Free radicals are unstable molecules with unpaired electrons that can cause oxidative damage. They are produced during normal metabolism but also due to environmental toxins and radiation. Antioxidants help stabilize free radicals and prevent cellular damage. The body has endogenous antioxidants like glutathione, vitamin C and E, and antioxidant enzymes. Exogenous antioxidants from foods and supplements can further support the antioxidant defense system. Oxidative stress occurs when there is an imbalance between free radicals and antioxidants leading to diseases. Maintaining adequate antioxidant levels through diet and lifestyle can help reduce oxidative damage and disease risk.

1. Dr Aswathi T

2. • Oxidation – removal of electrons
• Reduction – gain of electrons
Atomic stability of any orbital
• Atleast 2 electrons
• Spinning in opposite directions
• Most biological molecules
• Non-radicals (non-free radicals)

4. Free radicals
• Unpaired electron
• Unstable
• Highly reactive – oxidize biomolecules
• Formed constantly in human system
Accidental products
during metabolism
Deliberately during the
process of phagocytosis
Tissue injury
and
cell death

5. Sources of free radicals in the body
• Cyclooxygenation
• Lipooxygenation
• Lipid peroxidation (auto-oxidation on exposure to oxygen)
• Neutrophils (stimulated by exposure to microbes)
• Re-perfusion of ischemic organs

6. Other sources
• Toxic environmental pollutants (motor car exhaust)
• UV & Ionizing radiations
• Ozone
• Heavy metal poisoning
• Cigarette smoking
• Chronic alcohol intake

7. Types of Free Radicals
1. Superoxide Radicals (O₂●)
• Superoxide ion
• Oxygen molecule deficient of one electron
• Derived from
i) Inevitable leakage from the mitochondrial electron transport chain
reactions
ii) Metabolism of various drugs by cytochrome P-450 microsomal
oxidative system (eg: paracetamol or alcohol)
iii) Action of some enzymes

8. • Hypoxanthine Xanthine Uric acid
Oxidation
Superoxide ion Hydrogen peroxide

9. 2. Other Free Radicals
• Hydroxyl (OH●)
• Peroxyl (ROO●)
• Nitric oxide free radical (NO●)
Oxidative destruction of PUFAs
of cell membrane
(Lipid peroxidation)

10. • Directly disrupts the integrity and function of biological cell
membranes
• Destabilizes membrane receptors
• Damage to cellular components ( due to reactive aldehyde
production)
• Attack on proteins – inactivates enzymes
• Damage to nucleic acid – breaks in DNA strands
Free radicals are Carcinogenic

11. • Nitric oxide (NO) – cytotoxicity mechanism
• Very high affinity to even low concentrations of superoxide ion (O₂●)
• In oxidative stress – plenty of superoxide free radical – NO reacts very
rapidly
• Unstable peroxynitrite (ONOO●)
• Instantly gets degraded to hydroxyl (OH●) & nitrite free radical (NO₂●)

12. Nitric oxide (NO ) + Superoxide (O₂●)
Peroxynitrite (ONOO● - unstable)
Hydroxyl (OH●) + Nitrite (NO₂●)
Cytotoxicity of NO
During oxidative stress

13. How they are stabilized?
• Donate its unpaired electron
• Accept one from other biomolecules
• Non-free radical free radical
Disastrous chain reaction

14. But!
• If 2 unpaired electrons are shared by each other
• Covalent bond formation
• Reactivity terminated
X● + X● X-X

15. Reactive Oxygen Species (ROS)
• Highly reactive species – not free radicals in true sense
• Free radicals
positively charged
negatively charged
electrically neutral
Inorganic species (H₂O₂, HOCl, O●)

16. • H₂O₂ & HOCl – not free radicals - very powerful oxidizing agents
• Nascent oxygen – transient atomic state of oxygen – highly reactive
than O₂ - causation of oxidative processes & photosensitization
reactions
Symbol Name
¹O₂
R + O⁻
LOOH
Singlet oxygen
Alkoxy free radical
Lipid peroxide

