Zinc oxide

Toxicity to soil microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

Sludges were produced with ZnO ENPs, or their metal salts, added to both WWTP primary sludge and the activated sludge basin, followed by anaerobic digestion of primary clarifier solids and waste activated sludge. This resulted in three sludges: Control (SC) to which no metals were added, Metal salts (M) to which Zn(NO3)2 were added, and Nanoparticle (NP) to which ZnO ENP were added. The ZnO ENPs used had a primary particle size of 30 nm, but a hydrodynamic diameter in DI water of 253 nm. Zeta potential was þ15 mV at pH 7. The resulting wet sludges (approx. 25% dry weight) were air-dried and milled <2 mm. Soil was collected to a depth of 25 cm from a site inWoburn, Bedfordshire, UK, previously described by McGrath et al. (2012). The sandy soil (Arenosol, FAO) was sieved <2 mm and air-dried. Soil and sludges were mixed for 10 min using a Hobart bench mixer, leaving three bulk soil/sludge mixtures: SC, M and NP. Soil and sludge were mixed at a ratio of 58%:42%, targeting a total Zn concentration of 1400 mg/kg in both M and NP mixtures. The experiment was designed as a worst case scenario, based on the current U.S. EPA part 503 cumulative pollutant loading limit for Zn in soils amended with biosolids, combined with the guideline for soil/sewage sludge ratios of 1:1 in the top 15 cm of the soil and known concentrations in U.S. sewage
sludge (EPA, 1995)

GLP compliance:

not specified

Remarks:

GLP compliance not specified in the publication

Specific details on test material used for the study:

The primary particle size of the ZnO NPs was 30 ± 10 nm based on TEM measurements of 100 particles. The intensity average hydrodynamic diameter of the ZnO NPs in DI water at pH = 7 was 253 nm

Analytical monitoring:

yes

Vehicle:

no

Test organisms (inoculum):

soil

Total exposure duration:

176 d

Test temperature:

-9.8 to 13°C in January to 1.8 to 24.4 in June 2013

Nominal and measured concentrations:

1400mg/kg

Reference substance (positive control):

no

Duration:

176 d

Dose descriptor:

other: single dose effect concentration

Effect conc.:

1 400 mg/kg soil dw

Nominal / measured:

meas. (not specified)

Conc. based on:

element (total fraction)

Basis for effect:

nitrate formation rate

Remarks on result:

other: effect similar from zinc salt and ZnO nano

Reported statistics and error estimates:

All data were analysed using analysis of variance (ANOVA). Soil/sewage sludge Zn and Ag concentrations and total leachate amounts were square root transformed before analysis to establish homogeneity of variance. In cases where homogeneity could not be achieved by transformation, the non-parametric Mann-Whitney U-test was used (Genstat, VSN International, Hemel Hempstead, UK). PLFA and MSIR data were analysed using principal component analysis (PCA) followed by ANOVA of the factor scores to determine whether there were any significant effects of the experimental design on the PCA factor scores. PLFA and MSIR statistics were performed using Statsoft, Inc. (2012) STATISTICA version 11, stipulating an alpha value of 0.05.

Validity criteria fulfilled:

yes

Conclusions:

Study acceptable as weight of evidence.

Executive summary:

total leaching amounts were very low and there was no difference between ENP and salt forms of Zn and Ag. Inorganic N leaching indicated inhibition of nitrification by metals at the start of the experiment.

However, these microbial impacts were not accompanied by major changes in microbial community structure. Overall, leachate composition and soil microbial community responses were very similar in soils treated with sludges enriched with either ENPs or metal salts at the WWTP.

Reference 2

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)

Qualifier:

according to guideline

Guideline:

OECD Guideline 217 (Soil Microorganisms: Carbon Transformation Test)

GLP compliance:

not specified

Remarks:

GLP compliance not specified in publication

Specific details on test material used for the study:

Uncoated ZnO nanopowder (advertised particle size <100 nm diameter) and the ZnO bulk form were purchased from Sigma-Aldrich (Germany), and the ZnCl2 was from Panreac Spain).

Analytical monitoring:

yes

Vehicle:

no

Test organisms (inoculum):

soil

Total exposure duration:

28 d

Test temperature:

20±2 °C

Details on test conditions:

The Zn concentrations were always calculated on a dry soil (DW) basis. Untreated soil (natural Zn occurrence) was used as the control. The ZnONP and ZnO bulk powders were directly added to the soil and hand blended following previous studies (Hooper et al. 2011; Garcia-Gomez et al. 2014a). The powder application was selected because it was easier to make a homogenized mixing of the samples. The mixes were later sieved (2 mm) three times to ensure the homogenization of the samples. ZnCl2 was added to soil as an aqueous solution to guarantee that Zn2+ was the chemical form added to the soil. The volume of water added was equivalent to that required to achieve 50 % of the water holding capacity (WHC) of the soil. Control and treatment samples were wet to 50 % WHC and stored in the dark at 20±2 °C for 24 h before filling glass containers. Every container was filled with 100 g (DW) of control or treatment soil. Containers were incubated for 28 days (20±2 °C, dark, 40–60 % water holding capacity - WHC). The moisture of the samples was controlled by weight. pJ 7.48-8.29.

