.

1. Definition
2. Characteristics
3. Formation
4. Structure
5. Stability
6. Detection
7. Chemical Reactions
8. Pharmaceutical Importance
9. Antioxidants
Chemical species (atoms or molecules) that contain an
unpaired electron are called Free Radicals.

Chlorine free radical Cl

.

Hydrogen free radicals H

.
Bromine free radicals Br
.
Oxygen free radical O
.
 Free radicals are neutral species.
 Free radicals are named after the atom or molecule
from which they are formed
 Free radicals are represented by the symbol or
molecular formula and a dot on upper right on it.
 They have a very short half life.
 They are highly reactive and react indiscriminately with
the molecules in their immediate vicinity.
 Free radicals react with other free radicals or molecules to
form new free radicals or stable molecules.
 Free radicals may be colored even though their precursor
molecules may be colorless.
 They are formed by the homolytic fission of bond
 They are produced in both biological and chemical
reaction.
Homolytic bond fission:
Free Radicals
Heterolytic bond fission:
Cation and anion
.
Conjugation:
Free radicals of conjugated compounds are more stable
than non conjugated compounds because of delocalization
of unpaired electrons with the π electrons.
Steric effect:
Steric effect make the free radicals more stable by making
the attacking reagent difficult to approach the free radical.
Characteristics of Free Radical Reactions:
 Free radical reactions mostly take place in gaseous
phase, however these also occur in non polar solvents
and in presence of light as catalyst.
 They are less energy demanding
 Free radical reactions once initiated proceed with
great rapidity owing to their chain reactions of low
energy requirements
 Freeradical reactions can be inhibited by substances
which react with free radicals such as phenols, quinones,
diphenylamine, iodine etc. Such substances are called
Radical scavengers or inhibitors
Most free radical reactions proceed as chain reactions
and have three distinct phases viz.
 Initiation:
Generates a radical.
 Propagation:
The radical reacts with a stable molecule to produce
another reactive free radical and continues the chain
reaction.
 Termination:
Side reactions that destroy the reactive intermediate.
 Requires heat or light for initiation.
 The most effective wavelength is blue, which is absorbed
by chlorine gas.
A chlorine molecule splits homolytically into chlorine free
radicals.
The chlorine atom collides with a methane molecule
and abstracts a H, forming another free radical and one
of the products (HCl).
The methyl free radical collides with another chlorine
molecule, producing the organic product (methyl
chloride) and regenerating the chlorine radical.

.
A reaction is classified as a termination step when
any two free radicals join together producing a
non radical compound.
.
1. Heart diseases (32)* 6. Pneumonia & Influenza (4)
2. Cancers (23)* 7. Diabetes Mellitus (3)*
3. Strokes (7)* 8. AIDS (2)
4. Lung diseases (5) 9. Liver Disease (1)
5. Accidents (4) 10. Suicide (1)

 It indicates diet-related diseases (65%)

 >90% of disease incidence involves free radical damage

 Generation of Free Radical

1. Environmental effects:
a) Industrial and environmental pollution
b) Agricultural practices including use of herbicides and pesticides
c) Due to damages caused by UV or X-rays
d) Cigarette or alcohol.
e) Due to drug metabolism.
Industrial pollution

