Fate of silver nanoparticles in

constructed wetland microcosms

Hannele Auvinena, Diederik Rousseaua, Ralph Kaegib, Gijs Du Lainga

a Ghent University, Faculty of Bioscience Engineering, Ghent, BELGIUM b Eawag, SWITZERLAND

E-mail: Gijs.DuLaing@UGent.be

Introduction: silver nanoparticles (Ag NPs) in

wastewater and the environment

2

Auvinen, H., Gagnon, V., Rousseau, D.P.L., Du Laing, G. (2017). Fate of metallic engineered nanomaterials in constructed wetlands:

prospection and future research perspectives. Reviews in Environmental Science and Bio/Technology, 16, 207-222.

Silver nanoparticles in wastewater and environment

3

• Ag+ ions: bactericidal properties

• Ag NPs slowly release Ag+ ions

• use in e.g. high-performance

clothing, food packaging, bactericidal

coatings and wound bandages

4

• released from consumer products into wastewater during usage

• adverse effects on aquatic organisms

• in sewer systems: organic coating may keep them mobile

Silver nanoparticles in wastewater and environment

Constructed wetlands

5

Drawing by Vincent Gagnon

Ag NPs??

Ag NPs?? Ag NPs??

Research questions

6

How do Ag NPs behave in constructed wetlands and how can their removal be improved?

7

8

• What is the fate of silver nanoparticles

(Ag NPs) in constructed wetlands?

• Do constructed wetlands function as

sinks for these particles?

• Which transformations do they undergo?

Understanding the transformations and removal mechanisms

Understanding the effect of plants and substrate

9

• What is the role of substrate and plant in

removing the Ag NPs from the water

phase?

Understanding the toxicity of ENMs in the wetlands

10

• If Ag NPs are efficiently retained in

the wetland biofilm, can they have

adverse effects on the wetland

microbial community?

• Can the community adapt to the

contamination?

Fate of silver nanoparticles in constructed

wetlands – a microcosm study

11

Auvinen H. , Kaegi R., Rousseau D., Du Laing G. 2017. Fate of silver nanoparticles in constructed wetlands - a microcosm study. Water, Air and

Soil Pollution, 228, 97.

The aim

1. Determine removal efficiency

Effect of aeration

Effect of organic matter build-up (aging)

2. Identify partitioning and transformation mechanisms

12

Figure by: Mark Button

The setup

Influent (+ 50 µg ~10-15 nm citrate-coated AgNPs) and

effluent were sampled weekly for 18 weeks

Silver was quantified in all compartments: water, plant

material, attached on gravel&biofilm and microcosm walls

Electron microscopy was applied to imaging and to study

speciation of silver attached to biofilm

13

Microcosm Silver

nanoparticles

spiked Aeration applied

Organic matter

added

Negative control No No No

Positive control Yes No No

Air Yes Yes No

OM Yes No Yes

Results – removal efficiency

14

Very efficient removal was obtained in all microcosms

0

20

40

60

80

100

Positivecontrol

OM Air

Rem

ova

l eff

icie

ncy (

%)

Microcosm

Results – removal efficiency

15

Very efficient removal was obtained in all microcosms

Retention of silver correlated with solids removal (heteroaggregation)

0

20

40

60

80

100

Positivecontrol

OM Air

Rem

ova

l eff

icie

ncy (

%)

Microcosm

R² = 0.8068

0

10

20

30

40

50

60

70

80

0 100 200 300 400

Eff

luen

t A

g (

µg

/L)

Total suspended solids (mg/L)

Results – distribution within microcosms

16

Most silver was retained in the biofilm

Plant uptake was of minor importance

0

20

40

60

80

100

Dis

trib

uti

on

wit

hin

co

mp

art

men

ts (

%)

Positive control OM Air

Results – transformation of retained particles

17

Based on scanning transmission electron microscopy, silver (Ag) detected

within the biofilm was associated with sulfur (S) in all microcosms

Sulfidation reduces Ag availability and mobility

Positive control OM Air

Conclusions

18

Silver (nanoparticles) was associated with solids in the effluent

Improved removal of silver nanoparticles can be obtained in a

constructed wetland by enhancing solids removal

Conclusions

19

Silver (nanoparticles) was associated with solids in the effluent

Improved removal of silver nanoparticles can be obtained in a

constructed wetland by enhancing solids removal

Biofilm acted as a sink for silver nanoparticles

Silver may accumulate in long-term

can it be toxic to the wetland microbial community?

Conclusions

20

Silver (nanoparticles) was associated with solids in the effluent

Improved removal of silver nanoparticles can be obtained in a

constructed wetland by enhancing solids removal

Biofilm acted as a sink for silver nanoparticles

Silver may accumulate in long-term

can it be toxic to the wetland microbial community?

