benefits and Plant Sources 251
peroxide in the cell. Peroxides decompose to
form highly reactive free radicals, which can
damage the macromolecules like protein,
DNA and lipid. GPx enzyme plays an impor-
tant role in the protection of cell from this
damage, particularly lipid peroxidation. GSH
peroxidase contains selenium as a cofactor.
The synthesis of GSH peroxidase in humans
appears to be very important in scavenging
H2O2 (Cheng et al., 2003).
16.2.2 Antioxidant phytochemicals
There are more than a thousand phytochemi-
cals that have been identified with antioxidant
properties. Plants produce these chemicals to
protect themselves from microorganism and
oxidative stress, but now several evidences
suggest that these phytochemicals also protect
humans against various diseases caused by
FRs. Some of the well-known phytochemicals
are lycopene (tomatoes), isoflavones (in soy),
flavanoids (in fruits, vegetables), allyl sulfides
(onions, leeks, garlic), carotenoids (fruits, car-
rots) and polyphenols (tea, grapes). Medicinal
plant parts are commonly rich in phenolic
compounds, such as flavonoids, phenolic
acids, stilbenes, tannins, coumarins, lignans
and lignins. These compounds have multiple
biological effects including antioxidant activ-
ity (Shukla et al., 2009). The antioxidant activ-
ity of phytochemicals is mainly due to their
redox properties, which can play an impor-
tant role in adsorbing and neutralizing free
radicals, quenching oxygen, or decomposing
peroxides.
Flavonoids
Flavonoids are the most common secondary
metabolites in higher plants, and can directly
scavenge the superoxide ion, hydroxyl radi-
cal and H2O2. These include more than
4000 phenolic compounds that occur natu-
rally in plants.
Flavonols
The main flavonol is quercetin, followed
by myricetin, kaempferol, laricitrin, isor-
hamnetin and syringetin. The main sources
of flavonols are onion, kale, broccoli, lettuce,
tomato, apple, grape, berries, tea and red
wine. High contents of flavonols are pre-
sent in greener leaves (Manach et al., 2004).
Flavonols have multiple biological health
benefits. It reduces risk of cardiovascular
diseases, cancer, improve endothelial
function and reduce platelet activity. This
property is mainly attributed due to their
antioxidant properties (Patel, 2008). Furthe-
rmore, flavonols also help to prevent oxida-
tive damage to cells, lipids and DNA. The
antioxidant properties of flavonols are
drawn from the presence of aromatic rings
of the flavonoid molecule, which allows the
donation and acceptance of electrons from
FR species.
Anthocyanins
Anthocyanins are violet, blue and purple pig-
ments, which are mainly present in fruits,
berries and flowers. The major dietary antho-
cyanins include cyanidin, delphinidin, malvi-
din, pelargonidin, peonidin and petunidin
(Manach et al., 2004). Anthocyanins and their
derivatives have the capacity to scavenge FRs
through a number of mechanisms, thereby
reducing the oxidative stress. Anthocyanins
present in red cabbage reduce the oxidative
stress caused by the toxin paraquat (Igarashi
et al., 2000). Tsuda (2000) reported that cyani-
din, which is found in most fruit sources, has
potential antioxidant activity under in vivo
conditions. In another animal study, Tsuda
(1998) reported that cyanidins protect cell
membrane lipids from oxidation by a variety
of harmful substances.
Tannins
Tannins are commonly present in fruits
(grapes, persimmon, blueberry, etc.), tea,
chocolate, legume forages and legume trees
(Acacia sp., Sesbania spp. etc.) and grasses
(sorghum, maize, etc.). Tannins include proan-
thocyanidins, gallotannins and ellagitannins.
At high temperatures in alcohol solutions
or in a strong mineral acid, proanthocyanidins
release anthocyanidins, which have antioxi-
dant properties. Gallotannins and ellagitannins
are both hydrolysable tannins. Gallotannins
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252 R.L. Singh et al.
constitute galloyl esters of glucose or quinic
acid whereas ellagitannins are derivatives of
hexahydroxydiphenic acid (HHDP). Another
form of tannin is phloroglucinols, which are
subunits of phlorotannins and present in
marine brown algae only. Tannins give an
astringent or bitter taste to foods and bever-
ages (e.g. some red wines, teas and unripe
fruits). The basic function of tannin is not as a
primary antioxidant (i.e. they donate hydro-
gen atom or electrons) but they act as second-
ary antioxidants (i.e. interfere with the chain
reaction or by chelating the metal ions such as
Fe(II) thereby retarding oxidation or Fenton
reaction). Zhang et al. (2004) showed that
the inhibition of lipid peroxidation by tan-
nin constituents can act via the inhibition of
cyclooxygenase.
Phenolic acids
Phenolic acids are a major class of phenolic
compounds, widely occurring in the plant
kingdom. Predominant phenolic acids
include hydroxybenzoic acids (e.g. gallic
acid, p-hydroxybenzoic acid, protocatechuic
acid, vanillic acid and syringic acid) and
hydroxycinnamic acids (e.g. ferulic acid, caf-
feic acid, p-coumaric acid, chlorogenic acid
and sinapic acid) (Wrigstedt et al., 2010).
Ferulic, caffeic and p-coumaric acid are pre-
sent in many medicinal herbs and dietary
spices, fruits, vegetables and grains. Wheat
bran is a good source of ferulic acids.
Hydroxycinnamic acids (non-flavonoid phe-
nolics) are characterized by the C6–C3 struc-
ture. Plants use these compounds in both
structural and chemical defence strategies
against microbial flora as well as oxidative
stress (Cartea et al., 2011). Naturally occur-
ring hydroxycinnamic acids possess high
level of antioxidants in comparison to
hydroxybenzoic acid due to increased possi-
bilities for delocalization of the phenoxy rad-
ical (Beer et al., 2002). Phenolic compounds
have the potential to function as antioxidants
by scavenging the superoxide anion, hydroxyl
radical, peroxy radical or quenching singlet
oxygen and inhibiting lipid peroxidation in
biological systems (Izunya et al., 2010). At
low temperatures during the maturity of
leaves, the leaves have been shown to
increase the phenols and flavonoids content
(Singh, P. et al., 2008; Singh, B.N., 2009c).
16.3 Antioxidant Nutrients
16.3.1 Vitamin E
Vitamin E is the main lipid-soluble antioxi-
dant and plays a vital role in protecting
membranes from lipid peroxidation. Primary
function of vitamin E is to trap peroxy radi-
cal formation during lipid peroxidation
in cellular membranes. It is mainly present
in nuts, seeds, vegetables, fish oils, whole
grains (especially wheat germ), fortified
cereals and apricots (Glenville, 2006). Current
recommended daily allowance (RDA) is
15 IU day−1 for men and 12 IU day−1 for women.
16.3.2 Vitamin C or ascorbic acid
Vitamin C or ascorbic acid is a water-soluble
antioxidant that can reduce a variety of free
radicals. It acts as a synergist for tocopherol
by converting the oxidized tocopherols
back to their reduced status. Ascorbic acid
can also act as a pro-oxidant under certain
circumstances and helps regeneration of
membrane-bound oxidized vitamin E.
Vitamin C reacts with the a-tocopheroxyl
radical and is oxidized to dehydroascorbic
acid. Humans lack L-gulono-g-lactone oxi-
dase, which is a key enzyme in ascorbic acid
synthesis, hence it cannot be synthesized in
the body and must be acquired from dietary
sources. Ascorbic acid is mainly present in
citrus fruits and juices, kiwi, cabbage, green
peppers, spinach, broccoli, kale, cantaloupe
and strawberries. The RDA for vitamin C is
60 mg day−1. If taken in high dosages it may
be excreted out due to its water-soluble
nature but may cause adverse side effects in
some individuals. The efficiency of ascorbic
acid as scavenger of superoxide in mamma-
lian tissue is not less than the SOD enzyme.
