Disilver oxide

Toxicity to terrestrial plants

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to terrestrial plants: short-term

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study, available as unpublished report. Some limitations in method and/or reporting but otherwise adequate for assessment.

Qualifier:

according to guideline

Guideline:

other: International Standard ISO 11269-1 (ISO, 1993)

GLP compliance:

not specified

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the completion of the toxicity test, all of the remaining soil was dried at 40°C, homogenised and sieved to 2 mm for chemical analysis.

Vehicle:

no

Details on preparation and application of test substrate:

APPLICATION OF TEST SUBSTANCE TO SOIL
The soils were spiked with Ag by adding a concentrated AgNO3 solution to the soil samples at a rate of 50 mL of solution per kg dry soil. This addition was done using a fine spray onto the soils, followed by thorough mixing of the sample, to ensure that there was a homogeneous distribution of the solution throughout the soil. All of the soil spiking was conducted in duplicate to allow for a leached and unleached treatment.

LEACHING PROCEDURE:
The soils were left to equilibrate overnight prior to the commencement of soil leaching. The soils were leached with an artificial rain water (ARW) solution. Some modifications were made to the solution to remove Cl- salts to prevent precipitation of AgCl. On the initial day of leaching the spiked soils were transferred to containers with holes in the base, lined with mesh or filter paper. The containers were then submerged in a larger container that contained the ARW solution (ratio 800 mL of ARW : 1 kg soil), and left overnight for the soils to reach saturation. The following day, the soils were removed from the ARW solution and additional ARW was added to the surface of the soils at a rate equivalent to 600 mL ARW per 1 kg soil and allowed to drain through the base of the container overnight. The following day all soils (leached and unleached) were transferred to trays to allow them to air dry. The unleached soils were retained at room temperature while leaching of the other samples was being conducted. Approximately 10 days was allowed for sample drying. The dried samples were then homogenised and sieved to < 2 mm for the commencement of the toxicity test.

Species:

Hordeum vulgare

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: barley
- Plant family: Poaceae

Test type:

other: inhibition of root growth

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

5 d

Post exposure observation period:

None.

Test temperature:

The temperature was maintained at 23°C (±3°C) during the day and 17°C (±3°C) at night.

pH:

pH values for the soils used (in CaCl2):
Houthalen 3.6
Bordeaux 4.6
Inman Valley 5.0
Charleston 5.1
Kingaroy 5.5
Millicent 6.6
Balaklava 7.1
Port Kenny 8.0

Moisture:

70%

Details on test conditions:

TEST SYSTEM
- Testing facility: plant growth cabinet
- Test container (type, material, size): pot
- Amount of soil: 250 g
- Method of seeding: seeds were placed approximately 1 cm under the soil surface in each pot
- No. of seeds per container: 6
- No. of replicates per treatment group: 3
- No. of replicates per control: 3

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: eight study soils used, two of the soils were collected from Europe, Houthalen (Belgium) and Bordeaux (France) and the remaining six soils (Inman Valley, Charleston, Kingaroy, Millicent, Balaklava and Port Kenny) were collected from locations around Australia.
- Collection procedures: not reported
- Soil texture (if natural soil): Full details provided in Table 1.
- % sand: 19 - 93%
- % silt: 1.6 - 22%
- % clay: 1.4 - 60%
- Organic carbon: 0.9 - 12%
- Maximum water holding capacity (in % dry weigth): no data
- CEC: 5.3 - 42 cmol+/kg

GROWTH CONDITIONS
- Photoperiod: 16 hours of light and 8 hours of dark
- Light source: no data
- Light intensity and quality: no data
- Day/night temperatures: 23°C (±3°C) during the day and 17°C (±3°C) at night.
- Relative humidity (%): 70% of field capacity (FC)
- Watering regime and schedules: the water content of the soils was maintained by weight
- Water source/type: ultrapure water

ACCLIMATION PERIOD: no

EFFECT PARAMETERS MEASURED: The length of the plant roots and shoots were recorded at the completion of the test.

