LSU Chemistry Department 6/14/05 1

Stable Free Radicals and Potential Implications for Health Effects of Diesel

Emissions

Barry DellingerBarry Dellinger

Department of ChemistryDepartment of Chemistry

Louisiana State UniversityLouisiana State University

11th Annual Diesel Engine Emission Reduction (DEER) Conference11th Annual Diesel Engine Emission Reduction (DEER) ConferenceChicago, IllinoisChicago, Illinois

August 12 August 12 -- 15, 200515, 2005

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Chemical Reaction Zones in a Combustion Device

PersistentRadicals

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Chemical Reaction Zones in an Automotive Engine System

Pre-Flame and Flame Zone

Cool Zone

Post-Flame Zone

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Radical Stabilities/Reactivities

10-6 s 10-3 s103 s100 s

OH

H

O H2C CH

CH3

H

H

HH

H

H2C C CHOH

O

R

RO2PAN

O2

NO

OH

O

NO

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Resonance Stabilized Semiquinone Radicals

OH

OH

O

OH OH

O O

OH- H

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Oxidative Cycles of Semiquinones

OH

OH

O

O_

O

O

NADPH orNADP+ or

AscorbateDehydroascorbate

O2_

H2O2 O2_

HO

Fe2+

Fe3+

Cellular Damage

O2

NADPH or Ascorbate

NADP+ or Dehydroascorbate

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Tobacco Components Producing Radicals

Total Protein

Lignin

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EPR Spectra of Combustion-Generated Particles

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Source Distribution of PM2.5

Adapted from Gray and Cass, 1998

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EPR Spectra of PM2.5 Samples

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Lung Epithelial Cell DamageIncinerator Fly-ash Baton Rouge PM2.55 City PM2.5

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Stabilization Mechanisms

OH

OH

Cu) O

OH

OH

O

CuOO O

OH

H

Physisorption

- H2O

Chemisorption

OH

Cu(II)

O

O

O

e- transfer

RadicalFormation

OH

O.

OH

O

Cu(I)

O O

Cu(I)

O O

.

Mesomerization

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4000 3500 3000 2500 2000 1500

Aromatic ring-breathing1592 and 1493 cm-1

After 1-hour exposure

Cab-O-Sil before dosing

27% reduction inO-H stretching intensity

Isolated SiO-H group3747 cm-1

FTIR Spectra of 2-Chlorophenol on Silica at 350 C

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XANES Spectra of CuO/SilicaReduction by Phenol

8975 8980 8985 8990Photon Energy (eV)

DataBest fit: 3% Cu(0), 45% Cu(I) and 52% Cu(II)3% Cu(0), 55% Cu(I) and 42% Cu(II)3% Cu(0), 35% Cu(I) and 62% Cu(II)

Cu(I)Cu(0)

Cu(II)

Abs

orb a

nce

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Radical Yieldson 5% Cu(II)O/Silica

30025020015010050Adsorption Temperature, oC

1400

1200

1000

800

600

400

200

Spin

s/g

1,2-DichlorobenzeneChlorophenolChlorobenzeneHydroquinoneCatechol

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Radical g-Valueson 5% Cu(II)O/Silica

2.0080

2.0075

2.0070

2.0065

2.0060

2.0055

2.0050

2.0045

g va

lue

35030025020015010050Adsorption Temperature oC

Monochlorobenzene1,2-Dichlorobenzene2-ChlorophenolCatecholHydroquinone

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Substitution Effect on Radical Structure

CuO

Cu(I)

OCl

Cu(II)O

Cu(I)

OHCl

.

Chlorine Substitution

Hydroxyl Substitution

Cu(II)O

Cu(I)

O.HO

Cu(II) Cu(I)

OO

Cu(II)O

Cu(I)

O.O

H

H

HO

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Engineered Nanoparticle Surrogates~5 nm CuO ~30 nm CuO CuO/Carbon

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FMS0.5 suspended in 50 μl sterile, isotonic saline (1μg/μl) were instilled into the lungs of mice. At 24 h, , the mice were euthanized and exsanguinated. Peripheral blood and tissues were isolated. FMS were seen freely floating in the(a,d) peripheral blood (inset: FMS-laden macrophage; (b,e) within the myocardial tissue; and (c,f) within cells of the airways. (a,b,c) Brightfield images. (d,e,f) Fluorescent images: excitation/emission at 560/610 nm. Arrows indicate representative FMS0.5 in tissues. Scale bar = 20μm.

Nanoparticles Diffuse from the Lung into the Blood and Tissues

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A) Standard curve of FMS1 in isotonic saline containing 0.02% tween-80. The FMS1 standard curve was linear from 49 – 1562 particles/ml Mean values ( ) ± Cl plotted (n=3; r2=0.9987). B) Approximately 60%, 7%, and 2% of the total instilled dose was observed in the lungs, heart, and liver; respectively Values in the organs are percentages of the total dose of FMS1 administered to the mouse.

Biodistribution of FMS0.5 30 min After Intratracheal Instillation

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Surrogates Reduce Epithelial (and Myocardial) Cell Viability

5,000 cells/well plated 24 hrs prior to exposure176 ug exposure for 24 hrs3 wells/samplechemisorbed @ 230oC

C+ C- MCP/CuO DCBz/CuO MCBz/CuO Bz/CuO CT/CuO

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Radical/Particle SystemsSuppress P450 in the Lung

vehicle Cat./CuO DBBz/CuO DCBz/CuO

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Radical/Particle SystemsIncrease Free Radical Damage

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Myocardial ONNO .- is Evidence of Increased Oxidative Stress to Heart Tissue

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Increase in ROS in Heart Tissue by DHE Staining

Control Catechol/CuO

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Stabilization Mechanisms

OH

OH

Cu) O

OH

OH

O

CuOO O

OH

H

Physisorption

- H2O

Chemisorption

OH

Cu(II)

O

O

O

e- transfer

RadicalFormation

OH

O.

OH

O

Cu(I)

O O

Cu(I)

O O

.

Mesomerization

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Only Chemisorbed Chlorophenol Increases Lipid Peroxidation

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X

MetalO2

O

Men

O O

O

Men-1

O O

O

Me

ClPCDD/F

.

.

O.

O

HC Soot

O.2

-

ROS

CO

.OHOxidative

Stress

`

DNA damage

Lung disease

Heart disease

Liver disease

HO

HO

O

O

O

O

O2H2O2

O2

O2

NADPH or Ascorbate

NADP+ or Dehydroascorbate

Fe3+ Fe2+

OH

cellular damage

O

HOH2O H3O+

pH≥7.2

Airborne FineParticles

DNA Damage

Lung Disease

Heart Disease

Liver Disease

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WayneWayne BackesBackes Randy HallRandy Hall Cheri McFerrinCheri McFerrin Hieu Hieu TruongTruong Mikhail Bryukov

Steph Steph CormierCormier Lavri Lavri KhachatryanKhachatryan SofiaSofia MaskoMasko Kurt VarnerKurt Varner Shadrack Ndyaie

Catherine EvansCatherine Evans Slawo LomnickiSlawo Lomnicki ErwinErwin PoliakoffPoliakoff JudyJudy WornatWornat

Rafael CuetoAndy DeutschGuis Squadrito

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