The ascorbic acid level in extracellular fluids
is higher than those of glutathione. So, ascor-
bate probably plays a predominant role in
extracellular antioxidant protection. Vitamin C
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Antioxidants: Their Health Benefits and Plant Sources 253
reacts with the superoxide radical to form
dehydroascorbic acid and it returns to its
original state (vitamin C) with the help of
gluthathione (Prakash et al., 2012).
16.3.3 Glutathione
Glutathione, a tripeptide (glutamyl-cysteinyl-
glycine) antioxidant, is the most important
intracellular defence against damage by
ROS. It is widely distributed among living
cells and apparently involved in many bio-
logical functions. Glutathione present in the
oxidized (GSSH) form is converted to the
reduced GSH by enzyme glutathione reduc-
tase. It has been reported that reduced GSH is
mainly present in tissue. The free sulfhydryl
(SH) is a very reactive group in cysteine, pro-
viding target for radical attack. Reduced
glutathione is oxidized when it reacts with
free radicals and it gets back to the reduced
state by redox cycle involving GSH reduc-
tase and the electron acceptor NADPH
(Gassen and Youdim, 1999).
16.3.4 Selenium
Selenium, an essential element for antioxida-
tion reactions, is required only in small
amounts in humans and animals (Thomson,
2004). Selenoproteins (proteins containing
selenium) are important antioxidant enzymes.
There are nearly 30 known selenoproteins,
mainly containing selenocysteine. The active
site of GSH peroxidase (the most abun-
dant selenoprotein in mammals) and thiore-
doxin reductase enzyme has selenocysteine.
Thioredoxin reductase not only maintains cell
proteins in a reduced state but also provides
deoxyribonucleases required for DNA syn-
thesis (Holmgren, 1989). At low concentra-
tions it acts as an antioxidant, inhibiting lipid
peroxidation, whereas at higher concentra-
tions it behaves as pro- oxidant, enhancing the
accumulation of lipid peroxidation products.
The antioxidant properties of selenoproteins
help to regulate thyroid function, play impor-
tant role in the immune system and prevent
cellular damage from free radicals (Corvilain
et al., 1993). Selenium deficiency may cause a
form of heart disease, hypothyroidism and a
weakened immune system (Zimmerman and
Kohrle, 2002).
16.3.5 b-Carotene
b-carotene (precursor to vitamin A, retinol) is
present in liver, egg yolk, butter, milk, spin-
ach, squash, carrots, broccoli, tomato, yams,
cantaloupe, peaches and grains. b-carotene is
converted to Vitamin A by the body. The
carotenoids (fat-soluble antioxidant) are one
of the most common pigments found in
nature (Daun, 1988). b-carotene (one of the
best known carotenoids) is necessary for the
synthesis of vitamin A. Some other related
pigments include a-carotene, lutein, lycopene
and astaxanthin. There is evidence that diet
containing fruit and vegetables is associated
with lower incidences of cancer (Giovannucci,
1999). b-carotene has the capacity to quench
reactive oxygen (stop oxidative mechanisms),
making them chemoprotective against cancer.
There is strong evidence that b-carotene
increases the detoxification of carcinogens
present in the liver, thereby reducing the
development of cancer (Solomons, 2001).
16.3.5 Metal-binding protein
Transition metals are tightly bound to vari-
ous proteins that prevent them from reacting
with peroxides to form free radicals. These
include the following.
Ceruloplasmin
Ceruloplasmin is an effective antioxidant with
potent peroxidase property. It decomposes
hydrogen peroxide in the presence of reduced
glutathione. Ceruloplasmin is expressed mainly
in the liver but has been found to be expressed
in the lungs (Fleming et al., 1991) and mam-
mary glands. The role of ceruloplasmin as
antioxidant is against organic and inorganic
oxygen radicals from iron and ascorbate.
It contains 90–95% of the circulating copper
in normal mammals. The concentration of
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254 R.L. Singh et al.
ceruloplasmin increases by a factor of 2 to 3
during pregnancy and hormonal conditions.
It also inhibits lipid peroxidation induced by
ferrous ion by way of decomposing lipid per-
oxides (Verma et al., 2005).
Lactoferrin
Lactoferrin belongs to the iron transporter or
transferrin family of glycoproteins and is
mainly present in whey and exocrine secre-
tions from mammals and is released from
neutrophil granules during inflammation.
Human breast milk may contain as much as
15% lactoferrin while cow’s milk may have
only 0.5% to 1.0%. It has two important roles:
(i) it shows antibacterial, antiviral, antifungal,
anti-inflammatory, antioxidant and immu-
nomodulatory activities; and (ii) lactoferrin
plays an important role in the uptake and
absorption of iron through the intestinal
mucosa. Its ability to bind iron probably con-
tributes to both its antioxidant properties and
its antibacterial action (Gupta et al., 2012).
Metallothionein
Metallothionein (MT) consists of four low-
molecular-weight (6000–7000), metal-binding
proteins with high cysteine content. Metal-
lothioneins (MTs) are sulfhydryl-rich pro-
teins, which specifically neutralize hydroxyl
radicals (Viarengo et al., 2000). Antioxidant
properties of MTs are mainly due to sulfhy-
dryl nucleophilicity. In vitro studies have
revealed that it reacts directly with ROS
including superoxide and hydroxyl radicals
and hydrogen peroxide. Binding of transition
metals (Fe, Cu) to the protein reduce the
Fenton reactivity, resulting in reduced oxida-
tive stress.
Transferrin
Transferrin (iron-binding blood plasma gly-
coprotein) has a molecular weight of
approximately 80 kDa and bind iron very
tightly but reversibly and hence control the
level of free iron in biological fluids (Crichton
and Charloteaux-Wauters, 1987). It has two
specific high-affinity Fe(III) binding sites.
Iron present in the body is always found in
protein-bound form and never in a free state.
If iron is being transported or stored it must
be chelated in very specific ways by transfer-
rin or ferritin. Transferrin is mainly present
in serum, but it is also found in other body
fluids at lower concentrations (Chauhan et al.,
2004). The antioxidant activity of transferrin
is due to its reducing properties. It reduces
the concentration of free ferrous ion that
catalyses the conversion of hydrogen perox-
ide to highly toxic hydroxyl radical by Fenton
reaction. Transferrin is a universal iron carrier
and is able to deliver iron to cells without
formation of free radicals.
Ferritin
Ferritin (a globular protein complex consist-
ing of 24 protein subunits) is a ubiquitous
intracellular protein that stores iron and
releases it in a controlled fashion. Ferritin is
synthesized by almost all living organisms,
including algae, bacteria, higher plants and
animals. Intracellular iron is stored in the fer-
ritin in both prokaryotes and eukaryotes and
released into cells when needed; hence it acts
as buffer against iron deficiency. Ferritin that
is not combined with iron is called apoferri-
tin. Ferritin converts ferrous (Fe2+) to ferric
(Fe3+) form by ferroxidase activity, thereby
reducing the chance of the deleterious reac-
tion that occurs between ferrous iron and
hydrogen peroxide known as the Fenton
reaction, which produces the highly damag-
ing hydroxyl radical (Sarma et al., 2010).
16.4 Some Commonly Measured
Analytes with Antioxidant and
Pro-oxidant Activities
16.4.1 Gamma-glutamyltransferase
Gamma-glutamyl transpeptidase (also known
as g-glutamyltransferase, GGT, GGTP, gamma-
GT) (EC 2.3.2.2) is an enzyme that transfers
g-glutamyl functional groups. It is the first
enzyme of the g-glutamyl cycle that regulates
the antioxidant glutathione; hence it is a criti-
cal enzyme in glutathione homeostasis. GGT
is present in the cell membrane of many
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Antioxidants: Their Health Benefits and Plant Sources 255
tissues, including the kidney, bile duct, pan-
creas, gallbladder, spleen, heart, brain and
seminal vesicle (Sarma et al., 2010).