Pesticides &
herbicides

Excessive
Alcohol &
smoking

High fat foods

 Generation of Free Radical
2. Cellular metabolism:
About 1-5% of oxygen taken up in the body is converted to free
radicals.
They are constantly produced in mitochndria during the normal
oxidation of foodstuffs.
a) Due to leaks in the electron transport chain in mitochondria.
b) Some enzymes such as xanthine oxidase and aldehyde oxidase
form superoxide anion radical or hydrogen peroxide.
c) Macrophage also produces NO . from arginine by the enzyme
nitric oxide synthase. This is also an important anti-bacterial
mechanism.
 Types of Free Radicals Affecting
Biological System
Most free radicals in biological systems are derivatives of
oxygen (Reactive Oxygen Species, ROS),
and
Derivatives of nitrogen (Reactive Nitrogen Species, RNS).
Reactive Oxygen Species is used in a broad sense to
collectively free radicals (O2.-, OH.) and non-free
radicals (H2O2, 1O2, which are extremely reactive) of
the biological system.
1. Superoxide, O2.
2. Hydrogen peroxide, H2O2
3. Hydroxyl radical, OH.
4. Singlet oxygen, 1O2
5. Hydro peroxy radical, HOO.
6. Lipid peroxide radical, ROO.
7. Nitric oxide, NO.
8. peroxynitrite, ONOO.
When excess amounts of ROS is produced in cells e.g in
disease state or exercise the condition is called oxidative
stress.
These free radicals react indiscriminately with
biomolecules and cause damage to
 Proteins
 Lipids
 Carbohydrates
 DNA
 Harmful effects of free radicals
A. Free Radical and biomolecules
1. Proteins
Cause oxidation of sulfhydryl groups, and modification of
amino acids. ROS may damage protein by fragmentation,
aggregation results in the loss of biological activity of proteins.
2. Lipids
The polyunsaturated lipids of cell membranes are particularly
susceptible to damaging effects of free radicals.
This leads to the uncontrolled chain reaction called lipid
peroxidation.
 Lipid peroxidation refers to the
oxidative degradation of lipids.
 It is the process whereby free
radicals "steal" electrons from the
lipids in cell membranes, resulting
in cell damage.
 This process proceeds by a free
radical chain reaction mechanism.
 It most often affects
polyunsaturated fatty acids(PUFA).
 In addition, end products of
lipid peroxidation may be
mutagenic and carcinogenic
 Harmful effects of free radicals

3. Carbohydrates
Glycation increases the susceptibility of proteins to the
attack by free radicals.
4. Nucleic acid
Free radicals cause DNA strand breaks, fragmentation of bases
and deoxyribose results in cytotoxicity and mutations.
Sources Of Free Radical and damage to
biomolecules
 Harmful effects of free radicals
B. Diseases
1. Cardiovascular diseases (CHD): ox-LDL, formed by the action of
free radicals, promote CHD and atherosclerosis.
2. Cancers: damage DNA and cause mutation and cytotoxicity, play a
key role in carcinogenesis.
3. Inflammatory diseases: damage on the extracellular components
such as collagen and hyaluronic acid, promote glomerulonephritis
and ulcerative colitis.
4. Respiratory diseases: destroy endothelium and cause lung edema.
Cigarette smoke contains free radicals and promotes the production of
more free radicals.
 Harmful effects of free radicals
B. Free Radical and diseases
5. Diabetes mellitus: Destruction of islets results in pathogenesis.
6. Cataract
7. Male infertility: reduce sperm motility and viability.
8. Aging process
9. Others: such as Parkinson’s disease, Alzheimer’s disease,
multiple sclerosis, liver cirrhosis, muscular dystrophy.
 Severity of Biological
Oxidative Stress Consequences

A. Low level & gradual Aging

Carcinogenesis
B. Medium level & rapid Mutagenesis

C. Large level & rapid Stroke, Trauma,

,Death,
 Weapons of immune system providing anti bacterial
action
 Cell signaling e.g. isosorbide mononitrate
 Activation of gene-transcription factors
 Modulate metabolism
Free Radicals and Drug Stability:
Stability of drug is defined as;
“The state of drug formulation when it remains within
physical, chemical, microbiological and therapeutic
specifications set at the time of manufacture is called drug
stability.”
1. Hydrolysis
2. Oxidation Auto-oxidation
3. Isomerization
4. Polymerization
5. Decarboxylation
 The slow oxidation of a compound under the
influence of atmospheric oxygen is called auto oxidation.
 Fixed oil, volatile oils, fats, fat soluble and water-soluble
vitamins and phenols undergo auto-oxidation
 Auto-oxidation is a chain reaction involving formation of
free radicals.
 Antioxidants
 The substance present in low concentration relative to the
oxidizable substrate that significantly delays or reduces oxidation
of the substrate.

 They reduce the effect of dangerous oxidants (free radicals) by

binding together with these harmful molecules, decreasing their
destructive power.

 They can also help repair damage already sustained by cells.