17. Promoters of Free Radicals
• Transition metals – iron, copper
• Variable oxidation number (valency)
• Can accept a single electron or donate it
• Fe³⁺ + e⁻ = Fe²⁺
• Cu²⁺ + e⁻ = Cu⁺
• H₂O₂ OH● (Hydroxyl radical)
Promoters of free radicals
Iron & copper ions
Accelerate lipid
peroxidation
Accumulation of
highly cytotoxic
end products

18. How free radicals are removed?
Enzymatic and non-enzymatic antioxidant defense mechanisms
If it is inadequate
Free radical scavenging
Oxidative stress
an imbalance
between free
radicals and
antioxidants in
body

19. Lipid peroxidation
Protein and DNA damage
Tissue injury & cell death

20. Free Radicals and Diseases
• Autoimmune diseases
• Rheumatoid arthritis
• Paracetamol and alcohol toxicity
• Radiation injury
• Carcinogenesis
• Amyloidosis
• Aging

21. System / organ Disease
1. CNS Parkinsonism, Alzheimer’s disease, Huntington’s disease, multiple
sclerosis
2. CVS MI, Ischemic perfusion injury, Atherosclerosis
3. Endocrine Diabetes mellitus
4. GIT Peptic ulcer, Cirrhosis, Pancreatitis
5. Renal Nephrotoxicity (due to aminoglycosides & heavy metal poisoning)
6. RS Toxicity due to cigarette smoking
7. Eyes Cataract, Retinopathy

23. Antioxidants and their Clinical Use
• Free radical generation causation of tissue injury
i) Blockade of generation of toxic free radical
ii) Blockade of the chain reaction set by free radical
iii) Scavenging of free radical
iv) Blocking the secondary generation of toxic metabolite or mediator
V) Enhancing antioxidant capability

24. • At low concentration – prevent or delay the oxidation of lipids,
carbohydrates, proteins and nucleic acids
• Improve the quality of life by retarding the ageing process

25. Body defenses against oxidants
Intracellular enzymes
• Catalases, superoxide dismutases, glutathione peroxidases
• Dispose of the oxidants
Antioxidants obtained along with nutrients
• Vitamins A, C, E
• Trace elements – selenium
• Flavonoids and polyphenolic substances present in plant foods (fruits,
vegetables, amla, tea, garlic and others)
• Others present in spices (coriander, clove, cardamom)

26. Endogenous Antioxidants Exogenous (Pharmacological) Antioxidants
Superoxide Dismutase (SOD)
Glutathione Reduced form (G-SH)
Carotenoids (beta-carotene)
Melatonin
Alfa-Tocopherol (Vit. E)
Ascorbic acid (Vit. C)
Miscellaneous
Adenosine
Lactoferrin
Transferrin
Nicotinamide
Agents augmenting endogenous antioxidants
N-acetylcysteine
Ebselen
DesferoxamineCeruloplasmin
Allopurinol
Oxypurinol
Selegiline
Trace Elements and Minerals
Selenium
Manganese
Zinc
Copper
Chromium
From plant sources
Garlic, grape, soyabean (isoflavones), turmeric
(curcuminoids), tomato (lycopene, a red
carotenoid pigment), herbal preparations
containing bioflavonoids (phenolic chain
breaking anto-oxidant), spirulina

27. 1. Superoxide Dismutase (SOD)
• Found in all cells
• Scavenges superoxide ion - Prevents its accumulation
• Cells are protected from oxidative stress
• Catalyses dismutation reaction
• O₂● + O₂● + 2H H₂O₂ + O₂
• 2H₂O₂ 2 H₂O + O₂
H₂O₂ can still oxidise and
damage many cellular
components
Removed by catalases and
glutathione peroxidase

28. SOD afford protection against
• Ischemic reperfusion injury
• Pancreatitis
• Gastrointestinal ischemia
• Myocardial infarction
• Endotoxic shock
• Organ transplant

29. 2. Glutathione Reduced form (G-SH)
• Excellent antioxidant & cytoprotective
• Prevents free radical induced oxidation of SH groups of various
proteins to disulfide (-S-S-) derivatives
• Also protects hemoglobin from getting oxidized by H₂O₂
• Glutathione peroxidase (Selenium containing enzyme) scavenges toxic
amounts of peroxides, H₂O₂ and free radicals
• NADPH regenerates G-SH (reduced form) from G-S-S-G (oxidized
form) by glutathione reductase