VEHICLE CONTROL PERFORMED: no

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.0
- Test concentrations:
The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600

Nominal and measured concentrations:

The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600
There was close agreement between the measured and nominal concentrations which ranged from 80 – 117% of the expected values with an overall average of 95%. Based on these results, the nominal concentrations were used for all data analysis in all cases, except the Houthalen leached data where the measured concentrations were used.

Reference substance (positive control):

no

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

13 mg/kg soil dw

95% CI:

>= 10 - <= 16

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Houthalen soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

20 mg/kg soil dw

95% CI:

>= 15 - <= 25

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Bordeaux soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

146 mg/kg soil dw

95% CI:

>= 93 - <= 210

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Inman Valley soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

60 mg/kg soil dw

95% CI:

>= 50 - <= 71

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Charleston soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

25 mg/kg soil dw

95% CI:

>= 18 - <= 31

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Kingaroy soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

176 mg/kg soil dw

95% CI:

>= 143 - <= 214

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Millicent soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

61 mg/kg soil dw

95% CI:

>= 40 - <= 84

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Balaklava soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

25 mg/kg soil dw

95% CI:

>= 18 - <= 31

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Port Kenny soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

17 mg/kg soil dw

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Houthalen soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

20 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Bordeaux soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

249 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Inman Valley soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

157 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Charleston soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

215 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Millicent soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

47 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Balaklava soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

72 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: root growth

Remarks on result:

other: Port Kenny soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

109 mg/kg soil dw

95% CI:

>= 68 - <= 162

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Houthalen soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

130 mg/kg soil dw

95% CI:

>= 93 - <= 165

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Bordeaux soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

131 mg/kg soil dw

95% CI:

>= 110 - <= 156

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Inman Valley, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

62 mg/kg soil dw

95% CI:

>= 39 - <= 90

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Charleston soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

301 mg/kg soil dw

95% CI:

>= 220 - <= 394

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Millicent soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

215 mg/kg soil dw

95% CI:

>= 183 - <= 251

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Balaklava soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

45 mg/kg soil dw

95% CI:

>= 35 - <= 56

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot growth

Remarks on result:

other: Port Kenny soil, unleached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

101 mg/kg soil dw

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Houthalen soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

140 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Bordeaux soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

99 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Inman Valley, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

60 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Charleston soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

422 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Millicent soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

232 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot lenght

Remarks on result:

other: Balaklava soil, leached

Species:

Hordeum vulgare

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

183 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: shoot growth

Remarks on result:

other: Port Kenny soil, leached

Details on results:

SEED GERMINATION
- Percent seed germination: There was 100% germination observed in all of the control soils.
- Root length: The root length in the control soils varied across the different soils, however, was sufficient for all subsequent toxicity analysis. In most cases there was no significant difference in root length between the leached and unleached treatments with exceptions in the Houthalen and Charleston soils.
- Root discolouration/malformation: not reported

Reported statistics and error estimates:

No significant increase (p > 0.05) in the measured response at low Ag concentrations, a standard dose log-logistic model was used to fit the data and derive ECx values:
y=c+ (d-c)/(1+(x/e)^b )

The EC10 and EC50 values were then determined in each case through interpolation from the fitted curve at a 10% and 50% reduction from the fitted d values (i.e. fitted response in the control). EC10 values outside the concentration range tested, i.e., below or above the lowest or highest dose tested, respectively, were considered unreliable due to their large degree of uncertainty (and are thus not added here).

Table 1: Measured concentrations of silver (Ag) in soils used for the plant toxicity tests (inhibition of root growth). Values in parentheses represent the measured concentration as a percentage of the expected nominal concentration

