Formation of Reactive Oxygen Species - …duahn/teaching/Lipid oxidation/Formation of...Ground-state...

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Formation of Reactive Oxygen Species

Transcript of Formation of Reactive Oxygen Species - …duahn/teaching/Lipid oxidation/Formation of...Ground-state...

Formation of Reactive Oxygen Species

Ground-state oxygen (3O2 )

Oxygen is the most important factor on the development of lipid peroxidation.

Ground state oxygen is itself a radical, with two unpaired electrons each located in a π* antibonding orbital

Ground state oxygen has its outermost pair of electrons parallel spins: does not allow them to react with most molecules

Ground-state or triplet oxygen is not very reactive

Can be activated by the addition of energy, and transformed into reactive oxygen species

Singlet oxygen (1O2 )

Formed from triplet oxygen

Conversion of oxygen to singlet state can be accomplished by photosensitization in the presence of suitable sensitizers, such as chlorophyl, or heme pigments myoglobin or hemoglobin or by their derivatives

Has a pair of electrons with opposite spins

Singlet oxygen (1O2 )

1ΣgO2 has no unpaired electrons and thus does not qualify as a radical. 37.5 kCla above the ground state O2

1ΣgO2 usually decay to 1ΔgO2. 22.4 kCla above the ground state O2

1ΔgO2 is not a true radical but is reported to be an important ROS in reactions related to ultraviolet exposition (UVA, 320-400 nm)

Excess singlet oxygen formation can lead to certain diseases: porphyrias

Superoxide anion (O2-)

Monovalent reduction of triplet oxygen produces superoxide

Formed in almost all aerobic cells: mitochondriaOutside of mitochondria: ER through oxidation process of

cytochrome P-450 and NADPH-cytochrome c reductaseCu+2 + ascorbate + O2 --- O2

-

Fenton reaction also produces superoxide

Superoxide anion (O2-)

Not reactive enough to abstract hydrogen from lipids

Cannot enter the hydrophobic interior of membrane because of its charged nature

Can produce hydrogen peroxide (Dismutation)

Involved in hydroxyl radical formation

Can also react with nitric oxide (NO.) to produce peroxynitrate (OONO-)

Hydrogen peroxide (H2 O2 )

Not a radical

Important in biological systems because it can pass readily through cell membranes

Superoxide-generating systems produces H2 O2 by non- enzymatic or SOD-catalyzed dismutation

2 O2 - + 2 H+ -----> H2 O2 + O2 (SOD)

Hydroperoxyl radical (HO2.)

Protonation of O2 - yields the HO2.

Hydroperoxyl radical (HO2.) is more reactive than superoxide

and can enter membrane fairly easily.

The pKa of HO2. is 4.7-4.8, and so only 0.25% of O2 -

generated in physiological conditions is hydroperoxyl radical. Localized pH drop can exist

Hydroxyl radical (OH·)

The most reactive oxygen species known: site specific reaction

Can be produced by high-energy ionizing radiationH2 O -----> ·OH + ·H + H2 + H2 O2 + H3 O+ + e-aq

(ionizing radiation)In vivo production comes from metal-dependent (Fe, Cu)

breakdown of hydrogen peroxideFe2+-dependent decomposition of H2 O2 (Fenton reaction)Fe2+ + H2 O2 -----> Fe3+ + ·OH + OH-Fe2+ + H2 O2 -----> ferryl? -----> Fe3+ + ·OH + OH-

Hydroxyl radical (OH·)

Fe3+-dependent multi-stage decomposition of H2 O2

Fe3+ + H2 O2 -----> ferryl + H2 O2 -----> perferryl + H2 O2 --- --> .OH

Fe2+-EDTA dependent decomposition of H2 O2

Fe2+-EDTA + H2 O2 -----> intermediate species (ferryl) ----- > Fe3+-EDTA + ·OH + OH-

Ozone (O3 )

Ozone is not a free radical

Ionizing radiation of oxygen produces ozone

As singlet oxygen, it stimulates lipid peroxidation

Can induce damages at the lipid and proteins

Lipid (R.), peroxyl (ROO.) and alkoxyl radical (RO.)

Very strong reactivity and can abstract hydrogen atom from lipids

Can be formed from the lipid radicals by iron complexesROOH + Fe3+-complex -----> ROO. + H+ + Fe2+complexROOH + Fe2+-complex -----> RO. + OH- + Fe3+-complex

Iron-Oxygen Complexes

Ferryl (Fe4+) and perferryl (Fe5+) radicals

Powerful oxidants as a component of enzyme or simple iron complex

Ferryl species are generated by the interaction of H2 O2 with metmyoglobin

Fe(II)-complex + ROO•

+ H+ Fe(III)-complex + ROOH Fe(II)-complex + RO•

+ H+ Fe(III)-complex + ROH

Nitric oxide (.NO)

Produced in various types of cells

Is not too reactive (poorly oxidizing function), even antioxidant under physiological concentrations (up to 100 nM)

Reacts rapidly with oxygen to yield nitrogen dioxide (.NO2 ) which in turn may react with .NO to yield nitrogen trioxide (N2 O3 )

Rapidly react with O2- and produce extremely reactive peroxinitrite

(ONOO-) which mediates oxidation, nitrosation, and nitration reactions

ONOO- also decomposes to produce OH. radical

Thiyl radicals (RS.)

