Oxidants and Anti-Oxidant Systems

Overview

Cells continuously generate reactive oxygen species (ROS) and reactive nitrogen-oxygen species (RNOS) as part of normal metabolism and immune defense. These molecules play dual roles in physiology (signaling, immune defense) and pathology (oxidative damage). Survival depends on tightly regulated antioxidant control systems.

• ROS/RNOS: Generated during cellular metabolism and immune responses.

• Physiologic roles: Signaling, immune weapons.

• Pathologic roles: Oxidative damage to biomolecules.

• Antioxidant systems: Essential for cellular protection.

Radicals, Anions, and Oxygen Chemistry

Definitions and Properties

Understanding the chemistry of radicals and anions is fundamental to biochemistry and cell biology.

• Radical (free radical): Molecule with at least one unpaired electron.

• Anion: Molecule with a net negative charge; some molecules can be both (e.g., superoxide).

• Biradical nature of O2: Molecular oxygen has two unpaired electrons, making it relatively stable but prone to stepwise one-electron reductions.

Reactive Oxygen Species (ROS)

Definition and Behavior

ROS are oxygen-containing molecules with high chemical reactivity, including both radicals and non-radicals.

• Highly reactive radicals: Extremely short half-life, react near site of formation, cause chain reactions (especially in lipids).

Sources of Superoxide

Major Cellular Sources

Superoxide is generated by several cellular processes:

• Mitochondrial Electron Transport Chain (ETC): Electron leak at Complex I & III.

• NADPH oxidase: In phagocytes and endothelium.

• Xanthine oxidase

• Cytochrome P450 reactions

Reaction Example:

Sources of Hydrogen Peroxide

Generation and Features

• Superoxide dismutation: Catalyzed by superoxide dismutase (SOD).

• Oxidase enzymes: e.g., amino acid oxidases.

• Peroxisomal reactions

• Key features: Not a radical, membrane-permeable, precursor to hydroxyl radical.

Reaction:

Sources of Hydroxyl Radical

Metal-Catalyzed Reactions

• Fenton reaction:

• Haber-Weiss reaction:

• Clinical tie-in: Iron overload increases •OH formation.

Reactive Nitrogen-Oxygen Species (RNOS)

Key Molecules and Synthesis

• Nitric oxide (NO•)

• Peroxynitrite (ONOO−)

• Nitric oxide synthase (NOS) reaction:

• Required cofactors: NADPH, FAD, FMN, tetrahydrobiopterin (BH4), heme, O2

Peroxynitrite Formation

Reaction and Consequences

• Reaction:

• Consequences: Potent oxidant, causes protein nitration, damages mitochondria and DNA, functions as immune weapon.

Respiratory Burst

Enzymatic Reactions in Immune Cells

• NADPH oxidase:

• Myeloperoxidase (MPO): Uses to produce hypochlorous acid (HOCl).

• Key formulas: Chloride: , Hypochlorous acid: , Hypochlorite:

• Function: Microbial killing.

Oxidative Damage to Cells

Targets and Biomarkers

• Lipids: Peroxidation of polyunsaturated fatty acids (PUFA).

• Proteins: Oxidation, nitration.

• DNA: Strand breaks, base modification.

• Lipid peroxidation: Chain reaction leads to membrane rigidity and leakage.

• Malondialdehyde (MDA): Biomarker for lipid peroxidation.

Antioxidant Enzymes Overview

Superoxide Dismutase (SOD) and Catalase

• Superoxide Dismutase (SOD): Isoenzymes include Cu/Zn-SOD (cytosol), Mn-SOD (mitochondria), EC-SOD (extracellular).

• Catalase: Located in peroxisomes; high-capacity enzyme for H2O2 loads.

• Cofactor: Heme (iron).

Glutathione System

Enzymes and Cofactors

• Glutathione peroxidase (GPx):

• Reduced glutathione (GSH): γ-glutamyl-cysteinyl-glycine.

• Selenium (cofactor): Incorporated as selenocysteine in GPx active site.

• Glutathione reductase: (Cofactor: FAD)

Non-Enzymatic Antioxidants

Dietary and Endogenous Molecules

• Dietary: Vitamin E (α-tocopherol), Vitamin C, Carotenoids, Flavonoids.

• Endogenous: Uric acid, Melatonin.

• Role: Free radical scavenging, chain-breaking antioxidants.

Vitamin E (α-tocopherol)

Structure and Function

• Most biologically active form: α-tocopherol.

• Fat-soluble, lipid-phase antioxidant: Localizes to cell membranes and lipoproteins.

• Primary function: Chain-breaking antioxidant, protects PUFAs in membranes.

• Key reaction: Lipid radical (L•) + α-tocopherol → stable lipid + tocopheroxyl radical.

• Stops propagation phase of lipid peroxidation.

Vitamin C

Structure and Function

• Ascorbic acid: Water-soluble antioxidant.

• Primary function: Electron donor (reducing agent), directly scavenges superoxide, hydroxyl radical, peroxyl radicals.

• Regenerates oxidized antioxidants: Vitamin E, Glutathione.

• Enhances non-heme iron absorption:

Carotenoids

Structure and Function

• Lipid-soluble plant pigments: Found in fruits and vegetables.

• Major examples: β-carotene (provitamin A), lycopene, lutein, zeaxanthin.

• Primary function: Quench singlet oxygen (), scavenge peroxyl radicals (ROO•), act in lipid environments (cell membranes, LDL).

• Key mechanism: Physical quenching (energy transfer, not consumed), stabilize excited oxygen species.

Flavonoids

Structure and Function

• Large family of plant polyphenols: Characterized by multiple phenolic –OH groups.

• Major subclasses: Flavonols (quercetin), Flavones, Flavanols (catechins), Anthocyanins, Isoflavones.

Uric Acid

Structure and Function

• End product of purine metabolism: Formed from xanthine via xanthine oxidase.

• Major aqueous-phase antioxidant in plasma: Scavenges hydroxyl radical, peroxynitrite, singlet oxygen.

• Contributes up to ~50% of total plasma antioxidant capacity.

Melatonin

Unique Antioxidant Features

• Antioxidant cascade: Melatonin metabolites are also antioxidants.

• Does not become pro-oxidant.

• Crosses: Cell membranes, blood-brain barrier, mitochondrial membranes.

• Mitochondrial protection: Concentrates in mitochondria, reduces electron leak from ETC, ROS generation at source, preserves cardiolipin, mitochondrial DNA, ATP production.