Soil	Nominal concentration (mg/kg)	Average measured concentration (mg/kg)
		Leached	Unleached
Houthalen	5	4.3 (87)	4.3 (85)
	10	8.8 (87)	8.5 (85)
	20	17.0 (83)	18 (90)
	40	32 (81)	37 (92)
	80	62 (77)	80 (100)
	160	107 (67)	163 (102)
	320	185 (58)	320 (100)
Bordeaux	5	4.7 (94)	5.3 (105)
	10	9.4 (94)	9.9 (99)
	20	19 (97)	19 (94)
	40	41 (102)	38 (94)
	80	78 (97)	85 (107)
	160	144 (90)	180 (113)
	320	298 (93)	338 (106)
Inman Valley	25	23 (93)	20 (80)
	50	45 (91)	44 (88)
	100	92 (92)	93 (93)
	200	185 (92)	201 (101)
	400	374 (93)	370 (93)
	800	745 (93)	775 (97)
	1600	1430 (89)	1500 (94)
Charleston	25	23 (91)	24 (97)
	50	47 (93)	43 (86)
	100	80 (80)	82 (82)
	200	161 (80)	169 (85)
	400	368 (92)	364 (91)
	800	758 (95)	664 (83)
	1600	1560 (98)	1540 (96)
Kingaroy	25	26 (106)	26 (102)
	50	53 (106)	53 (106)
	100	97 (97)	86 (86)
	200	198 (99)	211 (106)
Kingaroy	400	396 (99)	398 (100)
	800	800 (100)	813 (102)
	1600	1610 (101)	1620 (101)
Millicent	50	43 (86)	45 (90)
	100	95 (95)	91 (91)
	200	180 (90)	189 (95)
	400	383 (96)	387 (97)
	800	805 (101)	738 (92)
	1600	1460 (91)	1650 (103)
	3200	2900 (90)	3170 (99)
Balaklava	25	23 (93)	23 (91)
	50	45 (90)	44.9 (90)
	100	88 (88)	89.6 (90)
	200	184 (92)	186 (93)
	400	384 (96)	387 (97)
	800	741 (93)	779 (97)
	1600	1530 (96)	1500 (94)
Port Kenny	25	27 (109)	29.3 (117)
	50	52 (104)	52 (105)
	100	80 (80)	103 (103)
	200	194 (97)	204 (102)
	400	342 (86)	373 (93)
	800	760 (95)	760 (95)
	1600	1400 (88)	1420 (89)

Validity criteria fulfilled:

yes

Remarks:

100% germination was observed in all of the control soils

Conclusions:

The most sensitive endpoint in the study was root length. The EC10 values based on this endpoint ranged from 13 (Houthalen soil) to 249 (Inman Valley soil) mg Ag/kg.

Executive summary:

The toxicity of Ag to barley (Hordeum vulgare), was tested according to ISO guideline 11269-1. For the inhibition of root growth test, both root and shoot length were measured as endpoints. In most cases the root length data were found to provide the most sensitive endpoint when deriving EC10 and EC50 values. Overall the EC10 values for root length were found to range from 13 (Houthalen soil) to 249 (Inman Valley soil) mg Ag/kg. The EC10 based on shoot length were between 2.3 mg Ag/kg (Kingaroy soil) and 422 mg Ag/kg (Millicent). All EC10 values were taken forward to the derivation of the PNEC according to the statistical extrapolation approach. Given the negligible effects of leaching on Ag toxicity to soil organisms, both results for leached and unleached soil treatments were used.

Reference 2

Endpoint:

toxicity to terrestrial plants: short-term

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study, available as unpublished report. Some limitations in method and/or reporting but otherwise adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 208 (Terrestrial Plants Test: Seedling Emergence and Seedling Growth Test)

Qualifier:

according to guideline

Guideline:

other: International Standard ISO 11269-2 (ISO, 1995)

GLP compliance:

not specified

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the completion of the toxicity test, all of the remaining soil was dried at 40°C, homogenised and sieved to 2 mm for chemical analysis.

Vehicle:

no

Details on preparation and application of test substrate:

APPLICATION OF TEST SUBSTANCE TO SOIL:
The soils were spiked with Ag by adding a concentrated AgNO3 solution to the soil samples at a rate of 50 mL of solution per kg dry soil. This addition was done using a fine spray onto the soils, followed by thorough mixing of the sample, to ensure that there was a homogeneous distribution of the solution throughout the soil. Only leached treatment was tested.

AGEING PROCEDURE:
Following spiking and leaching, the soils were dried and homogenised and separated into three replicates prior to rehydration with ultrapure water to 50% of FC. The samples were then retained in the dark at a constant temperature of 22°C for a 12 month period. The moisture content of the soils was maintained by weight and samples were aerated on a regular basis. At the completion of the 12 month ageing period the moisture content of the samples was increased to 70% FC (to be consistent with the previous plant toxicity testing) and the seedling emergence and early growth test was carried out.