Thiol compounds (RSH) are frequently oxidized in the presence of iron or copper ions:

RSH + Cu2+ ----> RS. + Cu+ + H+

These thiyl radicals have strong reactivity in combining with O2

RS. + O2 ---> RSO2 . They are able to oxidize NADH into NAD., ascorbic acid and

to generate various free radicals (.OH and O2-).

Thiyl radicals can also be formed by homolytic fission of disulfide bonds in proteins

Irradiation

Radiation: Energy moving through space in invisible wavesLight, infrared heat, microwaves, TVs all use radiant energy

Ionizing radiation: Shorter wavelengths in radiation. Capable of converting atoms and molecules to ions via the removal of electrons. Destroy DNA bonds in bacteria, pathogens and insects

Types of IrradiationRadio Active Nuclides

X-rayGamma Rays

Cobalt-60Cesium-137

Electron Beam

Linear Accelerator

E-beam Irradiation

Better Consumer Reception for Food Irradiation

X-ray can be generated

Other Industrial Uses

Physical and Chemical Modification and Cross-linking

Effects

Provides increased tensile, impact, and abrasion strength for wire, cable, and tubing

Improves product performance

Gamma Irradiation

The method of choice for

Sterilization of single-use medical supplies such as syringes, catheters, IV sets, gloves, face masks and more

Elimination of organisms from pharmaceuticals such as ointments and solutions

Advantages

Offers superior material penetration

Economical for high and/or low volume operation

Is highly time-efficient

Offers versatile irradiator designs Imposes minimal restrictions on product design and packaging

Irradiated Foods Marketed in the U.S.

Irradiated ground beef and poultry

18-20 M Lb sold in 2004

Fruits & vegetables

2 M lb sold annually

Mango, papaya, guava are currently sold by US retailers

Growing interest in blueberries, cherries, raspberries

The amount of irradiated tropical fruits will increase rapidly in the future

Spices and Botanics

Commercially irradiated since 1986

175 M Lb of (1/3 of commercial spices consumed in the US) are irradiated annually

Effect of Irradiation

Maximum Irradiation Dose

year Food Dose (kGy)

Purpose

1963 Wheat flour 0.2-0.5 Control molds

1986 Fresh fruit & vegetables 1.0 Inhibit sprouting Delay ripeningDisinfestation

1990 Poultry meat 3.0

Control pathogens1999 Refrigerated meatFrozen meat

4.57.0

Dehydrated enzymes 10

1986 Dehydrated spices & herbs 30

Shelf-lives of Meat Products after Irradiation

Dose Untreated shelf Irradiated shelf-

Meat Products (kGy ) life (days) life (days)

Beef top round 2.0 8-11 28

Beef burgers 1.54 8-10 26-28

Beef cuts under vacuum 2.0 NA 70

Corned beef 4.0 14-21 35

Whole and minced Lamb 2.5 7 28-35

Andrew et al. (1998)

Exhaustive chemical, biological, and feeding studies have shown irradiated foods to be both safe and wholesome

In 1980, FDA and a Joint FAO/IAEA/WHO Expert Committee had concluded on the wholesomeness of irradiated food that irradiation of any food at <

10 kGy causes no toxicological

hazard and nutritional or microbiological problems

In 1983, a worldwide standard covering irradiated foods was adopted by the Codex Alimentarius Commission

2-ACBs, benzene and methyl benzene (toluene)

Is Irradiated Food Safe?

Lipid Oxidation in Precooked Turkey Breast

0

1

2

3

4

5

TBA

RS

(mg

MD

A/k

g m

eat)

0 kGy 2.5 kGy 5 kGy 0 kGy 2.5 kGy 5 kGy

Aerobic Vacuum

Frozen storage for 3 months

Cholesterol Oxidation in Turkey Thigh

Day 7

Vacuum pkg Aerobic pkg0 kGy 4.5 kGy 0 kGy 4.5 kGy

& 7β-hydroxychol. 36.0c 30.0c 51.9b 86.7a

β-epoxide 0b 0b 0b 7.2a

α-epoxide 0c 0c 6.4b 11.5a

20α-hydroxycholesterol 0b 0b 0b 1.4a

cholestanetriol 0 0 0 1.07-ketocholesterol 2.7c 1.5c 19.0b 27.1a

Total* 38.7c 31.6c 77.3b 134.7a

* μg COPs/g lipid

Vacuum-Packaged Raw Turkey Breast

Color of Irradiated Ground Beef

Nonirradiated 4.5 kGy irradiated

Production of Sulfur Volatiles

COO-

IH3 N-CH

ICH2ICH2ISICH3

Methionine

COOCOO--

IIHH33 NN--CHCH

IICHCH22IISHSH

CysteineCysteine

CH3 -S-CH3 (dimethyl sulfide)

H3 C-S-S-CH3 (dimethyl disulfide)

S=C=S (carbon disulfide)

H3 C-S-S-S-CH3 (dimethyl trisulfide)