16.4.2 Uric acid
Uric acid, the end product of purine metabo-
lism, works as an antioxidant. It is the most
abundant aqueous antioxidant in humans
and contributes as much as two-thirds of all
free-radical scavenging capacity in plasma. It
is particularly effective in quenching hydroxyl,
superoxide and singlet oxygen and peroxyni-
trite radicals and may play a protective physi-
ological role by preventing lipid peroxidation.
The major antioxidant role of uric acid is its
ability to bind and inactivate peroxynitrite.
At physiological concentrations, urate pro-
tects erythrocyte ghosts against lipid peroxi-
dation leading to lysis of erythrocytes. Urate
is found to be about as effective an antioxi-
dant as ascorbate in these experiments. Urate
is much more easily oxidized than deoxynu-
cleosides by singlet oxygen and is destroyed
by hydroxyl radicals at a comparable rate
(Nieto et al., 2000).
16.4.3 Bilirubin
Bilirubin, the end product of haem metabo-
lism, has the ability to function as an antioxi-
dant in the brain, scavenging free radicals and
reducing oxidative damage. It is reported that
bilirubin protects oxidation of lipids such as
linoleic acid and vitamin A. Stocker et al. (1987)
demonstrated that bilirubin has more of an
antioxidant effect than vitamin E towards lipid
peroxidation. It has also been experimentally
proved that higher concentration of serum bili-
rubin increases its antioxidant capacity.
16.4.4 High-density lipoprotein
High-density lipoprotein (HDL) has long
been known as the ‘good cholesterol’, protect-
ing against heart disease and atherosclerosis.
It has been experimentally found that HDL
has powerful antioxidant properties, similar
to vitamin C and vitamin E. An enzyme related
to synthesis of HDL cholesterol, lecithin-
cholesterol acyltransferase, is a powerful anti-
oxidant enzyme that blocks the oxidization of
low-density lipoprotein (LDL) cholesterol.
Cholesterol is beneficial if it is not oxidized.
Barter et al. (2007) suggested that a low level
of HDL increases the risk of diseases even
in people with very low LDL levels. Jafri
et al. (2010) suggested that there is an inverse
relationship between high HDL and cancer
occurrence.
16.4.5 Nitric oxide
Nitric oxide is an uncharged lipophilic mole-
cule that behaves like amphoteric molecule,
i.e. NO could function as an electron donor
(oxidant) or an electron acceptor (antioxi-
dant) (Drew and Leeuwenburgh, 2002). It
contains a single unpaired electron (NO•),
which reacts with other molecules, such as
oxygen, GSH and superoxide radicals. They
prevent free radicals from stealing electrons
from other molecules.
16.5 Sources of Natural Antioxidants
Dietary antioxidants include ascorbate, toco-
pherols, carotenoids and bioactive plant
phenols. The health benefits of fruits and
vegetables are largely due to the antioxidant
vitamins supported by the large number of
phytochemicals, some with greater antioxi-
dant properties. Sources of tocopherols,
carotenoids and ascorbic acid are well
known and there are plenty of publications
related to their roles in health. Exogenous
dietary antioxidants capable of scavenging
free radicals are of great interest in combat-
ing oxidative stress-induced cell damage.
Plants containing a high content of polyphe-
nols and flavanoids are considered as poten-
tial antioxidants and can be used as adjuvant
therapy. These plant polyphenols and flava-
noids are multifunctional and can act as
reducing agents, hydrogen donors, singlet
oxygen quenchers and metal ion chelators
(Gassen and Youdim, 1999).
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256 R.L. Singh et al.
Several natural antioxidants such as sily-
marin, grape seed extract, resveratrol, cur-
cumin etc., are known to reduce oxidative
stress and protect from hepatic damage.
Ulusoy et al. (2012) reported antioxidant and
anti-apoptotic effects of proanthocyanidine
from grape seed extract. Silymarin, a flavonoid
complex from Silybum marianum, has been
used in the treatment of hepatitis, liver cirrho-
sis, viral hepatitis and fatty liver. It has been
shown to have antioxidant, antilipid peroxida-
tive, anti-inflammatory and liver regenerative
effects. Lupeol, a pentacyclic triterpenoid,
found in many plants such as crataeva, mango,
olive etc., received much attention due to its
wide spectrum of medicinal properties that
include antiprotozoal, anti-inflammatory, anti-
carcinogenic, cardioprotective and antimicro-
bial activities. Hepatoprotective action of
lupeol against aflatoxin B1-induced toxicity
has been reported by Preetha et al. (2006).
Cymbopogon citratus D. Stapf., commonly
known as lemongrass, contains volatile oil
consisting of citral, a monoterpene (a mixture
of two isomeric aldehydes, neral and geranial
in the ratio of 2:3), as a major component,
which is used in various perfume and cos-
metic industries (Rauber et al., 2005). The
plant is reported to possess antifungal, mos-
quito repellent, insecticidal, antidiabetic, anti-
septic, antimutagenic and anticarcinogenic
activity (Masuda et al., 2008).
Fumaria parviflora Lam. (Fp) is used for
dermatological diseases, stimulation of liver
function and gall bladder, as antiscabies, anti-
scorbite, antibronchite, diuretic, expectorant,
antipyretic, diaphoretic, appetizer and anti-
neoplastic agent. Its antinoceceptive effect
has also been worked out (Heidari et al.,
2004). Phytochemical analysis of Fp indicated
presence of organic acids and isoquinoline
alkaloids, namely: fumaric acid, protropine,
cryptopine, sinactine, stylopine, dihydro-
fumariline, per-fumidine and dihydrosan-
guirine (Suau et al., 2002). Acetyl-cholinesterase
and butyrylcholinesterase inhibitory activity
of Fp has also been reported (Orhan et al.,
2004). Significant oral antipyretic activity has
been shown by hexane-chloroform and water-
soluble extracts of Fp in rabbits (Akhtar et al.,
1984). A 50% ethanolic extract of Fp was also
tested to discover the role of mitochondria
and ROS/oxidative stress in cytotoprotective
and anti-apoptotic effects against nimesulide-
induced hepatotoxicity (Tripathi et al., 2010).
Glycyrrhiza glabra (liquorice) possesses
triterpene, saponins, glycyrrhizin/glycyrrhi-
zic acid and glycyrrhetic acid. Glycyrrhizic
acid (GA), a biologically active constituent of
liquorice root with a structure of 20b-carboxy-
11-oxo-30-norolean-12en-3-b-yl-2-o-b-D-glu-
copyranosiduronic acid, is believed to be partly
responsible for anti-ulcer, anti-inflammatory,
antidiuretic, anti-epileptic, anti-allergic, anti-
dote, antitumour, antiviral, antihypotensive
and several other properties of the plant
(Baltina, 2003). Hypocholesterolaemic and
hypoglycaemic activities have also been
reported (Sitohy et al., 1991).
Bacopa monnieri Linn. (syn. Herpestis mon-
nieri Linn. H.B. and K) is used as a nerve tonic,
brain tonic, memory enhancer, laxative, astrin-
gent, antipyretic, anti-inflammatory and lep-
rosy healer. It is also useful in renal disorders,
blood diseases, cough, anaemia and poison-
ing. The plant also finds various applications
in central nervous system depressant activity.
Its major constituents including two saponins
(bacoside A and bacoside B) have been isolated
and characterized (Chowdhuri et al., 2002).
Geraniol, an acyclic monoterpenoid, is an
important constituent of essential oils of gin-
ger, lemon, lime, lavender, nutmeg, orange,
rose and palmarosa. It is reported to prevent
cancer. Camphene, another component, is a
bicyclic monoterpene with a pungent smell.
