OXYGEN METABOLISM AND TOXICITY

Oxygen
• Molecular O2 – two unpaired electrons with parallel spins (antibonding character) Free Radicals (FR)
• To accept e- one electron (either O2 or substrate) must spin invert • Chemical species with unpaired e- in valence shell
• Spin inversion – large thermodynamic / kinetic barrier, multistep, ↑ Ea; protective • ↓ chemical specificity = abstracts e- indiscriminately
• One e- reductions of O2 are not subject to kinetic barrier • Generation
o Homolytic covalent bond cleavage: A:B  A· + B·
• Oxidative Stress – state wherein level of ROS > endogenous antioxidant defenses 
↑ FR  local injury (lipids most prone) organ dysfunction
o Single e- loss: X  e- + X+·
o e- addition: Y  e- + Y-·
Reactive Oxygen Species Properties
• major O2 metabolites by 1e- reduction
• Cytotoxic oxidative chain reaction: indiscriminately extracts e- from molecules forming new FRs
ROS Formation Notes
O2- Superoxide ETC, leakage @ CoQ, accidental nonspecific CoQH + O2 central role in formation of other reactive intermediates
Hydrogen 2-electron reduction of O2 not a FR, but an oxidizing agent, main source of OH· in presence of transition metals,
H2O2
Peroxide Dismutation Rxn: 2O2- + 2H+  H2O2 + O2 via SOD Dismutation: radical  nonradical
Fenton Rxn: Fe2+ + H2O2  Fe3+ + OH- + OH·; probable initiator of chain reactions, forms lipid peroxides, most reactive against biological
OH· Hydroxyl radical molecules (membrane lipids)
Haber-Weiss Rxn: O2- + H2O2  O2 + H2O + OH·
1O2 Singlet Oxygen Pigment rxn non radical, strong oxidant, electronically excited, mutagenic form of O2: e- paired in π orbital
NO Nitric oxide arginine  citrulline + NO via NOS endothelium derived relaxing factor, gaseous FR
ONOO- Peroxynitrite NO + O2-
HOCl Hypochlorous acid H2O2 + Cl-  HOCL + OH- via myeloperoxidase major bactericidal agent in activated polys

Source Mechanism Examples

reduction of O2 diradical, abstraction from Fe2+ by O2 catecholamines, hemoglobin, myoglobin, reduced CytC, thiol
Autoxidation
Oxidation xanthine oxidase (XO) (ischemia-reperfusion), prostaglandin synthase, lipoxygenase;
Enzymatic oxidation
Endogenous

H2O2 + Cl-  HOCl myeloperoxidase (neutrophils)

↑ O2 in phagocytes, flavoprotein Cyt-b-245 NADPH oxidase + Chronic granulomatous disease (CGD) – defective NADPH-oxidase system  prone to
Respiratory burst
immune response  O2-  dismutate to H2O2  OH· + HOCl bacterial infections by catalase producers
Subcellular Organelles ETC leaks  ↑ O2- during ↑ O2 or ischemia Mitochondria, microsomes & peroxisomes (H2O2)
Transition metal ions Fenton reaction Fe, Cu, and other e- donors
Ischemia reperfusion Ischemia  ↑ xanthine & XO, ↓ SOD & GSHP Xanthine oxidase
injury Reperfusion  ↑ O2 = e- acceptor & XO cofactor  O2- and H2O2
 ↑ FR during hyperoxia Antibiotics w/ quinoid or metals (nitrofurantoin), antineoplastics (bleomycin),
Drugs
anthracycline (andriamycin), pro-oxidants (methotrexate)
Transfers energy to cellular components = 1° radicals  2° rxn Electromagnetic (X-rays, γ-rays) or particulate radiation (e-, photons, neutrons, α & β
Radiation
particles)
Exogenous

Alveolar damage, FRs in gas phase, semiquinones in tar phase Contains aldehydes, epoxides, peroxides
Tobacco smoking
Lung microhemorrhages  Fe  Fenton ↑ neutrophils = ↑ FR NO, peroxyl radicals, carbon-centered radical
Phagocytosis  cell rupture  proteolytic enzymes  ↑ Mineral dust, silica, asbestos
Inorganic particles
neutrophil  ↑ FR
Degradation  hydroxyl radicals O3 Ozone – not FR, oxidizing agent, 2 unpaired e-
Gases CFCs
Photodissociation  chlorine radicals

Defense Mechanisms
O2-  H2O2 + O2, requires Cu, Zn, Mn
Superoxide Dismutase (SOD)
1° defense, CuZnSOD in plasma and cytosol, MnSOD in mitochondria
Enzymes

H2O2 + H2O2  H2O + O2

Catalase in peroxisomes > cytosol and microsome, Fe @ active site,
↑ in liver and RBC, ↓ in brain heart, and skel muscle
Glutathione peroxidase (GSHP) H2O2 and lipid peroxides + Glutathione (GSH)  H2O + GSSG, requires selenium
Low MW scavengers / Vitamins Metal chelators

Ferritin Multi-protein shell surrounding FE3+ core, prevents Fenton

Transferrin Binds Fe3+
Ceruloplasmin Converts Fe2+ to Fe3+
Albumin Binds to Cu2+ > Fe
Urate Chelates Fe and Cu, from purine catabolism

Vitamin E (α tocopherol)
Vitamin C
Carotenoids / VitA / β-carotene
Major membrane bound chain-breaking antioxidant, fat soluble, reacts with peroxyl and other radicals
Terminates FR lipid peroxidation by e- donation  tocopheryl radical  stable, nonfunctional, oxidized tocopheryl quinone + VitC  reduced
Water soluble, most effective aqueous phase antioxidant
Accepts e- from O2-, H2O2, HOCl, OH·, OOH radicals
L-ascorbate  e-, H+  ascorbyl radical  -e  dehydro-L-ascorbic acid
Chain-breaking antioxidant, fat-soluble, from carrots, tomato (lycopene)
Accepts e- from lipid peroxy radicals  free radical intermediate
Maintains reduced cysteinyl in Hb, prevents methmyoglobin formation

Glutathione

Cellular compartmentation Separation of species and sites involved in ROS formation, ferritin, peroxisomes
Repair mechanisms DNA repair, oxidized FA removal from membrane, oxidized protein degradation and resynthesis

Cellular Damage Causes Targets Effects

OH· + LH  LH· 
Lipid polyunsaturated lipid propagated by O2  LOO· (lipid peroxy radical) + LOOH (lipid peroxide) 
initiated by FR (i.e. OH·)  chain reaction
peroxidation (LH) degraded LOOH + malondialdehyde (blood soluble)
Termination: antioxidant e- donation in 2 steps = stabilized oxidized compound
Protein damage OH· attack  oxidation Pro, His, Arg, Cys, Met protein fragmentation, cross-linking, aggregation, proteolysis (“foreign” recognition)
non-specific binding of Fe2+ to DNA  OH· strand breaks, base alteration
DNA damage DNA
(Fenton)