Plants played a minor role at retaining silver nanoparticles

more details in the following experiment

Substrate- and plant-mediated removal of

silver nanoparticles in constructed wetlands

21

Auvinen H., Sepulveda Vasquez V., Rousseau D., Du Laing, G. 2016. Substrate- and plant-mediated removal of citrate-coated silver nanoparticles in

constructed wetlands. Environmental Science and Pollution Research. 23(21): 21920-21926.

The aim

1. Determine the effect of substrate type on the removal of Ag NPs from

the water phase

2. Verify the effect of biofilm on the removal

3. Identify possible plant-mediated processes in removing Ag NPs from

the water phase

22

The setups

Substrate experiment

Silver nanoparticles in the water phase (100 µg/L) brought

in contact with different substrates

Gravel, sand, zeolite, biofilm-covered gravel

For 24 h – no sulfidation!?

Hydroponic culture

Phragmites australis (common reed) cultivated for 4

weeks in water containing silver nanoparticles (100 µg/L)

Analysis in different plant parts & extraction of Ag

adsorbed to roots

23

Results – removal by different substrates

Removal depended on the type of substrate:

Sand biofilm-covered gravel zeolite gravel

24

a a

b

c

a

b, c

0

20

40

60

80

100

Control MQ Control WW Sand Zeolite Gravel Gravel withbiofilm

Rem

oval

fro

m

wate

r p

hase (

%)

Results – removal by different substrates

Removal depended on the type of substrate:

Sand biofilm-covered gravel zeolite gravel

Possible removing mechanisms of nanoparticle (24 h – no sulfidation!?)

Attachment to surfaces

Aggregation and sedimentation

25

a a

b

c

a

b, c

0

20

40

60

80

100

Control MQ Control WW Sand Zeolite Gravel Gravel withbiofilm

Rem

oval

fro

m

wate

r p

hase (

%)

Results – plant-mediated processes

Most silver remained in the water phase

Roots were able to attach and take up silver nanoparticles to some

extent

Only a very low fraction (1%) of total silver was translocated to the shoots

26

50

30

16

1 0

Water phase

Root surface

Uptake by roots

Translocation

Glass recipient

Conclusions

Presence of biofilm increased the removal of silver nanoparticles from the

water phase significantly

The type of substrate also played a role in the removal. Possibly through:

Attachment to its surface

Aggregation-sedimentation

Immobilization of silver nanoparticles by plant roots occurred

Translocation to aboveground tissue was negligible

27

Susceptibility of constructed wetland

microbial communities to silver nanoparticles

28

Button M., Auvinen H., Van Koetsem F., Hosseinkhani B., Rousseau D., Weber K., Du Laing G. 2016. Susceptibility of

constructed wetland microbial communities to silver nanoparticles: A microcosm study. Ecological Engineering 97: 476-485.

The aim

Silver nanoparticles were efficiently retained by the biofilm, but how is the

toxicity to wetland microbial community at low concentration?

1. Study the toxicity of silver ions and two different types of silver

nanoparticles (PVP and citrate coating)

Compare different silver species

Define toxic dose (dose-response test)

2. Study the adaptation of the community to silver nanoparticles

29

The setup

Microcosms simulating constructed wetlands

Analysis of microbial community function and

structure:

Catabolic capacity: community-level

physiological profiling (CLPP)

30

The setup

Microcosms simulating constructed wetlands

Analysis of microbial community function and

structure:

Catabolic capacity: community-level

physiological profiling (CLPP)

Structure: denaturing gradient gel

electrophoresis (DGGE)

31

Results – Development of microcosm community

32

The biofilm community in the microcosms developed differently from the

community in the water phase (IW)

The largest impact was caused by silver ions in the water phase Ag IW

Results – dose response test and acclimatization

Silver nanoparticles (PVP & citrate-coated) were less toxic than silver ions

The unexposed community (Ag naïve) was not significantly more sensitive to

silver than the previously exposed community

33

Conclusions

Low doses of Ag did not exert significant toxic effects in the short-term

Ionic form led to both functional and structural changes in the microbial

community (water phase)

Higher doses of Ag-NPs (>1 mg/L) significantly reduced microbial

community function in the case of citrate-coated Ag-NPs

PVP coated Ag-NPs were shown to have limited toxicity in this study

34

Synthesis

35

36

ENM containing

influent Effluent

TSS TSS

Synthesis – Removal of ENMs in constructed wetlands

• Silver nanoparticles are

efficiently retained by biofilm

• High affinity for organic

matter typical for ENMs in

general

37

ENM containing

influent Effluent

TSS TSS

Synthesis – Removal of ENMs in constructed wetlands

• Silver nanoparticles are

efficiently retained by biofilm

• High affinity for organic

matter typical for ENMs in

general

• Nanoparticles less toxic than

silver ions

• Effects on the microbial

community function and

structure only at high

concentrations (>500 µg/L)

