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NANOPARTICLE SURFACE ACTIVITY
UNDERSTANDING MEASURING AND
INTEGRATING IT INTO DOSIMETRY
Dhimiter Bello ScD MSc
Assoc Professor Exposure Biology
Work Environment Department
College of Health Sciences
UMass Lowell
Visiting Scientist Harvard School of Public Health
Molecular amp Integrative Physiological Sciences
Department of Environmental Health
Dhimiter_Belloumledu SNO Venice 03112015
The CHALLENGE
THE VIAL THE FILTER
dhimiter_belloumledu 2
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 3
Dose Metrics A Historical Perspective
dhimiter_belloumledu 4
MASS(ugm3)
RESPIRABLE
FIBERS
(fm3)
Viable amp Total
Microorganisms
(m3)
Departing from Mass (OR NOT)
dhimiter_belloumledu 5
Maynard amp Kuempel 2005
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
The CHALLENGE
THE VIAL THE FILTER
dhimiter_belloumledu 2
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 3
Dose Metrics A Historical Perspective
dhimiter_belloumledu 4
MASS(ugm3)
RESPIRABLE
FIBERS
(fm3)
Viable amp Total
Microorganisms
(m3)
Departing from Mass (OR NOT)
dhimiter_belloumledu 5
Maynard amp Kuempel 2005
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 3
Dose Metrics A Historical Perspective
dhimiter_belloumledu 4
MASS(ugm3)
RESPIRABLE
FIBERS
(fm3)
Viable amp Total
Microorganisms
(m3)
Departing from Mass (OR NOT)
dhimiter_belloumledu 5
Maynard amp Kuempel 2005
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Dose Metrics A Historical Perspective
dhimiter_belloumledu 4
MASS(ugm3)
RESPIRABLE
FIBERS
(fm3)
Viable amp Total
Microorganisms
(m3)
Departing from Mass (OR NOT)
dhimiter_belloumledu 5
Maynard amp Kuempel 2005
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Departing from Mass (OR NOT)
dhimiter_belloumledu 5
Maynard amp Kuempel 2005
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
SURFACE AREA METRIC
dhimiter_belloumledu 6
Maynard amp Kuempel 2005
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Surface Area as a Dose Metric
7
BET Surface Area [cm2]Lung [g]
Inflam
ma
tion P
MN
()
DOSE = f(SURFACE AREA SURFACE ACTIVITY)
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
bull Size Distribution
bull Surface Area
bull Surface Chemistry
bull Surface Charge
bull Bulk Chemical Composition
bull Metals amp Impurities
bull Morphology
bull Crystalinity
bull Biopersistence
bull Metal Leachinghellip
28 Properties in all (ICON 2007)
Physicochemical amp Morphological Properties Influencing
Toxicity
8
Surfaces are NOT
equal
Multiple parameters
related to surface
properties (SP)
How to measure these
surface properties
How do these
measures relate to
biologytoxicology
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
(1) Sotiriou et al Curr Opin Chem Eng 2011 1 3 ndash 10
(2) Xia et al ACS Nano 2011 5 1223 ndash 1235
(3) Napierska et al Particle and Fibre Toxicology 2010 739
(4) Teleki et al Chem Mater 2009 21 2094ndash2100
(5) Sotiriou et al Adv Funct Mater 2010 20 4250ndash4257Demokritou ES Nano 2013
Next generation ENM Safer-by design
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Impurities amp Bioactivity
Doping of SiO2 NPs with
MeOx NP or Men+
CB +Fe2O3
Synergistic ROS generation
10
Guo et al PFT 2009Limbah et al ESampT 2007 41
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Examples illustrate the importance of material composition electronic structure bonded
surface species (eg metal-containing) surface coatings (active or passive) and solubility
including the contribution of surface species and coatings and interactions with other