LEACHING PROCEDURE:
The soils were left to equilibrate overnight prior to the commencement of soil leaching. The soils were leached with an artificial rain water (ARW) solution. Some modifications were made to the solution to remove Cl- salts to prevent precipitation of AgCl. On the initial day of leaching the spiked soils were transferred to containers with holes in the base, lined with mesh or filter paper. The containers were then submerged in a larger container that contained the ARW solution (ratio 800 mL of ARW : 1 kg soil), and left overnight for the soils to reach saturation. The following day, the soils were removed from the ARW solution and additional ARW was added to the surface of the soils at a rate equivalent to 600 mL ARW per 1 kg soil and allowed to drain through the base of the container overnight. The following day all soils (leached and unleached) were transferred to trays to allow them to air dry. The unleached soils were retained at room temperature while leaching of the other samples was being conducted. Approximately 10 days was allowed for sample drying. The dried samples were then homogenised and sieved to < 2 mm for the commencement of the toxicity test.

Species:

Lycopersicon esculentum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: tomato
- Plant family: Solanaceae

Test type:

seedling emergence toxicity test

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

21 d

Remarks:

The test was terminated three weeks after 50% emergence was observed in the controls.

Post exposure observation period:

None.

Test temperature:

The test temperature was maintained at 23°C (±3°C) during the day and at 17°C (±3°C) at night.

pH:

pH values for the soils used (in CaCl2):
Houthalen 3.6
Bordeaux 4.6
Inman Valley 5.0
Charleston 5.1
Kingaroy 5.5
Millicent 6.6
Balaklava 7.1
Port Kenny* 8.0

Moisture:

Each replicate was hydrated to 70% of field capacity (FC) with ultrapure water.

Details on test conditions:

TEST SYSTEM
- Testing facility: plant growth cabinet
- Test container (type, material, size): pot. The soil surface was covered with alkathene granules to limit evaporation of water from the pots.
- Amount of soil: 500 g
- Method of seeding: tomato seeds (not germinated) were planted in each replicate pot
- No. of seeds per container: 10
- No. of replicates per treatment group: 1
- No. of replicates per control: 1

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: eight study soils used, two of the soils were collected from Europe, Houthalen (Belgium) and Bordeaux (France) and the remaining six soils (Inman Valley, Charleston, Kingaroy, South East, Balaklava and Port Kenny) were collected from locations around Australia.
- Collection procedures: not reported
- Soil texture (if natural soil): Full details provided in Table 1.
- % sand: 19 - 93%
- % silt: 1.6 - 22%
- % clay: 1.4 - 60%
- Organic carbon: 0.9 - 12%
- Maximum water holding capacity (in % dry weigth): no data
- CEC: 5.3 - 42 cmol+/kg

NUTRIENT MEDIUM (if used)
- Description: Fertiliser pellets (Nitrophoska®) were added to each of the pots at a rate of 0.5 g per pot.

GROWTH CONDITIONS
- Photoperiod: 16 hours light and 8 hours dark
- Light source: not reported
- Light intensity and quality: not reported
- Day/night temperatures: 23°C (±3°C) (day) and 17°C (±3°C) (night
- Watering regime and schedules: Each replicate was hydrated to 70% of field capacity (FC) with ultrapure water prior to start of the test.
- Water source/type: ultrapure water

ACCLIMATION PERIOD: no

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : emergence, plant height and plant biomass (dry weight).

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.0
- Range finding study: no
- Test concentrations:
The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600

Nominal and measured concentrations:

The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600
The measured concentrations of Ag in all of the aged soils were in close agreement with the nominal concentrations (measured values averaged 94% of the expected nominal concentrations), with the exception of Houthalen, due to loss through the leaching process. Overall there were no significant differences observed between the 1 month and 12 month measured concentrations of Ag. Based on this, all data analysis was done using nominal Ag concentrations, with the exception of the Houthalen soil where measured concentrations were used.

