CHRONIC EFFECTS OF ZnO NANOPARTICLES TO EISENIA FETIDA
Chamila Samarasinghe
10/09/2019
The University of Newcastle
OUTLINE
• Introduction – Nanoparticles, Need for toxicity data, earthworms • Objectives of the experiment • Experimental design • Findings of the experiment • Comparison of our results with data available in literature • Challenges • Considerations for risk assessment
NANOPARTICLES
McNeil, SE 2005
• Increased relative surface area • Most of the atoms lay close or at the surface • Different/ enhanced properties compared to their bulk counterparts • Unique physico-chemical characteristics (eg. higher reactivity)
ZnO NANOPARTICLES
• Zn – essential trace element for living organisms • Properties:
ohigh catalytic / UV attenuation oWhitening oAntimicrobial
• Nanoparticle use largely increased in the last decade
ZnO APPLICATIONS
• 3rd Highest global production volume (550 tons) • Cosmetics, sunscreens, toothpastes, beauty products • Industrial coating – Textiles, plastic, wood • Sensors, Sensing devices, Catalysts • Solar cells, LCDs, electronics • Antifouling paints • Antimicrobial usage – Biocides? • Fertilizer
Bondarenko et al., 2013
• Improved properties - more efficiency in industrial applications • Manufacture and utilization – not properly regulated, industry does not
recognize as new chemicals, regulated as conventional bulk chemical compounds
• Volume of the NP enabled products increased • Higher volumes may release to the environment in future • Unique physico-chemical properties – enhanced reactivity, higher stability in
the environment??? • Potential harmful effects to living organisms
NEED FOR NANOECOTOXICITY
Amede et al., 2017
Environmental exposure, common transformations, transport and bioavailability of metal oxide nanoparticles in terrestrial environment
EARTHWORMS
• Important element in the terrestrial ecosystem, Key trophic link • Represent ~ 60 -80% of total soil biomass (Bouche, 1992) • Play key role in soil formation and organic matter breakdown • Maintain & improve soil structure • Contribute in nutrient cycling • Biological indicators of soil health • High potential of large scale exposure of earthworms to any nanoparticle • Model organisms to investigate toxicity of environmental pollutants
https://www.naturespot.org.uk/species/common-earthworm
EARTHWORM LIFE CYCLE
OBJECTIVES OF THE EXPERIMENT
• To assess the impact of ZnO nanoparticles to earthworms • Growth • Survival • Reproduction
• Tissue Zn bioaccumulation due to of ZnO nanoparticle exposure.
OECD PROTOCOLS
Earthworms are introduced to contaminated / spiked soil
Acute toxicity measurement Survival Morphological / behavioural symptoms
Chronic toxicity measurement Survival Reproduction (Cocoon & juvenile count) Biomass change Bioaccumulation
EXPERIMENTAL DESIGN
ZnO Nanoparticles
Uncontaminated soil
Collection (<10 cm – top layer) Processing Characterization
Spiking & Aging ~ 3 months
Introduction of Parent Earthworms
₊
Survival, Weight change &
Bioaccumulation
28 days
56 days Reproduction
SOIL CHARACTERIZATION
pH Sand (%)
Slit (%)
Clay (%)
Texture Total Carbon (%)
Background Zn (mg Kg-1) 6.5
38 33
28 Clay loam
4.99 106.79 ± 3.20
NANOPARTICLE CHARACTERIZATION
(A) XRD Diffractogram and (B) Scanning electron micrographs of ZnO dry nanopowder (Original magnifications: x 10, 000; Scale bar: 200 nm)
0 20 40 60 800
5000
10000
15000
20000
Position [02 θ]
Counts
(A) (B)
SURVIVAL & WEIGHT CHANGE • No significant effects were observed on survival and growth of earthworms
exposed to the ZnO nanoparticle - spiked soils when compared to the control soils.
• Any sublethal effects??
REPRODUCTION - COCOONS & JUVENILES
Reproductive response of the earthworms exposed to ZnO nanoparticles in soil
0 0.1 1 10 100 500 10000
5
10
15
20
ZnO nanoparticle Treatment (mg/ Kg)
Coco
on C
ount
0
0.1
1
10
100
500
1000
**
**
0 0.1 1 10 100 500 10000
10
20
30
40
ZnO nanoparticle Treatment (mg/ Kg)
Juve
nile
Cou
nt
***
***
BIOACCUMULATION OF Zn
• No considerable tissue accumulation of Zn metal was observed in the body tissues of earthworms exposed to ZnO nanoparticle spiked soil.
0 0.1 1 10 100 500 10000
20
40
60
80
100
ZnO nanoparticle Treatment (mg/ Kg)Mea
sure
d t
issu
e Z
n C
on
cen
trat
ion
(µg
/g w
et w
t.)
0
0.1
1
10
100
500
1000
FINDINGS OF THIS STUDY - SUMMARY
• ZnO nanoparticles did not affect earthworm survival and growth. • Above 500 mg Kg-1 concentration, ZnO nanoparticles are likely to cause
reproductive toxicity to the terrestrial organisms.
• > 500 mg Kg-1 – Exceed the expected environmental concentrations
INHIBITION OF EARTHWORM REPRODUCTION AT 1000 mg/ Kg OF ZnO NANOPARTICLES – COMARISON OF DATA
Our Stu
dy
Garcia-Gom
ez, 2014
Alves, 2019
Hooper, 2019
Garcia-Gom
ez, 2019
0
50
100
150
Inhi
biti
on ra
te o
f Coc
oon
prod
ucti
on (%
)
NR
97%
59%50%
100%
Our Stu
dy
Garcia-Gom
ez, 2014
Alves, 2019
Hooper, 2019
Garcia-Gom
ez, 2019
0
50
100
150
Inhi
biti
on ra
te o
f Juv
enile
pro
duct
ion
(%)
NR
92%
72%
40%
100%
CHALLENGES
• Inconsistent data • Different experimental approaches, difficulties in comparison • Interactions with organic material in the environment – soil characteristics • Potential solubility in aqueous medium
• Unavailability of common parameters – additional specific requirements??
• Dispersion conditions • Characterization of nanoparticles in the environmental matrix • Consideration of test conditions for potential artefacts due to their properties)
CONSIDERATIONS FOR RISK ASSESSMENT
• Extensive work already done for the risk assessment of Zn metal / bulk form is a useful starting basis to asses the potential environmental risk
• For initial analysis – no apparent need to go beyond for single nanoparticle risks
• Combined effects of NPs and co-existing contaminants in soil?? • Comparison of data with different types of soils – influence of soil
characteristics • More research is needed to provide insight to the ecotoxicological effects of
exposure to these nanoparticles.
• Continuous exposure • Long exposure period
• Simultaneous exposure from multiple chemical stressors
• Subtle effects • Cumulative and synergistic effects
• Long term impact on the ecosystem
Professor Megh Mallavarapu Professor Ravi Naidu Professor John Aitken Dr. Kannan Krishnan CRC CARE – PhD Scholarship CleanUp 2019 organizing committee GCER &The University of Newcastle