 They may be considered as the scavengers of free radicals.

 Prevents the transfer of electron from organic molecules to free
radicals.
 During this reaction the antioxidant sacrifices itself by donating
electron to free radical and thus is oxidized. Thus it prevents the
oxidation of organic molecules so it is called as antioxidant.
 Stabilizes free radicals.
 Terminates free radical reactions.
 As one antioxidant molecule can only react with a single free
radical. Therefore, there is a constant need to replenish antioxidant
resources.
Qualities of a good antioxidants:
 Decays to harmless products

 Relatively un-reactive

 Repaired Rapidly
 Classification of antioxidant
Ⅱ. According to their chemical
nature and action: Metabolic antioxidants:
a)Enzymatic antioxidants:  Bilirubin
 SOD  Uric acid
 Catalase  Transferrin
 Glutathione reductase  Albumin
b)Non-enzymatic antioxidants: Chemicals:
Nutrient antioxidants:  Sodium bisulphite,
 β-carotene (Vitamin A)  Butylated hodroxy
 α-tocopherol (Vitamin E) toluene (BHT)
 Ascorbic acid (Vitamin C)
 Classification Of Antioxidant
Ⅰ. According to their location:
a) Plasma antioxidants: b) Cell membrane antioxidants:
 Ascorbic acid (Vitamin C)  α- tocopherol (Vitamin E)
 Bilirubin c) Intracellular antioxidants:
 Uric acid  Superoxide dismutase
 Transferrin (SOD)
 Ceruloplasmin  Catalase
 β-carotene  Glutathione peroxidase
(GPx)
 Enzymatic Antioxidants
1. Superoxide dismutase (SOD)

2O2·⁻+ 2H+ SOD H2O2 + O2

SOD is present in essentially every cell in the body .

This is the first line of defence to protect cells from the injurious

effects of superoxide.
 Enzymatic Antioxidants
2. Catalase, CAT

2H2O2 catalase 2H2O + O2

Catalase is present in all body organs being especially

concentrated in the liver and erythrocytes.
The brain, heart and skeletal muscle contains low amounts.
 Nutrient Antioxidants
1. alpha-tocopherol (vitamin E)

 The most important lipid-soluble antioxidant.

 Present in all cellular membranes.
 Vitamin E prevents the peroxidation of membrane phospholipids
and prevents cell membrane damage through its antioxidant action.
 Nutrient Antioxidants
2. Ascorbic acid (vitamin C)

+ 2O2·⁻ + 2H+ H2O2 +

Dehydroascorbate, DHA
It is a water-soluble, antioxidant present in citrus fruits, potatoes,
tomatoes and green leafy vegetables.
It is a chain breaking antioxidant as a reducing agent or electron
donor. It scavenges free radicals and inhibits lipid per oxidation. It
also promotes the regeneration of α-tocopherol.
 Nutrient Antioxidants
3. Carotenoids Vitamin A:

Carotenoids consist of C-40 chains with conjugated double bonds,

they show strong light absorption and often are brightly colored
(orange).
They occur as pigments in bacteria, algae and higher plants.
β-carotene is the most important.
It is composed of two molecules of vitamin A (retinol) joined together.
It can quench singlet oxygen
 Other Important Nutrient Antioxidants
Antioxidant Source

1.Coenzyme Q10 Organ meats (best heart), beef, chicken

2. Selenium Sea foods, meats, whole grains
3. Catechins Green tea
4. Quercetin Onions, red wine, green tea
 Metabolic Antioxidant
1. Glutathione, GSH

GSH is also a scavenger of hydroxyl radicals and singlet oxygen.

 Metabolic antioxidant

1. Uric acid: Scavenge singlet oxygen and hydroxyl radical

2. Ceruloplasmin: Inhibit iron and copper dependent lipid per oxidation
3. Transferrin: Prevents iron-catalyzed radical formation
4. Albumin: Scavenge radicals on its surface
5. Bilirubin : Protects albumin bound free fatty Acid from peroxidation
Normally, cellular homeostasis is a delicate balance between the
production of free radicals and our antioxidant defenses.