environmental factors
Nel et al Science 311 622 (2006)
Oxidative stress ndash An important mechanism of NP
toxicity
Oxidative stress (OS) has
been recognized in vivo and
in vitro systems as one
such major pathway and is
being explored for ENM
toxicity screening purposes
(Nel et al 2006 Xia et al
2006 Borm et al 2007
Ayres et al 2008 Rogers et
al 2008 Bello et al 2009
Lu et al 2009 Meng et al
2009)
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
In vivo InhalationOral Administr Skin irritationAquatic animals
hellip
hellip
hellip
hellip
hellip
hellip
hellip
hellip
In vitro Cell free System
Cellular System
System Markers (of Oxidative Stress)
ROS - Reactive oxygen species DTT-Dithiothreitol DCFH-DA-Dichlorofluorescin diacetate HO-1 Heme
oxygenase-1 SOD-superoxide dismutase GSH-Glutathione EPR-Electron paramagnetic resonance ESR-
electron spin resonance
hellip
hellip
Measure ROS generationDTT assay
DCFH-DA assay
EPRESR
Cell viabilityMitochondrial dysfunction
ROS ndash DCFH-DA assay
Activation of pro-inflammatory pathway
Inflammatory factors cytokine production
Redox enzyme expression (HO-1 SOD)
DNA damagecell mutagenesisproliferation
Luciferase Reporter Cyt C
Similar to Cellular system
GSH depletion
Hematological biochemical
and pathologic change
12
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Approaches
Acellular
ESREPR
o Spin trapping with select
agents (eg DMPO)
DTT
o Colorimetric
DCFH
o Fluorescence (RO2 RO OH
HOCl and ONOO but not O2- and H2O2 )
FRAS
o Human Serum - Colorimetric
Property
dhimiter_belloumledu 13
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Human Blood Serum(containing endogenous
antioxidantseg VITE VITC)
Residual Serum Antioxidants
Fe3+
IN FRAS REAGENTFe2+
+TPTZ
BLUE COLORCOMPLEX
MEASURE ABSORBANCE AT 593nm BY VISIBLE
SPECTROPHOTOMETRY
FRAS REACTION
HOmiddot
ANALYTICAL APPROACH TO DETERMINE THE DEGREE OF OXIDATIVE STRESS EXERTED BY NANOPARTICLES IN HUMAN BLOOD SERUM BY FERRIC REDUCTION ACTIVITY OF SERUM (FRAS ASSAY)
O
CH3
CH3
OH
CH3
H3C
-Tocopherol(Reduced form)
R
O
OHO
HO
OHHOH2C
L-Ascorbic Acid(Reduced form)
O
OO
O
OHHOH2C
+ H2O
L-Dehydroascorbic Acid (Oxidized form)
O
CH3
CH3
OCH3
H3CR
Stabilized Tocopherol Radical
Oxidatively Damaging Nanoparticles+
+ ROOH
L-Ascorbic acid(Reduced form)
L-Dehydroascorbic acid(Oxidized form)
α-Tocopherol(Reduced form) Stabilized Tocopheroxyl Radical
ROOmiddot
O
TPTZ-246-
Tripyridyl-135zine
RED
BLUE
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Oxidative Stress vs Inflammation
X2012
dhimiter_belloumledu 16
P
M
N
c
m
2
DCFH ESR
AMcell -ESR Luciferase
Activity
R2=074 R2=079
R2=095
R2=088
Rushton et al 2010
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
FRAS vs ESR
dhimiter_belloumledu 17
y = 00273x + 67895Rsup2 = 00013
0
20
40
60
80
100
120
140
160
0 50 100 150
ES
R (
H2
O2
eq
)
FRAS TEU (mM)
ESR vs FRAS
Bello D unpublished data
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Protocols
dhimiter_belloumledu 18Pal et al 2011 Dose-Response
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
dhimiter_belloumledu 19
(A) (C)
(B) (D)
mR
OS
(micro
mo
l H
2O
2E
q
mg
)sR
OS
(micro
mo
l H
2O
2E
q
m2)
mR
OS
(micro
mo
l H
2O
2E
qm
g)
sRO
S (micro
mo
l H
2O
2E
q
m2)
mBOD (micromol TEU Eqmg)mBOD (micromol TEU Eqmg)
sBOD (micromol TEU Eqm2) sBOD (micromol TEU Eqm2)
Spearman 085
R2 Linear 078
Spearman 062
R2 Linear 068
Spearman 058
R2 Linear 032
Spearman 030
R2 Linear 011
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
FRAS
Positive
(2128)
Negative