Reference substance (positive control):

no

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

116 mg/kg soil dw

95% CI:

>= 41 - <= 217

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Bordeaux soil, aged 12 months

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

110 mg/kg soil dw

95% CI:

>= 51 - <= 227

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Millicent soil, aged 12 months

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

180 mg/kg soil dw

95% CI:

>= 134 - <= 234

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Bakalava soil, aged 12 months

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

121 mg/kg soil dw

95% CI:

>= 74 - <= 192

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Port Kenny soil, aged 12 months

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

60 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Charleston soil, aged 12 months

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

253 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Port Kenny soil, aged 12 months

Details on results:

The growth of plants in the control soils was similar in the test conducted 1 month post spiking and the test conducted in the soils that had been aged for 12 months. Some significant differences were however observed in the Inman Valley, Charleston and Balaklava soils.

In all soils clear dose response relationships were evident in the soils that had been aged 12 months. When the dose response curves for the 12 months aged soils are compared with the 1 month aged soils it is clear in all cases that there is a shift to right, indicating a decrease in the toxicity of Ag in the aged soils.
The effect of ageing on the toxicity of Ag is also evident when the EC10 and EC50 values are compared. In all cases, the ECx values are either significantly lower in the 12 months aged soils or show no significant difference. The AFs that were determined from these values were found to range from 1.7 to 22 for the EC10 values and 1.3 to 2.8 for the EC50 values. To calculate an overall average AF, the Bordeaux EC10 AF was removed as this value was considerably higher than the other values. Following the removal of this value, the average overall AF in these soils was 2.4, indicating that after a 12-month ageing period the toxicity of Ag decreased twofold.

Reported statistics and error estimates:

EC10 values outside the concentration range tested, i.e., below or above the lowest or highest dose tested, respectively, were considered unreliable due to their large degree of uncertainty (and are thus not added here).

Table1: Results from analyses of the selected study soils for pH, organic carbon, cation exchange capacity, particle size distribution and total silver concentration.

Soil	pH (CaCl2)	pH (water)	Organic carbon (%)	Cation exchange capacity (cmol^+/kg)	Particle size distribution (%)			Total Silver (mg/kg)
					clay	silt	Sand
Houthalen	3.6	4.9	1.5	5.3	1.4	1.7	93	< 0.04
Bordeaux	4.6	5.6	1.9	6.4	2.5	1.6	93	< 0.04
Inman Valley	5.0	6.0	5.3	25	42	22	26	< 0.04
Charleston	5.1	6.6	6.9	12	14	12	63	< 0.04
Kingaroy	5.5	6.1	0.9	13	60	17	19	< 0.04
Millicent	6.6	6.9	12	42	19	5.2	48	< 0.04
Balaklava	7.1	8.5	1.9	27	30	20	47	< 0.04
Port Kenny*	8.0	8.8	1.8	13	12	4.2	21	< 0.04

* note that particle size distribution for Port Kenny is equal to 37.2% due to the high concentration of CaCO₃that is present in this soil (60%)

 

Table 2: Silver (Ag) concentrations corresponding to a 10% and 50% reduction in plant biomass (EC10 and EC50, respectively) in soils aged 12 months and 1 month. All concentrations are shown as mg Ag/kg and values in parentheses represent the 95% confidence intervals.

SOIL	EC10				EC50
	12 month	1 month	AF		12 month	1 month	AF
Houthalen	1.0 (na-3.2)	0.47 (na-1.2)	2.1		16 (9.8-23)	8.5 (5.7-11)	1.9
Bordeaux	8.1 (na-28)	5.3 (na-9.2)	1.5		77 (37-167)	19* (14-25)	4.1
Inman Valley	11 (na-32)	6.9 (na-18)	1.6		85 (48-124)	50 (31-72)	1.7
Charleston	60 (43-74)	7.0* (na-16)	8.6		123 (110-137)	50* (34-68)	2.5
Kingaroy	22 (na-43)	1.1 (0.24-4.0)	20		58 (35-80)	12* (5.6-18)	4.8
Millicent	41 (na-125)	34 (na-61)	1.2		317 (171-535)	139 (104-184)	2.3
Balaklava	17 (na-43)	45 (41-50)	0.4		121 (76-176)	83 (76-91)	1.5
Port Kenny	253 (203-305)	108* (70-156)	2.3		371 (310-440)	282 (173-585)	1.3