It constitutes a minor part of many essential
oils including turpentine oil, cypress oil, cit-
ronella oil, ginger oil etc., and is known to
possess antilithic and expectorant properties.
Camphene is also present in apricot, carrots,
cinnamon, ginger, cumin seed, nutmeg, car-
damom and turmeric. It is used as a food
additive for flavouring as well as in the prep-
aration of fragrances, plasticizers for resins
and lacquers (Verschueren, 2001).
Free radicals generated in diabetes may
lead to several kinds of diabetic complica-
tions including nephropathy, neuropathy,
cardiopathy and many more. Many herbal
medicines as single agents or in different
oral formulations have been recommended
for diabetes mellitus due to the fact that
they are less toxic than oral hypoglycaemic
Prakash_Ch16.indd 256 11/27/2013 10:07:55 AM

Antioxidants: Their Health Benefits and Plant Sources 257
agents such as sulfonylureas, metformin, etc.
(Ponnachan et al., 1993).
Anthocyanins have been shown to be
natural anti-inflammatory agents and pain
relievers. Chronic inflammation has also
been associated with an increased risk of can-
cer, but anti-inflammatory drugs are not
effective for reducing this type of inflamma-
tion (Singh, B.N. et al., 2009b). Some impor-
tant sources of antioxidants are presented in
Table 16.2.
16.6 Roles of Antioxidants in the
Prevention of Diseases
Plants have numerous natural antioxidants to
control the oxidative stress induced by these
free radicals (Pacher et al., 1997; Sarma et al.,
2010). Free radicals have been implicated in the
pathogenesis of over 100 human diseases such
as cancer, heart disease, stroke, Alzheimer’s
disease, diabetes, premature ageing, high
blood pressure and sepsis, to name a few.
16.6.1 Cancer
Antioxidants protect DNA thereby reducing
the oxidative DNA damage caused by the
free radical and ultimately control the
increased abnormal cell division, the main
characteristic of carcinogenesis. Experimenta l
evidence using cell culture and animal mod-
els indicate that antioxidants either slow or
prevent the development of cancer through
its action as free-radical scavenger (Rock
et al., 1996). Using in vitro and an animal
model system, it was experimentally found
that plant-derived phytochemicals, such as
allyl sulfides, isothiocyanates and sulfora-
phene, inhibit the various step of tumour
development (Milner, 1994). Blot et al. (1993)
and Sardas (2003) reported that a combina-
tion of b-carotene, vitamin E and selenium
significantly reduces the chance of cancer
development especially in the case of gastric
cancer. Experimental evidence also suggests
that b-carotene with a-tocopherol/retinol
significantly reduced the chance of lung can-
cer (Omenn et al., 1994).
16.6.2 Alzheimer’s disease
Alzheimer’s disease (AD) is characterized by
progressive loss of memory as the major clini-
cal manifestation. Studies on free radicals
suggest that oxidative stress causes neurode-
generative disorders, including AD. Metal
ion also plays an important role in the devel-
opment of AD. Nutraceutical antioxidants
such as b-carotene, curcumin, lutein, lyco-
pene, turmerin etc., showed positive effects
by reducing oxidative stress, mitochondrial
dysfunction and various forms of neural
degeneration (Glenville, 2006). It has been
observed that a lower activity of antioxidant
enzyme such as superoxide dismutase is
related to occurrence of Alzheimer’s disease
in humans (Thome et al., 1997). Kontush et al.
(2001) reported that supplementation with
vitamins E and C to the patient significantly
increases the concentration of vitamins in
plasma and decreases the oxidation of lipo-
protein, while vitamin E alone does not have
any significant effects. High intake of nutraceu-
tical postpones the development of demen-
tias such as AD (Haider and Bhutta, 2006).
16.6.3 Atherosclerosis
Atherosclerosis is a common cardiovascular
disease, which occurs due to deposition of
oxidized fatty acid to the arteries in the form
of plaque. Approximately two-thirds of the
serum cholesterol pool in a normal subject is
low-density lipoprotein-cholesterol (LDL-C),
which is believed to play an important role
in the development of atherosclerosis (Shukla
et al., 2011).
Flavonoids and other plant-derived poly-
phenols, present in fresh fruits and vegetables,
have been shown to be powerful antioxidants
capable of preventing LDL oxidation induced
by free radicals. Recommended daily allow-
ance for the flavonoids is 1 g in an ordinary
diet, which is sufficient for the antioxidant
defence system. Interestingly, it has been
found that the antioxidant activity of some of
flavonoids synergistically increases when they
are supplemented with ascorbic acid to pre-
vent LDL oxidation. The beneficial properties
Prakash_Ch16.indd 257 11/27/2013 10:07:55 AM

258 R.L. Singh et al.
Table 16.2. Some important sources of antioxidants.
Plant Antioxidants References
Medicinal plants
Terminalia chebula (Bahera) Casuarinin, chebulanin and chebulinic acid Cheng et al., 2003
Cassia fistula (Amaltas) Lupeol, b-sitosterol, hexacosanol, kaempferol, proanthocyanidin,
bianthraquinone glycoside, anthraquinones, flavonoids,
flavan-3-ol derivatives, sennoside A, sennoside B
Akiremi et al., 2000
Withania somnifera
(Ashwagandha)
Withanolides, cuscohygrine, anahygrine, tropane, pseudotropine,
anaferine, dl-iso-pllatierine, withanine, withasominine, withaninine,
somniferin, pseudowithanine, tropanol, pseudotopanol,
cuscokygrene, 3-tigioyloxytropana, isopelletierine
Sangwan, 2004; Mohammad and Elisabeth, 2009;
Kushwaha and Karanjekar, 2011
Fruits
Berries (Sarashphal) Flavanols, hydroxycinammic acids, hydroxybenzoic acids,
anthocyanins
Wang and Lin, 2000; Yanishlieva-Maslarova and
Heinonen, 2001
Citrus fruits Flavanones, flavonols, phenolic acids Yanishlieva-Maslarova and Heinonen, 2001;
Manach et al., 2004
Black grapes Anthocyanins, flavonols Belitz and Grosch, 1999; Yanishlieva-Maslarova
and Heinonen, 2001
Cherries Hydroxycinnamic acids, anthocyanins Belitz and Grosch, 1999; Yanishlieva-Maslarova
and Heinonen, 2001
Plums (Jamun), apples,
pears
Hydroxycinnamic acids, catechin Belitz and Grosch, 1999; Yanishlieva-Maslarova
and Heinonen, 2001
Vegetables
Allium sativum (Garlic) Aliin, allicin, ajoene, allylpropyl disulfide, diallyl trisulfide, sallylcysteine,
vinyldithiines, S-allylmercaptocystein, S-allylcysteine, S-allyl
mercaptocysteine and saponins
Kemper, 2000; Amagase, 2006
Allium cepa (Onion) Phenolic acids, flavonoids, cepaenes, thiosulfinates, anthocyanins,
sulfur compounds, saponins, quercetrin
Singh, B.N. et al., 2009a; Panduranga Murthy
et al., 2011
Trigonella foenum-graecum
(Fenugreek)
Coumarin, fenugreekine, nicotinic acid, sapogenins, phytic acid,
scopoletin, trigonelline, L-tryptophan-rich proteins and saponins
Yoshikawa et al., 1997
Daucus carota (Carrot) Carotol, daucene, germacrene D, bergamotene, selinene, carotol,
daucol, copaenol
Ozcan and Chalchat, 2007
Sweet potato leaves Flavonols, flavones, Chu et al., 2000
Yellow onion Flavonols Manach et al., 2004
Beans Flavanols Manach et al., 2004
Spinach Flavonoids, p-coumaric acid Bergman et al., 2001
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Antioxidants: Their Health Benefits and Plant Sources 259
Flours
Oats, wheat, rice Caffeic, ferulic acids Yanishlieva-Maslarova and Heinonen, 2001
Drinks
Orange juice Flavanols Manach et al., 2004
Coffee Hydroxycinnamic acids Manach et al., 2004
Chocolate Flavanols Manach et al., 2004
Red wine Flavan-3-ols, flavonols, anthocyanins Manach et al., 2004
Herbs and spices
Sage, carnosol Carnosic acid, lateolin, rosmanul, rosmarinic acid Yanishlieva-Maslarova and Heinonen, (2001)
Foeniculum vulgare (Fennel) Essential oil (trans-anethole, α-phellandrene, α-pinene), dipentene,
methyl chavicol, feniculun, anisaldehyde and anisic acid
Piccaglia and Marotti, 2001; Mimica-Dukic et al.,
2003; Araque et al., 2007
Rosemary Carnosic acid, carnosol, Rosmarinic acid rosmanol Yanishlieva-Maslarova and Heinonen, 2001;
Ibanez et al., 2003
Thyme Thymol, carvacrol, flavonoids, lubeolin Exarchou et al., 2002
Ginger Gingerol and related compounds Moure et al., 2001; Yanishlieva-Maslarova and
Heinonen, 2001
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260 R.L. Singh et al.