38

ENM containing

influent Effluent

TSS TSS

Synthesis – Removal of Ag NPs in constructed wetlands

• Plant roots can immobilize

Ag NPs

• Translocation is negligible

• Silver nanoparticles are

efficiently retained by biofilm

• High affinity for organic

matter typical for ENMs in

general

• Nanoparticles less toxic than

silver ions

• Effects on the microbial

community function and

structure only at high

concentrations (>500 µg/L)

39

ENM containing

influent Effluent

TSS TSS

Synthesis – Removal of Ag NPs in constructed wetlands

• Plant roots can immobilize

Ag NPs

• Translocation is negligible

• Silver nanoparticles are

efficiently retained by biofilm

• High affinity for organic

matter typical for ENMs in

general

• Transformations, e.g.

• Sulfidation

• Adsorption

• Aggregation &

precipitation

• Dissolution

• Nanoparticles less toxic than

silver ions

• Effects on the microbial

community function and

structure only at high

concentrations (>500 µg/L)

40

Synthesis – Possible release of Ag NPs from constructed wetlands

• Sludge not usually

recovered from

constructed wetlands

• In special cases, the

concentrations of Ag

NPs in the sludge

layer may be high

40

ENM containing

influent Effluent

TSS TSS

41

Synthesis – Possible release of Ag NPs from constructed wetlands

• Low concentrations of Ag NPs in plant

tissues

• Not likely to impact composting of

harvested plant material • Sludge not usually

recovered from

constructed wetlands

• In special cases, the

concentrations of Ag

NPs in the sludge

layer may be high

41

ENM containing

influent Effluent

TSS TSS

42

Synthesis – Possible release of Ag NPs from constructed wetlands

• Low concentrations of Ag NPs in plant

tissues

• Not likely to impact composting of

harvested plant material

• Controlling the release of

solids (TSS) with the

effluent = a means to

control Ag NPs discharge

• Design, monitoring for

clogging, maintenance

• Sludge not usually

recovered from

constructed wetlands

• In special cases, the

concentrations of Ag

NPs in the sludge

layer may be high

42

ENM containing

influent Effluent

TSS TSS

Future perspectives

43

Future perspectives: Engineered nanomaterials

44

Accumulation of

nanoparticles over time:

Toxic effects in the long-

term?

TSS

Future perspectives: Engineered nanomaterials

45

Accumulation of

nanoparticles over time:

Toxic effects in the long-

term?

Release of ENMs controlled

by release of solids with

effluent:

Effect of malfunctions such

as clogging and short-

circuiting?

TSS

Thank you

46

Acknowledgements • Research Foundation Flanders (travel grant)

• European MSCA Joint Doctorate Programme on resource

recovery from wastewater - SuPER-W

www.superw.ugent.be (GA N° 676070)

• European COST Action ENTER (ES1205)

• Mark Button & Kela Weber

• Vincent Gagnon

• Baharak Hosseinkhani

• Frederik Van Koetsem

• V. Vasquez Sepulveda

Contact: Gijs.DuLaing@UGent.be

References • Auvinen H. , Kaegi R., Rousseau D., Du Laing G. 2017. Fate of silver

nanoparticles in constructed wetlands - a microcosm study. Water, Air and Soil

Pollution, 228, 97.

• Auvinen, H., Gagnon, V., Rousseau, D.P.L., Du Laing, G. (2017). Fate of

metallic engineered nanomaterials in constructed wetlands: prospection and

future research perspectives. Reviews in Environmental Science and

Bio/Technology, 16, 207-222.

• Auvinen H., Sepulveda Vasquez V., Rousseau D., Du Laing, G. 2016.

Substrate- and plant-mediated removal of citrate-coated silver nanoparticles in

constructed wetlands. Environmental Science and Pollution Research. 23(21):

21920-21926.

• Button, M., Auvinen, H., Van Koetsem, F., Hosseinkhani, B., Rousseau, D.,

Weber, K.P., Du Laing, G. (2016). Susceptibility of constructed wetland

microbial communities to silver nanoparticles: A microcosm study. Ecological

Engineering, 97, 476–485.

• Van Koetsem, F., Rinklebe, J., Du Laing, G. (2017). Analysis and Fate of Metal-

Based Engineered Nanoparticles in Aquatic Environments, Wetlands, and

Floodplain Soils. In : Rinklebe, J., Knox, A., Paller, M. (Eds.). Trace Elements in

Waterlogged Soils and Sediments, CRC Press, Taylor&Francis Group, Boca

Raton, US, pp. 101-134.