31. G-SH improves the outcome of
• Organ transplant
• Cystic fibrosis
• Reperfusion injury
• Diseases in AIDS patients

32. 3. Carotenoids (Beta-Carotene)
• Vitamin-A precursor
• Reduces the risk of cancers due to free radical damage
• Provide protection from heart attack and photo-sensitisation

33. 4. Melatonin
• Hormone from pineal gland
• Potent scavenger of OH● radicals
• Neutralizes nascent oxygen
• Removes H₂O₂ and NO
• Inactivates peroxynitrite (ONOO●)
• Catalyses action of various enzymes involved in the activity of
endogenous antioxidants

34. 5. Alfa-Tocopherol (Vit. E)
Protection against
• Neurological damage
• Atherosclerosis
• Thrombotic vascular disease
• Retrolental fibroplasia

35. 6. Ascorbic Acid (Vit. C)
• Reacts with H₂O₂ , peroxide and superoxide free radicals
• Itself gets oxidized to Dehydroascorbate
Ascorbic Acid
Deficiency of Vit. C predisposes to
Atherosclerosis
Carcinogenicity

36. Tobacco (chewing and smoking) – rich in free radicals
Damage DNA
Throat and lung cancer
Vit. C and E inhibits Nitrosamine formation

37. 7. Miscellaneous
i) Adenosine
• Endogenous anti-inflammatory-antioxidant
• Inhibits superoxide generation by NADPH oxidase in neutrophils and
macrophages
• Affords protection against reperfusion injury
ii) Lactoferrin and Transferrin
• Circulating iron binding proteins
• Reduce iron-dependent free radical toxicity
• Afford natural protection against lung cancer
iii) Nicotinamide

38. Agents Augmenting Endogenous Antioxidants
i) N-acetylcysteine – glutathione precursor
ii) Ebselen – congener of glutathione peroxidase
• N-acetylcysteine – treating paracetamol toxicity & in AIDS patients
iii) Desferoxamine
Iv) Ceruloplasmin
. Augment endogenous
glutathione peroxidase activity
. Promote endogenous
antioxidant activity
Bind free Fe²⁺
Detoxify superoxide anion
Inhibit iron-dependent lipid peroxidation

39. Exogenous (Pharmacological) Antioxidants
i) Allopurinol
ii) Oxypurinol
• Reperfusion injury
• Myocardial infarction
• Cerebral and gastrointestinal reoxygenation injury
iii) Selegiline
Antigout drugs
Xanthine oxidase inhibitors
Inhibiting the process of superoxidation
Xanthine
oxidase
promotes
superoxidation

40. Trace Elements and Minerals
• Selenium
• Enhance antioxidant activity of vitamin E
• Promotes synthesis of glutathione peroxidase
• Stimulate the immune system of the body
• Most favoured ingredient in various antioxidant formulations
• Manganese, Zinc, Copper and Chromium
• Good sources for generating SOD, glutathione peroxidase and
catalase enzymes

41. Antioxidants From Plant Sources
• Garlic, grape fruit juice, soyabean (isoflavones), turmeric
(curcuminoids), tomato (lycopene, a red carotenoid pigment), herbal
preparations containing bioflavonoids (phenolic chain breaking anto-
oxidant), spirulina

42. Coenzyme Q10 (Ubiquinone)
• Fat-soluble antioxidant
• Humans synthesize as well as obtained it from dietary sources
• Present in mitochondria of human cells
• High concentrations found in heart, liver, kidney and skeletal
muscle
• Tissue concentration decline with age
• Sold as a dietary supplement
• Involved in ATP generation

43. • Usual dose: 100-200 mg/day in divided doses
• Beneficial effects in
- Parkinson's disease
- CHF
• Well tolerated
• Adverse reactions – mild – nausea, vomiting & abdominal discomfort
• Decrease response to warfarin

44. Spirulina
• Marine blue-green algae
• “Most powerful food on earth”
• Good source of SOD, beta-carotene and
B-complex vitamins
• Contain 70% by weight of protein and many
micronutrients
• 2 tablets per day (each containing 500 mg
of spirulina)

45. Summary…..

46. THANK YOU