of certain plants may be explained by the pres-
ence of some especially effective flavonoids
like resveratrol, which has also been found in
red wines. Probucol, a hypocholesterolaemic
drug, has significant antioxidant activity and
in vivo study on rabbit showed that it has pro-
tective effects against atherosclerosis. In ani-
mal studies, aspirin has also been shown to
prevent atherosclerosis (Jaichander et al., 2008).
16.6.4 Heart diseases
There are several factors such as high choles-
terol level, hypertension, diabetes, cigarette
smoking etc. that provide a platform for the
development of heart disease. Oxidation of
low density lipoprotein (LDL-cholesterol)
causes deposition of fatty acid in arteries lead-
ing to development of atherosclerosis, which
ultimately causes heart disease (Anderson
et al., 1995). Heart disease is acquired with age
because oxidized fatty acid gets more ‘sticky’
and easier to adhere to the artery walls. It is
believed that high intake of ascorbic acid
reconstitutes the endothelial dysfunctions
(Ting et al., 1997) and protects the circulating
lipoprotein from free radicals.
16.6.5 Diabetes
Diabetes mellitus (DM) is characterized by
hyperglycaemia (Grill and Bjorklund, 2000).
Oxidative stress due to lack of antioxidant
defences may also cause diabetes (Cross et al.,
1987; Maxwell et al., 1997; Keaney and
Loscalzo, 1999; Bonnefont-Rousselot et al.,
2000; West, 2000). It is hypothesized that if
ROS are involved in the genesis of diabetes,
then antioxidants may be an effective approach
in prevention of diabetes (Giugliano et al.,
1996). Reaven (1995) revealed that supplemen-
tation of vitamin E reduces the sensitivity of
LDL to in vitro oxidation and availability of
oxidized LDL in type-2 diabetics as well as in
healthy subjects (Liao et al., 1995). It is hypoth-
esized that imbalance between generation and
scavenging of free radicals is the main cause
associated with diabetes. Insulin increases
the uptake of vitamin C in to the cell but in
hyperglycaemic conditions this process is
inhibited resulting in a condition known as
‘tissue scurvy’. Supplementation of vitamin C
alone controls the blood glucose level,
improves endothelium-dependent vasodila-
tion and increases the resistance of lipoprotein
towards oxidation in the patient with either
type-1 or type-2 diabetes mellitus (Ting et al.,
1996; Timimi et al., 1998; Kawano et al., 1999).
16.6.6 Parkinson’s disease
Parkinson’s disease (PD) results from damage
in neuronal cells in certain regions of the
brain, and is characterized by muscle rigidity,
shaking and difficulty in walking (Losso,
2003). Latif et al. (2007) reported that vitamin E
in food may be protective against PD.
Glutathione has also shown some promising
results in preliminary studies to treat PD but
appropriate long-term dosing, side-effects and
the most effective method of administration
are not yet clear.
16.7 Conclusions
Antioxidants may be a promising source for
the prevention and or treatment of free radical-
generated diseases such as atherosclerosis,
hypertension, diabetes, cancer, Parkinson’s
and Alzheimer’s diseases etc. Evidence also
indicates that antioxidants protect/cure the
diseases by involving a number of biological
processes, including signal transduction path-
ways, activation of antioxidant defences, cell
proliferation, cell survival-associated gene
expression, differentiation and preservation
of mitochondrial integrity. To protect the cells
and organ systems of the body against reac-
tive oxygen species, humans have evolved a
highly sophisticated and complex antioxidant
protection system. It involves a variety of anti-
oxidant components, both endogenous and
exogenous in origin, that function interac-
tively and synergistically to neutralize free
radicals. Increasing dietary intake of antioxi-
dants may help to maintain an adequate anti-
oxidant status and, therefore, the normal
physiological function of human beings.
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Antioxidants: Their Health Benefits and Plant Sources 261
References
Akhtar, M.S., Khan, Q.M. and Khaliq, T. (1984) Effects of Euphorbia prostrata and Fumaria parviflora in nor-
moglycemic and alloxan-treated hyperglycaemic rabbits. Planta Medica 50, 138–142.
Akiremi, A.A., Omobuwajo, O.R. and Elujoba, A.A. (2000) Pharmcopieal standards for the fruits of Cassia
fistula and Cassia podocarpa. Nigerian Journal of Natural Products and Medicine 4, 23–26.
Amagase, H. (2006) Clarifying the real bioactive constituents of garlic. Journal of Nutrition 136, 716S–725S.
Anderson, T.J., Meredith, I.T., Yeung, A.C., Frei, B., Selwyn, A.P. and Ganz, P. (1995) The effect of cholesterol-
lowering and antioxidant therapy on endothelium-dependent coronary vasomotion. The New England
Journal of Medicine 332, 488–493.
Araque, M., Rojas, L.B. and Usubillaga, A.D. (2007) Antibacterial activity of essential oil of Foeniculum
vulgare miller against multi resistant gram-negative bacilli from nosocomial infections. Ciencia 15,
366–370.
Baltina, L.A. (2003) Chemical modification of glycyrrhizic acid as a route to new bioactive compounds for
medicine. Current Medicinal Chemistry 10, 155–171.
Barter, P., Gotto, A.M., LaRosa, J.C., Maroni, J., Szarek, M., Grundy, S.M., Kastelein, J.J.P., Bittner, V., Fruchart, J.C.
and Treating to New Targets Investigators (2007) HDL cholesterol, very low levels of LDL cholesterol,
and cardiovascular events. New England Journal of Medicine 357, 1301–1310.
Beer, D.D., Joubert, E., Gelderblom, W.C.A. and Mandey, M. (2002) Phenolic compounds: a review of their
possible role as in vivo antioxidant of wine. South African Journal of Enology 23, 48–61.
Belitz, H.D. and Grosch, W. (1999) Phenolic compounds. In: Food chemistry, 2nd edn. Springer, Berlin,
pp. 764–775.
Bergman, M., Vershavsky, L., Gottlieb, H.E. and Grossman, S. (2001) The antioxidant activity of aqueous
spinach extract: chemical identification of active fractions. Phytochemistry 58, 143–152.
Blot, W.J., Li, J.Y., Taylor, P.R., Guo, W., Dawsey, S., Qingang, G., Yang, C.S., Zheng, S.F., Gail, M., Li, G.Y.,
Yu, Y., Liu, B., Tangrea, J., Sun, Y.H., Liu, F., Fraumeni, J.F., Zhang, Jr Y.H. and Li, B. (1993) Nutrition
intervention trials in Linxian, China: supplementation with specific vitamin/mineral combinations, can-
cer incidence, and disease-specific mortality in the general population. Journal of the National Cancer
Institute 85, 1483–1491.