(728)
DCFH
Positive
(1028)10 0
Negative
(1828)11 7
N=28
Pal et al 2013 J Nano Research
Comparison of FRAS amp DCFH for the 28 ENM
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Acellular vs
Intracellular
GSHGSSG
dhimiter_belloumledu 21
Pal et al 2014
J Nano Research
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
dhimiter_belloumledu 22
Pal et al J Nano Research 2013
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Oxidative Stress vs Inflammation
dhimiter_belloumledu 23
TiO2_F
TiO2_D
Ag-35
Cu-60
MWCNT_MitsuiNi-60
Cristobalite
1
10
100
1000
10000
1 10 100 1000 10000
PM
N N
um
be
r x
10
^7m
^2
sBOD (nmol TEUsm2)
FRAS vs Inflammation
Bello Unpublished data PMN data Alison Elder U Rochester
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Physicochemical Properties amp FRAS Oxidative Stress
24
BOD ~ with SSA and the total
content of Redox-active metalsBello Hsieh et al 2010 Nanotoxicology
BO
D (
TE
Us
n
mo
l1
0m
g)
BOD (TEUs) = 075 SSA (m2g-1) + 0418TMe (ppm) R2=093
Hsieh Bello et al 2013
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
25
Distribution of FRAS BOD Values of 145 Tested
ENMs
1 BOD is expressed as Trolox
equivalent units (TEUs
μmol L-1)
2 Total antioxidant capacity of
normal blood serum is 535 plusmn15 μmol L-1 TEUs
3 1000 TEUs BOD near
complete depletion of the
antioxidants pool (5mgmL)
4 15 TEUs = The non-
significant BOD
5 25 of ENM lt15 TEUs
Hsieh et al 2013 Small
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Between-Class Variations in BOD
26
Hsieh Bello et al Small 2013
N=138
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Within-Class Variation in BOD CNTs
27
Hsieh Bello et al Small 2013
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Metal Oxides
28
TiO2
Hsieh Bello et al Small 2013
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
100 10 1
00
01
02
03
04
05
MWCNTs
SWCNTs
sB
OD
(T
EU
s micro
mo
l m
-2 m
L-1)
Outer Diameter (nm)
Effect of CNT OD size on BOD
Surface Activity of narrow CNTs is
much higher than for larger CNTsHsieh Bello et al 2011 Nanotoxicology
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Surface Activity vs ENM Class
dhimiter_belloumledu 30Hsieh Bello et al Small 2013
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Outline
Surface in Inhalation Dosimetry
o Surface Area amp Activity
o Bench top Technologies and Options
Surface in ENM Exposure Assessment
o Gaps amp Needs
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 31
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
dhimiter_belloumledu 32
Brouwer et al J Nanoparticle Res
2013 15 2090
Exposures to ENM by
Task
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Surface Area vs Number Concentration
dhimiter_belloumledu 33
Brouwer et al Journal of Nanoparticle Research 2013 15(11)
DOI101007s11051-013-2090-7
NSAM
SMPS
lt100 nm amp
ALVEOLAR
O gt100 nm amp TB
200-1000 nm amp
ACTIVE SA (LQ1-DC)
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
ENM Exposures
320201
534
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
35
Nanoparticle Emissions from Commercial Photocopiers
Bello et al Nanotoxicology 2012
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Morphology amp Chemistry
36Bello et al Nanotoxicology 2012
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Important Real-World Lessons
NP Exposures are often to MIXTURES
PCM Properties along the life cycle of NEPs may be DIFFERENT from input ENMso Toxicological properties ndash likely DIFFERENT (by how much amp what
direction)