* indicate significantly (p < 0.05) lower values when the leached and unleached ECx values are compared

AF: ageing factor calculated as 12 month ECx/1 month ECx

na, confidence interval(s) not available due to large variability in data

Effect of soil ageing on plant uptake of silver

The BAFs that were calculated from the Ag concentrations in the above ground plant tissue showed a general decreasing trend when the 1 month results were compared to the 12 month results. Due to the high variability in the data, significantly lower BAFs were only evident in the Bordeaux and Port Kenny soils. In contrast, at one Ag rate in the Houthalen and two rates in the Kingaroy soils, the opposite trend was significant, whereby a significantly lower BAF was observed in the 1 month samples compared to the 12 month samples. Overall, there was an average 15% reduction in BAF values of the 12 month aged samples compared to the 1 month aged samples. The overall decreasing trend in the BAFs values indicates that there is a decrease in the uptake of Ag into the plants with time, which is also consistent with the decreasing toxicity that was observed as evidenced by the AFs greater than 1.

Validity criteria fulfilled:

not specified

Conclusions:

The EC10 and EC50 values were found in all cases to increase in soils that had been aged for 12 months, compared to those generated in soils that had been aged for 1 month. Overall, the ageing factors (AFs) were found to range from 1.3 to 4.4 (and additional value of 22 was observed for one soil) with an overall average AF of 2.4.

Executive summary:

An affect of soil ageing on the toxicity of silver, using tomatoes (Lycopersicum esculentum) as the test species was tested using soils spiked with silver that were subsequently allowed to age for either one or 12 months. In all cases, the EC10 and EC50 values were significantly lower or there was no significant difference compared to those generated in soils that had been aged for one month. EC10 values in soils aged for 12 months were between 60 (Charleston soil) and 253 mg Ag/kg (Port Kenny soil) for the endpoin biomass. For plant height, the EC10 values ranged from 110 (Millicent soil) to 180 mg Ag/kg (Balaklava soil) in 12 months aged soils. All EC10 values were taken forward to the derivation of the PNEC according to the statistical extrapolation approach.

Reference 3

Endpoint:

toxicity to terrestrial plants: short-term

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study, available as unpublished report. Some limitations in method and/or reporting but otherwise adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 208 (Terrestrial Plants Test: Seedling Emergence and Seedling Growth Test)

Qualifier:

according to guideline

Guideline:

other: International Standard ISO 11269-2 (ISO, 1995)

GLP compliance:

not specified

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the completion of the toxicity test, all of the remaining soil was dried at 40°C, homogenised and sieved to 2 mm for chemical analysis.

Vehicle:

no

Details on preparation and application of test substrate:

APPLICATION OF TEST SUBSTANCE TO SOIL
The soils were spiked with Ag by adding a concentrated AgNO3 solution to the soil samples at a rate of 50 mL of solution per kg dry soil by using a fine spray onto the soils, followed by thorough mixing of the sample, to ensure that there was a homogeneous distribution of the solution throughout the soil. All of the soil spiking was conducted in duplicate to allow for a leached and unleached treatment.

LEACHING PROCEDURE:
The soils were left to equilibrate overnight prior to the commencement of soil leaching. The soils were leached with an artificial rain water (ARW) solution. Some modifications were made to the solution to remove Cl- salts to prevent precipitation of AgCl. On the initial day of leaching the spiked soils were transferred to containers with holes in the base, lined with mesh or filter paper. The containers were then submerged in a larger container that contained the ARW solution (ratio 800 mL of ARW : 1 kg soil), and left overnight for the soils to reach saturation. The following day, the soils were removed from the ARW solution and additional ARW was added to the surface of the soils at a rate equivalent to 600 mL ARW per 1 kg soil and allowed to drain through the base of the container overnight. The following day all soils (leached and unleached) were transferred to trays to allow them to air dry. The unleached soils were retained at room temperature while leaching of the other samples was being conducted. Approximately 10 days was allowed for sample drying. The dried samples were then homogenised and sieved to < 2 mm for the commencement of the toxicity test.