Nominal and measured concentrations:

Measured. Soil was contaminated with 1000 mg Zn kg−1 soil as ZnONPs, ZnO bulk, or ZnCl2.

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO nano

Basis for effect:

other: C transformation % inhibition of control

Remarks on result:

other: 20%

Remarks:

Not significantly different from ZnCl2

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO bulk

Basis for effect:

other: C transformation % inhibition of control

Remarks on result:

other: 20%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnCl2

Basis for effect:

other: C transformation % inhibition of control

Remarks on result:

other: 27%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO nano

Basis for effect:

nitrate formation rate

Remarks:

% inhibition of control

Remarks on result:

other: 0%

Remarks:

Significantly different from ZnCl2

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO bulk

Basis for effect:

nitrate formation rate

Remarks:

% inhibition of control

Remarks on result:

other: 0%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnCl2

Basis for effect:

nitrate formation rate

Remarks:

% inhibition of control

Remarks on result:

other: 35%

Remarks:

Significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO nano

Basis for effect:

other: DH activity

Remarks:

% inhibition of control

Remarks on result:

other: 52%

Remarks:

Not significantly different from ZnCl2

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO bulk

Basis for effect:

other: DH activity

Remarks:

% inhibition of control

Remarks on result:

other: 40%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnCl2

Basis for effect:

other: DH activity

Remarks:

% inhibition of control

Remarks on result:

other: 65%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO nano

Basis for effect:

other: PH activity

Remarks:

% inhibition of control

Remarks on result:

other: 33%

Remarks:

Not significantly different from ZnCl2

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnO bulk

Basis for effect:

other: PH activity

Remarks:

% inhibition of control

Remarks on result:

other: 28%

Remarks:

Not significantly different from ZnO-nano

Duration:

28 d

Dose descriptor:

other: single dose concentration

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

element (total fraction)

Remarks:

ZnCl2

Basis for effect:

other: PH activity

Remarks:

% inhibition of control

Remarks on result:

other: 30%

Remarks:

Not significantly different from ZnO-nano

Details on results:

The organic carbon mineralization in all soil treatments at both test times was reduced compared to the respective controls, although the differences were less than 30 %. The maximum effect was observed for ZnCl2, and there was no difference between ZnO-NPs and Zn bulk. Similarly, N transformation processes were only affected by ZnCl2, which inhibited N turnover by more than 30 % compared with the control. Both DH and PH (to a lesser extent) activities were significantly affected by the Zn treatments. DH activity was reduced immediately following the addition of any Zn chemical to the soil, and this reduction increased with the incubation period. Among treatments, there were not significant differences in DH activity between microorganisms exposed to ZnO-NPs and those exposed to both ZnO bulk or ZnCl2 after 28 days; however, the responses of the later (ZnO bulk and ZnCl2) were statistically different from each other (p<0.01). The PH activity at day 1 was not affected by any ZnO form (NP or bulk), but it was reduced by the Zn ion. After 28 days, all treatments inhibited PH activities by a similar amount.

Reported statistics and error estimates:

The data were analyzed statistically using the STAT GRAPHICS software (Version 5.0). Statistically significant differences between individual means for chemical and toxicological data were identified by one-way analysis of variance (ANOVA) with Fisher’s least significant difference procedure (LSD, p<0.05).

Total Zn uptake was calculated as the product of dry matter production and the total Zn concentration in roots or shoots. The transference factor (Tf) was defined as the ratio of the Zn concentration in shoots to that in roots. The bioconcentration factor (BCF) was calculated as the ratio of the Zn concentration measured in roots or shoots to that of the Zn in soil. Zinc concentrations are reported on a DW basis.

Validity criteria fulfilled:

yes

Conclusions:

Good quality study useful for assessing terrestrial toxicity.

Executive summary:

Good study, followed OECD guidelines. ZnCl2 generally presented the highest toxicity (greatest level of inhibition); ZnO-nano and ZnO bulk effects were similar.

Description of key information

The toxicity of the nano-ZnO form is in general lower than the toxicity of the zinc ion. Ingestion of solids is a potential risk pathway of greater importance for organisms in soils/sediments than in aquatic systems – potentially a greater chance for exposure to dispersed/aggregated MNPs, and nano particles can be more stable due to e.g. coating or given environmental conditions. However, there is no indication of toxicity deviating from the zinc-ion in the present dataset.

Additional information

There are data on the effect of nano-ZnO (compared to zinc ion toxicity) available for soil invertebrates (4 species), plants (10 species) and soil microorganisms. These data are summarized in the attached table in the attached background material section, where the effect concentrations observed for the Zn2+form are normalized (100%).

From these data, it follows that the toxicity of the nano-ZnO form is in general lower than the toxicity of the zinc ion. Ingestion of solids is a potential risk pathway of greater importance for organisms in soils/sediments than in aquatic systems – potentially a greater chance for exposure to dispersed/aggregated MNPs, and nano particles can be more stable due to e.g. coating or given environmental conditions. However, there is no indication of toxicity deviating from the zinc-ion in the present dataset.