Bonnefont-Rousselot, D., Bastard, J.P., Jaudon, M.C. and Delattre, J. (2000) Consequences of the diabetic
status on the oxidant/antioxidant balance. Diabetes & Metabolism 26, 163–176.
Cartea, M.E., Francisco, M., Soengas, P. and Velasco, P. (2011) Phenolic Compounds in Brassica Vegetables.
Molecules 16, 251–280.
Chauhan, A., Chauhan, V., Brown, W.T. and Cohen, I. (2004) Oxidative stress in autism: Increased lipid per-
oxidation and reduced serum levels of ceruloplasmin and transferrin – the antioxidant proteins. Life
Sciences 75, 2539–2549.
Cheng, H.Y., Lin, T.C., Yu, K.H., Yang, C.M. and Lin, C.C. (2003) Antioxidant and Free Radical Scavenging
activities of Terminalia chebula. Biological and Pharmaceutical Bulletin 26, 1331–1335.
Chowdhuri, D.K., Parmar, D., Kakkar, P., Shukla, R., Seth, P.K. and Srimal, R.C. (2002) Antistress effects of
bacosides of Bacopa monnieri: modulation of Hsp70 expression, superoxide dismutase and cytochrome
P450 activity in rat brain. Phytotherapy Research 16, 639–645.
Chu, Y.H., Chang, C.L. and Hsu, H.F. (2000) Flavonoid content of several vegetables and their antioxidant
activity. Journal of the Science of Food and Agriculture 80, 561–566.
Corvilain, B., Contempre, B., Longombe, A.O., Goyens, P., Gervy-Decoster, C., Lamy, F., Vanderpas, J.B. and
Dumont, J.E. (1993) Selenium and the thyroid: How the relationship was established. American Journal
of Clinical Nutrition 57, 244–248.
Crichton, R.R. and Charloteaux-Wauters, M. (1987) Iron transport and storage. European Journal of
Biochemistry 164, 485–506.
Cross, C.E., Halliwell, B. and Borish, E.T. (1987) Oxygen radicals and human disease. Annals of Internal
Medicine 107, 526–545.
Daun, H. (1988) The chemistry of carotenoids and their importance in food. Clinical Nutrition 7, 97.
Dhakarey, R., Uppadhyay, G., Singh, B.N., Singh, H.B., Prakash, D., Kumar, S., Singh, K.K. and Singh, R.L.
(2005) Phenolic content and antioxidant potential of Rhododendron species. Indian Society of
Agricultural Biochemists 18, 40–43.
Drew, B. and Leeuwenburgh, C. (2002) Aging and the role of reactive nitrogen species. Annals of the New York
Academy of Sciences 959, 66–81.
Prakash_Ch16.indd 261 11/27/2013 10:07:56 AM

262 R.L. Singh et al.
Exarchou, V., Nenadis, N., Tsimidou, M., Gerothanassis, I.P., Troganis, A. and Boskou, D. (2002) Antioxidant
activities and phenolic composition of extracts from Greek oregano, Greek sage and summer savory.
Journal of Agricultural and Food Chemistry 50, 5294–5299.
Fleming, R.E., Whitman, I.P. and Gitlin, J.D. (1991) Induction of ceruloplasmin in rat lung during hyperoxia.
American Journal of Physiology 260, 68–74.
Gassen, M. and Youdim, M.B. (1999) Free radical scavengers: chemical concept and chemical relevance.
Journal of Neural Transmission. Supplementa 56, 193–210.
Giovannucci, E. (1999) Tomatoes, tomato-based products, lycopene, and cancer: review of the epidemiologic
literature. Journal of the National Cancer Institute 91, 317.
Giugliano, D., Ceriello, A. and Paolisso, G. (1996) Oxidative stress and diabetic vascular complications.
Diabetes Care 19, 257–267.
Glenville, M. (2006) Nutritional supplements in pregnancy: commercial push or evidence based. Current
Opinion in Obstetrics and Gynecology 18, 642–647.
Grill, V. and Bjorklund, A. (2000) Dysfunctional insulin secretion in type 2 diabetes: role of metabolic abnor-
malities. Cellular and Molecular Life Sciences 57, 429–440.
Gupta, C., Prakash, D., Garg, A.P. and Gupta, S. (2012) Whey Proteins: A novel source of Bioceuticals.
Middle-East Journal of Scientific Research 12, 365–375.
Haider, B.A. and Bhutta, Z.A. (2006) Multiple-micronutrient supplementation for women during pregnancy.
Cochrane Database of Systematic Reviews 18, CD004905.
Halliwell, B. and Gutteridge, J.M.C. (1989) Free Radicals in Biology and Medicine, 2nd edn. Clarendon Press,
Oxford, UK.
Heidari, R.M., Mandgary, A. and Enayati, M. (2004) Antinociceptive effects and toxicity of Fumaria parviflora
Lam. in mice and rats. DARU 12, 136–140.
Holmgren, A. (1989) Thioredoxin and glutaredoxin systems. Journal of Biological Chemistry 264, 13963–13966.
Ibanez, E., Kubatova, A., Senorans, F.J., Cavero, S., Regiero, G. and Hawthorn, S.B. (2003) Subcritical water
extraction of antioxidant compounds from rosemary plants. Journal of Agricultural and Food Chemistry
571, 375–382.
Igarashi, K., Kimura, Y. and Takenaka, A. (2000) Preventive effects of dietary cabbage acylated anthocyanins
on paraquat-induced oxidative stress in rats. Bioscience Biotechnology and Biochemistry 64,
1600–1607.
Izunya, A.M., Nwaopara, A.O., Aigbiremolen, A., Odike, M.A.C., Oaikhena, G.A., Bankole, J.K. and Ogarah, P.A.
(2010) Morphological and biochemical effects of crude aqueous extract of mangifera indica l. (mango)
stem bark on the liver in wistar rats. Research Journal of Applied Sciences, Engineering and Technology 2,
460–465.
Jafri, H., Alsheikh-Ali, A.A. and Karas, R.H. (2010) Baseline and on-treatment high-density lipoprotein choles-
terol and the risk of cancer in randomized controlled trials of lipid-altering therapy. Journal of the
American College of Cardiology 55, 2846–2854.
Jaichander, P., Selvarajan, K., Garelnabi, M. and Parthasarathy, S. (2008) Induction of paraoxonase 1 and
apolipoprotein A1 gene expression by aspirin. Journal of Lipid Research 49, 2142–2148.
Kakkar, P. and Singh, B.K. (2007) Mitochondria: a hub of redox activities and cellular distress control.
Molecular and Cellular Biochemistry 305, 235–253.
Kawano, H., Motoyama, T., Hirashima, O., Hirai, N., Miyao, Y., Sakamoto, T., Kugiyama, K., Ogawa, H. and
Yasue, H. (1999) Hyperglycemia rapidly suppresses flow-mediated endothelium-dependent vasodilation
of brachial artery. Journal of the American College of Cardiology 34, 146–154.
Keaney, J.F. and Loscalzo, J. (1999) Diabetes, Oxidative Stress, and Platelet Activation. Circulation 99, 89–191.
Kemper, K.J. (2000) Garlic (Allium sativum). Available at: http://www.mcp.edu/herbal/default.htm (accessed
25 April 2013).
Kontush, A., Mann, U., Arlt, S., Ujeyl, A., Lührs, C., Müller-Thomsen, T. and Beisiegel, U. (2001) Influence of
vitamin E and C supplementation on lipoprotein oxidation in patients with Alzheimer’s disease. Free
Radical Biology and Medicine 31, 345–354.
Kubola, J. and Siriamornpun, S. (2008) Phenolic contents and antioxidant activities of bitter gourd (Momordica
charantia L.) leaf, stem and fruit fraction extracts in vitro. Food Chemistry 110, 881–890.
