Time-resolved and Online Determination of Reactive Oxygen Species … · 2018. 8. 14. · Barcelona, 25 April 2012 M. Kalberer Measuring ROS concentrations online • Many current

Time-resolved and Online Determination of

Reactive Oxygen Species (ROS) in Ambient Air

Markus Kalberer, Stephen Fuller

Department of Chemistry

University of Cambridge, UK

M. Kalberer AirMonTech, Barcelona, 25 April 2012

Overview

• Oxidative stress - a possible mechanism of particle-induced

health effects

• A new instrument measuring reactive oxygen species (ROS)

concentration on-line: design and instrument characterization

• First applications of the instrument


Ambient particle cause negative biological/health effects

as observed in epidemiological and laboratory studies

Health effects of aerosols

London-Smog, 1952 Deaths SO2 aerosol particles (dust)

Six cities study, Dockery et al. 1993


What particle properties are relevant for

biological effects?

particles e.g.,

from combustion

sources

1m

potential damaging properties

- organic components

- metals

- mass, size, surface properties

effects in the cell

- ROS?

- oxidative stress?

- inflammation


Oxidative stress in the lung

H2O2

O2-

OH

Respiratory

tract lining

fluid

Lung epicelial

cells

Circulatory

system

glutathione

vitamin C vitamin E

Antioxidants ROS

• Oxidative stress = disturbance of prooxidant- antioxidant balance leading to potential

biomolecular damage

• Oxidative stress can be induced by ROS in the lung lining layer

• Antioxidant depletion may cause damage to cell membranes, proteins

& DNA

Antioxidants

ROS

anti-ox

proteins org. peroxides & radicals


PM induced ROS in the lung

Formation routes of ROS from aerosol particles:

1. Particle bound ROS – e.g. H2O2, ROOH, radicals

= reactive and short-lived

2. (Catalytic) ROS production in the lung lining fluid by

redox active components; e.g., quinones and metals

3. Metabolism of organics (e.g. PAHs) may lead to ROS

formation


Measuring ROS concentrations in solution

- the reaction system

• DCFH is oxidised by ROS in the presence of Horseradish

Peroxidase to the fluorescent product DCF; excited at 470 nm

and emitting at 520 nm.

• DCFH reacts with almost all ROS


HRP

HRP-I

HRP-II

ROS (ox) (H2O2)

DCF-H

DCF

DCF-H

DCF

ROS (red) (H2O)

• The fluorescence is directly proportional to the concentration of DCF

• While the DCFH assay gives a linear relationship with H2O2, organic

peroxides react much slower with HRP.

• This assay gives an indication of ROS activity related to an

equivalent H2O2 concentration. NOT an absolute ROS

concentration.

I [DCF] [H2O2]

(HRP = Horesradish Peroxidase)

Measuring ROS concentrations in solution

- the reaction system


Measuring ROS concentrations online

• Many current aerosol studies measuring composition use

off-line filter collection techniques

• The time delay between collection and analysis possibly

allows for degradation and loss of reactive components

• Online analysis allows for fast analysis that minimizes

loss of reactive components and allows for higher time

resolution

• Online analysis allows for high time resolution


Fast Online Quantification

of Oxidizing Particle Components


Water Bath

37°C

hydrophobic

filter

Continuous

collection and

solvent

extraction

Connected to LabView program

0.25 ml / min

5 slpm

sampling

Aqueous

Aqueous

To vacuum

pump

HRP solution

To mix with

DCFH solution

Aerosol inlet

15 c

m

Fast Online Quantification

of Oxidizing Particle Components


Particle into liquid sampling

0

0,2

0,4

0,6

0,8

1

0 100 200 300 400 500

Co

llecti

on

Eff

ecie

ncy

Aerodynamic Diameter / nm

Collection Effeciency


ROS quantification

Water Bath

37°C

Hydrophilic

filter

-Continuous

filter collection

and solvent

extraction


0.25 ml / min

5 slpm

sampling

Aqueous

Aqueous

Fan cooled 5 W 470 nm LED

Spectrometer


Experimental set up

Water Bath

37°C

Hydrophilic

filter

Continuous

filter collection

and solvent

extraction


0.25 ml / min

5 slpm

sampling

Aqueous

Aqueous

Fan cooled 5 W 470 nm LED

Spectrometer


Sensitivity calibration with H2O2

Fluorescence intensity after reaction has gone to completion after


Normalised to equivalent

fluorescein fluorescence to

account for non linear

spectrometer behaviour at

higher concentrations




R2 = 0.99

0

500

1000

1500

2000

0 100 200 300 400 500 600 700

[Flu

ore

sc

ien

] / n

M

[H2O2] / nM

Dynamic range:

ca. 2 orders of magnitude

Detection limit:

in liquid H2O2: ~ 10 nM

ambient air (5lpm):

4 nmol / m3


Smog Chamber Experiments

• Experiments at Paul Scherrer Institut (PSI), Switzerland

• Primary moped emissions photochemically aged in smog

chamber

• Mopeds conforming to both Euro 1 and Euro 2 emissions

regulations

27 m3

Chamber

UV + NOx + O3



Secondary Organic Aerosol Formation

Primary organic aerosol

(POA)

Soot

Gaseous volatile organic

compounds (VOCs)

Oxidation by ozone and

OH in the presence of

UV light and NOx

Oxidised VOCs

condense to form

secondary organic

aerosol (SOA)

Primary

Processing

Secondary



Charcoal Denuder

Online ROS Instrument

Remove gaseous

oxidants



With photochemical

SOA formation:

increase of ROS

ROS =

ca. 1 nmol μg-1 SOA

50

40

30

20

10

0

SO

A m

ass / µ

g m

-3

2.52.01.51.00.50.0

Time after lights on / hours

50

40

30

20

10

RO

S c

on

ce

ntr

atio

n /

nM

ole

s m

-3

Euro 1 SOA mass

Euro 2 SOA mass

Euro 1 ROS concentration

Euro 2 ROS concentration


Smog Chamber Experiments: POA vs. SOA

ROS concentration in POA (before photochemical aging): negligible


Smog Chamber Experiments: Euro1 vs. Euro2

ROS / g POA large differences between Euro 1 and Euro 2

8

6

4

2

RO

S c

on

ce

ntr

atio

n n

orm

alis

ed

to

PO

A m

ass

/ n

Mo

les m

-3 µ

g-1

2.52.01.51.00.50.0

Time after lights on / hours

Euro 1

Euro 2


Ambient measurements

ROS in ambient urban air (Cambridge)

0

20

40

60

80

100

120

140

160

180

200

11:52:48 12:21:36 12:50:24 13:19:12 13:48:00 14:16:48 14:45:36 15:14:24 15:43:12

nM

ole

s m

-3

Time

clean air ambient air clean

air

F

luo

res

cen

ce s

ign

al

11:50 12:50 13:50 14:50 15:50


Conclusions

• Design of online instrument to quantify ROS

• ROS “scavenged” within seconds: reactive ROS are not lost

• Time resolution: ca. 10min

• Detection limit: 10nmol (H2O2), 4 nmol / m3

• ROS only observed in SOA from moped emissions

ROS concentrations of ca. 1 nmol μg-1 SOA

• No ROS in primary moped emissions


Acknowledgements

PSI, Villigen – Smog Chamber Experiments

Stephen Platt

Josef Dommen

Andre Prevot

Urs Baltensperger


Thanks for your attention

Documents

Time-resolved and Online Determination of Reactive Oxygen Species … · 2018. 8. 14. · Barcelona, 25 April 2012 M. Kalberer Measuring ROS concentrations online • Many current