Multi-metric approach ndash necessaryo Sufficient
o Interpretation
Exposure- dose equivalency for in vitro or in vivo workhellipo mass number surface area elemental composition
dhimiter_belloumledu 37
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Outline
Current Metrology amp Exposure Monitoring for ENM
Surface Area
Surface Activity ndash what does it tell us
o Benchtop Technologies and Options
o Validity of the Concept
Near Real-time monitoring of SAr and SAc
dhimiter_belloumledu 38
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Nanodevices
(FP7 project)httpwwwnano-
deviceeuindexphpid=328
dhimiter_belloumledu 39
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Near-Real Time ROS Is Almost Here
dhimiter_belloumledu 40
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
dhimiter_belloumledu 41
Vogel 2013
3 min cycle
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Direct On-filter FLD detection
dhimiter_belloumledu 42
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
CONCLUSIONS
dhimiter_belloumledu 43
Surface activity appears to integrate
across multiple PCM parameters including
surface area
Therefore it is a critical parameter to
capture preferably in near-real time
Additional parameter to SA
The challenge- to develop the right
technology
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
My Collaborators
dhimiter_belloumledu 44
Prof P
Demokritou HSPHProf E Rogers
UMass LowellProf D F Schmidt
UMass LowellProf P Gaines
UMass Lowell
Prof B Wardle
MIT Prof J Isaacs
NEU
Dr D
Brouwer TNOProf Redlich Yale
Prof J Mead
UMass Lowell
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Questions
X 2012DHIMITER_BELLOUMLEDU
45
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
112920
1046
Monitoring NP Exposures Instrumentation
CCI
FMPS
APS
NPS-500
Nano-ID
CPC
Q-Trak
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
Personal Samplers
Personal size selective
impactors
o Naneum Aerosol PS 300
o Several miniaturized impactors
Quazi Personal Real-Time
Monitors
o Philips NanoTracer (10-300 nm
TNC SD)
oDiSCmini (Matter-Aerosol Inc)
dhimiter_belloumledu 47
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
BOD Correlates Well with TELI in E-Coli
dhimiter_belloumledu 48
Guo amp Gu et al 2011 EST
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
ldquoToxicity Screening tests for new nanomaterials
products are urgently needed Whilst recognizing that
oxidative stress potential may not be predictive of all
possible adverse outcomes tests based upon oxidative
potential maybe an invaluable tool for initial screening
and classification of the relative biohazard of such
materialsrdquo
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
OXIDATIVE STRESS ANTIOXIDANTS
bullAscorbic acidbullTocopherolbullFolate
bullCatalasebullGlutathionie PeroxidasebullSuperoxide Dismutase
ENDOGENOUS PRODUCTION OFFREE RADICALS ROS AND RNS
eg bullOH bullO2- ROObull RObull H2O2
ONOO-
SMOKING
AIR POLLUTIONRADIATION
CHEMICALS
INCREASED PRODUCTION OFFREE RADICALSREACTIVE OXYGEN AND
NITROGEN SPECIES (ROS RNS)eg bullOH bullO2- ROObull RObull H2O2
ONOO-
OXIDATIVE DAMAGE
DNA DamageProtein
DenaturationInflammation
Lipid Peroxidation
Mutation
METAPLASIA CARCINOGENESIS
Defective Protein Structures
Cytokine and Chemokine Release
Mitochondrial andother Membrane Damage
Complement Activation and WBC Infiltration
DEVELOPMENT OF CHRONIC DISEASES
NANOPARTICLES
Biological Oxidative Stress
Key Metric Biological Oxidative Stress
What SAMPLE TYPE do we use to check for this Oxidative Stress
top related