Kumar, M., Kumar, S. and Kaur, S. (2011) Investigations on DNA protective and antioxidant potential of chlo-
roform and ethyl acetate fractions of Koelreuteria paniculata Laxm. African Journal of Pharmacy and
Pharmacology 5, 421–427.
Kushwaha, R. and Karanjekar, S. (2011) Standardization of Ashwagandharishta formulation by TLC Method.
International Journal of Chem Tech Research 3, 1033–1036.
Prakash_Ch16.indd 262 11/27/2013 10:07:56 AM

Antioxidants: Their Health Benefits and Plant Sources 263
Latif, S., Anwar, F., Ashraf, M. and Gilani, A.H. (2007) Moringa oleifera: a food plant with multiple medicinal
uses. Phytotherapy Research 21, 17–25.
Liao, J.K., Shin, W.S., Lee, W.Y. and Clark, S. (1995) Oxidized LDL decreases the expression of eNOS. Journal
of Biological Chemistry 270, 319–324.
Losso, J.N. (2003) Targeting excessive angiogenesis with functional foods and nutraceuticals. Trends in Food
Science & Technology 14, 455–468.
Manach, C., Scalbert, A., Morand, C., Remesy, C. and Jimenez, L. (2004) Polyphenols: food sources and bio-
availability. American Journal of Clinical Nutrition 79, 727–747.
Masuda, T., Odaka, Y., Ogawa, N., Nakamoto, K. and Kuninaga, H. (2008) Identification of geranic acid, a
tyrosinase inhibitor in lemongrass Cymbopogon citratus. Journal of Agricultural and Food Chemistry 56,
597–601.
Maxwell, S.R., Thomason, H., Sandler, D., Leguen, C., Baxter, M.A., Thorpe, G.H., Jones, A.F. and Barnett, A.H.
(1997) Antioxidant status in patients with uncomplicated insulin-dependent and non-insulin-dependent
diabetes mellitus. European Society for Clinical Investigation 27, 484–490.
Milner, J.A. (1994) Reducing the Risk of Cancer. In: Goldberg, I. (ed.) Functional Foods. Chapman and Hall,
New York, pp. 39–70.
Mimica-Dukic, N., Kujundzic, S., Sokovic, M. and Couladis, M. (2003) Essential oil composition and anti-
fungal activity of Foeniculum vulgare Mill obtained by different distillation conditions. Phytotherapy
Research 17, 368–371.
Mohammad, H.M. and Elisabeth, M. (2009) Steroidal Lactones from Withania somnifera, an Ancient Plant for
Novel Medicine. Molecules 14, 2373–2393.
Moure, A., Cruz, J.M., Franco, D., Dominguez, J.M., Sineiro, J., Dominguez, H., Numez, M.J. and Parajo, J.C.
(2001) Natural antioxidants from residual sources. Food Chemistry 72, 145–171.
Nieto, F.J., Iribarren, C., Gross, M.D., Comstock, G.W. and Cutler, R.G. (2000) Uric acid and serum antioxi-
dant capacity: a reaction to atherosclerosis? Atherosclerosis 148, 131–139.
Omenn, G.S., Goodman, G., Thornquist, M., Grizzle, J., Rosenstock, L., Barnhart, S., Balmes, J., Cherniack, M.G.,
Cullen, M.R. and Glass, A. (1994) The beta-carotene and retinol efficacy trial (CARET) for chemopreven-
tion of lung cancer in high risk populations: smokers and asbestos-exposed workers. Cancer Research
54, 2038s–2043s.
Orhan, I., Sener, B., Choudhary, M.I. and Khalid, A. (2004) Acetylcholinesterase and butyrylcholinesterase
inhibitory activity of some Turkish medicinal plants. Journal of Ethnopharmacology 91, 57–60.
Ozcan, M.M. and Chalchat, J.C. (2007) Chemical composition of carrot seeds (Daucus carota L.) cultivated
in Turkey: characterization of the seed oil and essential oil. Grasasy Aceites 58, 359–365.
Pacher, P., Beckman, J.S. and Liaudet, L. (1997) Nitric oxide and peroxynitrite in health and disease.
Physiological Reviews 87, 315–424.
Panduranga Murthy, G., Mamtharani, D.R., Tejas, T.S. and Suarlikerimath, N.M. (2011) Phytochemical analysis,
in vitro anti-bacterial and antioxidant activities of wild onion sps. International Journal of Pharma and
Bio Sciences 2, 230–237.
Patel, J.M. (2008) A Review of Potential Health Benefits of Flavonoids. Lethbridge Undergraduate Research
Journal. Vol. 3, Number 2. Available at: http://www.lurj.org/article.php/vol3n2/flavonoids.xml (accessed
25 April 2013).
Piccaglia, R. and Marotti, M. (2001) Characterization of some Italian types of wild fennel (Foeniculum
vulgare Mill.). Journal of Agricultural and Food Chemistry 49, 239–244.
Ponnachan, P.T.C., Paulose, C.S. and Panikar, K.R. (1993) Effect of leaf extract of Aegle marmelose in diabetic
rats. Indian Journal of Experimental Biology 31, 345–347.
Prakash, D., Upadhyay, G., Gupta, C., Pushpangadan, P. and Singh, K.K. (2012) Antioxidant and Free Radical
scavenging activities of some promising wild edible fruits. International Food Research Journal 19,
1109–1116.
Preetha, S.P., Kanniappan, M., Selvakumar, E., Nagaraj, M. and Varalakshmi, P. (2006) Lupeol ameliorates
aflatoxin B1-induced peroxidative hepatic damage in rats. Comparative Biochemistry and Physiology
Part C: Toxicology & Pharmacology 143, 333–339.
Rauber, C.S., Guterrs, S.S. and Schapoval, E.E.S. (2005) LC determination of citral in Cymbopogon citratus
volatile oil. Journal of Pharmaceutical and Biomedical Analysis 37, 597–601.
Reaven, P. (1995) Dietary and pharmacologic regimens to reduce lipid peroxidation in noninsulin-dependent
diabetes mellitus. American Journal of Clinical Nutrition 62, 1483S–1489S.
Rice-Evans, C. and Burdon, R. (1993) Free radical-lipid interactions and their pathological consequences.
Progress in Lipid Research 32, 71–110.
Prakash_Ch16.indd 263 11/27/2013 10:07:56 AM

264 R.L. Singh et al.
Richards, R.T. and Sharma, H.M. (1991) Free radicals in health and disease. Indian Journal of Clinical Practice 2,
15–26.
Rock, C.L., Jacob, R.A. and Bowen, P.E. (1996) Update on the biological characteristics of the antioxidant
micronutrients: vitamin C, vitamin E, and the carotenoids. Journal of the American Dietetic Association
96, 693–702.
Sangwan, R.S. (2004) Photochemical variability in commercial herbal products and preparations of Withania
somnifera. Current Science 10, 461.
Sardas, S. (2003) The role of antioxidants in cancer prevention and treatment. Indoor Built Environment 12,
401–404.
Sarma, A.D., Mallick, A.R. and Ghosh, A.K. (2010) Free Radicals and Their Role in Different Clinical
Conditions: An Overview. International Journal of Pharma Sciences and Research 1, 185–192.
Sharma, H.M., Hanna, A.N., Kauffman, E.M. and Newman, H.A.I. (1992) Inhibition of human low-density
lipoprotein oxidation in vitro by Maharishi Ayurveda herbal mixtures. Pharmacology Biochemistry and
Behavior 43, 1175–1182.
Shukla, M., Singh, U., Singh, P. and Singh, R.L. (2009) Nutraceutical properties of agrowaste part of some
citrus plants. Journal of Ecophysiology and Occupational Health 9, 97–103.
Shukla, M., Singh, S.V., Singh, P., Singh, U., Vishwakerma, S.P., Khanna, A.A., Sexana, J.K. and Singh, R.L.
(2011) Anti-dyslipidimic and antioxidant activity of hydro-ethanolic fruit extract of Ficus glomerota.
Asian Journal of Pharmaceutical and Clinical Research 4, 145–148.
Singh, B.N., Singh, B.R., Singh, R.L., Prakash, D., Dhakarey, R., Uppadhyay, G. and Singh, H.B. (2009a)
Oxidative DNA damage protective activity, antioxidant and antiquorum sensing potential of Moringa
olifera. Food Chemistry and Toxicology 47, 1109-–1116.
Singh, B.N., Singh, B.R., Singh, R.L., Prakash, D., Sarma, B.K. and Singh, H.B. (2009b) Antioxidant antiquo-
rum sensing activities of green pod of Acacia nilotica L. Food Chemistry and Toxicology 47,
778–786.
Singh, B.N., Singh, B.R., Singh, R.L., Prakash, D., Singh, D.P., Sarma, B.K., Uppadhyay, G. and Singh, H.B.
(2009c) Polyphenolics from various extracts/fractions of red onion (Allium cepa) peel with potent anti-
oxidant and antimutagenic activities. Food Chemistry and Toxicology 47, 1161–1167.
Singh, P., Singh, U., Shukla, M. and Singh, R.L. (2008) Antioxidant activity imparting biomolecules in Cassia
fistula. Advances in Life Sciences 2, 23–28.
Singh, R.H. (1992) Rasayana and Vajikarana. In: Sharma, P.V. (ed.) History of Medicine in India. Indian
National Science Academy, New Delhi.
Singh, U., Singh, P., Shukla, M. and Singh, R.L. (2008) Antioxidant activity of vegetables belonging to Papilionaceae
family. Advances in Life Sciences 2, 31–36.
Sitohy, M.Z., el-Massry, R.A., el-Saadany, S.S. and Labib, S.M. (1991) Metabolic effect of licorice roots
(Glycyrrhiza glabra) on lipid distribution pattern, liver and renal functions of albino rats. Nahrung 35,
799–806.
Solomons, N.W. (2001) Vitamin A and carotenoids In: Present Knowledge in Nutrition, 8th edn. ISLI Press,
Washington, DC.
Stocker, R., Glazer, A.N. and Ames, B.N. (1987) Antioxidant activity of albumin-bound bilirubin. Proceedings
of the National Academy of Sciences of the United States of America 84, 5918–5922.
Suau, R., Cabezudo, B., Rico, R., Nájera, F. and López-Romero, J.M. (2002) Direct determination of alkaloid
contents in Fumaria species by GC–MS. Phytochemical Analysis 13, 363–367.
Thome, J., Gsell, W., Rösler, M., Kornhuber, J., Frölich, L., Hashimoto, E., Zielke, B., Wiesbeck, G.A. and
Riederer, P. (1997) Oxidative-stress associated parameters (lactoferrin, superoxide dismutases) in serum
of patients with Alzheimer’s disease. Life Science 60, 13–19.
Thomson, C.D. (2004) Assessment of requirements for selenium and adequacy of selenium status: a review.
European Journal of Clinical Nutrition 58, 391–402.
Timimi, F.K., Ting, H.H., Haley, E.A., Roddy, M.A., Ganz, P. and Creager, M.A. (1998) Vitamin C improves
endothelium-dependent vasodilation in patients with insulin-dependent diabetes mellitus. Journal of the
American College of Cardiology 31, 552–557.
Ting, H.H.,Timimi, F.K., Boles, K.S., Creager, S.H.J., Gans, P. and Creager, M.A. (1996) Vitamin C improves
endothelium-dependent vasodilation in patients with non-insulin-dependent diabetes mellitus. Journal
of Clinical Investigation 97, 22–28.
Ting, H.H., Timimi, F.K., Haley, E.A., Roddy, M.A., Ganz, P. and Creager, M.A. (1997) Vitamin C improves
endothelium-dependent vasodilation in forearm resistance vessels of humans with hypercholester-
olemia. Circulation 95, 2617–2622.
Prakash_Ch16.indd 264 11/27/2013 10:07:56 AM

Antioxidants: Their Health Benefits and Plant Sources 265
Tripathi, M., Singh, B.K., Mishra, C., Raisuddin, S. and Kakkar, P. (2010) Involvement of mitochondria mediated
pathways in hepatoprotection conferred by Fumaria parviflora Lam. extract against nimesulide induced
apoptosis in vitro. Toxicology In Vitro 24, 495–508.
Tsuda, T. (1998) Dietary cyanidin 3-0-beta-D-glucoside increases ex vivo oxidative resistance of serum in rats.
Lipids 33, 583–588.
Tsuda, T. (2000) The role of anthocyanins as an antioxidant under oxidative stress in rats. Biofactors 13,
133–139.
Ulusoy, S., Ozkan, G., Yucesan, F.B., Ersoz, S., Orem, A., Alkanat, M., Yulug, E., Kaynar, K. and Al, S. (2012)
Anti-apoptotic and antioxidant effects of grape seed proanthocyanidin extract (GSPE) in preventing
cyclosporine A-induced nephropathy. Nephrology http://dx.doi.org/10.1111/j.1440-1797.2012.01565.x.
Verma, V.K., Ramesh, V., Tewari, S., Gupta, R.K., Sinha, N. and Pandey, C.M. (2005) Role of bilirubin, vitamin c
and ceruloplasmin as antioxidants in coronary artery disease [CAD]. Indian Journal of Clinical
Biochemistry 20, 68–74.
Verschueren, K. (2001) Handbook of Environmental Data on Organic Chemicals, 4th edn, vols 1–2. John
Wiley and Sons, New York, 419 pp.
Viarengo, A., Burlando, B., Ceratto, N. and Panfoli, I. (2000) Antioxidant role of metallothioneins: a compara-
tive overview. Cellular and Molecular Biology (Noisy-le-grand) 46, 407–417.
Wang, S.Y. and Lin, H.S. (2000) Antioxidant activity in fruits and leaves of blackberry, raspberry, and strawberry
varies with cultivar and development stage. Journal of Agricultural and Food Chemistry 48, 140–146.
West, I.C. (2000) Radicals and oxidative stress in diabetes. Diabetic Medicine 17, 171–180.
Wrigstedt, P., Kylli, P., Pitkanen, L., Nousiainen, P., Tenkanen, M. and Sipila, J. (2010) Synthesis and antioxi-
dant activity of hydroxycinnamic acid xylan esters. Journal of Agricultural and Food Chemistry 58,
6937–6943.
Yanishlieva-Maslarova, N.N. and Heinonen, M. (2001) Sources of natural antioxidants. In: Pokorny, J.,
Yanishlieva, N. and Gordon, M. (eds) Antioxidants in Food. CRC Press, Boca Raton, Florida,
pp. 210–249
Yoshikawa, M., Murakami, T., Komatsu, H., Murakami, N., Yamahara, J. and Matsuda, H. (1997) Medicinal
Foodstuffs: IV. Fenugreek seeds (1): structures of trigoneosides Ia, Ib, IIb, IIIa and IIIb new furostanol
saponins from the seeds of Indian Trigonella foenum- graecum L. Chemistry and Pharmacology Bulletin
45, 81–87.
Zhang, Y.J., DeWitt, D.L., Murugesan, S. and Nair, M.G. (2004) Novel lipid-peroxidation and cyclooxygenase
inhibitory tannins from Picrorhiza kurrora seeds. Chemistry & Biodiversity 1, 426–441.
Zimmerman, M.B. and Kohrle, J. (2002) The impact of iron and selenium deficiencies on iodine and thyroid
metabolism: biochemistry and relevance to public health. Thyroid 12, 867–878.
Prakash_Ch16.indd 265 11/27/2013 10:07:56 AM