Species:

Lycopersicon esculentum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: tomato
- Plant family: Solanaceae

Test type:

seedling emergence toxicity test

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

21 d

Remarks:

The test was terminated three weeks after 50% emergence was observed in the controls.

Post exposure observation period:

None.

Test temperature:

The temperature was maintained at 23°C (±3°C) (day) and at 17°C (±3°C) (night).

pH:

pH values for the soils used (in CaCl2):
Houthalen 3.6
Bordeaux 4.6
Inman Valley 5.0
Charleston 5.1
Kingaroy 5.5
Millicent 6.6
Balaklava 7.1
Port Kenny 8.0

Moisture:

The soils were hydrated to 70% of FC.

Details on test conditions:

TEST SYSTEM
- Testing facility: plant growth cabinet
- Test container (type, material, size): pot. The soil surface was covered with alkathene granules to limit evaporation of water from the pots.
- Amount of soil: 500 g
- Method of seeding: tomato seeds (not germinated) were planted in each replicate pot
- No. of seeds per container: 10
- No. of plants (retained after thinning):
- No. of replicates per treatment group: 1
- No. of replicates per control: 1

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: eight study soils used, two of the soils were collected from Europe, Houthalen (Belgium) and Bordeaux (France) and the remaining six soils (Inman Valley, Charleston, Kingaroy, South East, Balaklava and Port Kenny) were collected from locations around Australia.
- Collection procedures: not reported
- Soil texture (if natural soil): Full details provided in Table 1.
- % sand: 19 - 93%
- % silt: 1.6 - 22%
- % clay: 1.4 - 60%
- Organic carbon: 0.9 - 12%
- Maximum water holding capacity (in % dry weigth): no data
- CEC: 5.3 - 42 cmol+/kg

NUTRIENT MEDIUM (if used)
- Description: Fertiliser pellets (Nitrophoska®) were added to each of the pots at a rate of 0.5 g per pot.

GROWTH CONDITIONS
- Photoperiod: 16 hours light and 8 hours dark
- Light source: not reported
- Light intensity and quality: not reported
- Day/night temperatures: 23°C (±3°C) (day) and 17°C (±3°C) (night
- Watering regime and schedules: Each replicate was hydrated to 70% of field capacity (FC) with ultrapure water prior to start of the test.
- Water source/type: ultrapure water

ACCLIMATION PERIOD: no

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : emergence, plant height and plant biomass (dry weight).

VEHICLE CONTROL PERFORMED: no

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.0
- Range finding study: no
- Test concentrations:
The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600

Nominal and measured concentrations:

The nominal Ag concentrations used for this toxicity testing were as follows (all concentrations are shown as mg Ag/kg):
Houthalen 0, 5, 10, 20, 40, 80, 160, 320
Bordeaux 0, 5, 10, 20, 40, 80, 160, 320
Inman Valley 0, 25, 50, 100, 200, 400, 800, 1600
Charleston 0, 25, 50, 100, 200, 400, 800, 1600
Kingaroy 0, 25, 50, 100, 200, 400, 800, 1600
Millicent 0, 50, 100, 200, 400, 800, 1600, 3200
Balaklava 0, 25, 50, 100, 200, 400, 800, 1600
Port Kenny 0, 25, 50, 100, 200, 400, 800, 1600
There was close agreement between the measured and nominal concentrations which ranged from 80 – 117% of the expected values with an overall average of 95%. Based on these results, the nominal concentrations were used for all data analysis in all cases, except the Houthalen leached data where the measured concentrations were used.

Reference substance (positive control):

no

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

8.7 mg/kg soil dw

95% CI:

<= 18

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Houthalen soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

17 mg/kg soil dw

95% CI:

>= 7.9 - <= 27

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Bordeaux soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

54 mg/kg soil dw

95% CI:

>= 34 - <= 74

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Charleston soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

42 mg/kg soil dw

95% CI:

>= 26 - <= 57

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Balaklava soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

53 mg/kg soil dw

95% CI:

<= 98

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Port Kenny soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

5.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Bordeaux soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

28 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Charleston soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

62 mg/kg soil dw

Nominal / measured:

meas. (not specified)

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Millicent soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

52 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Balaklava soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

61 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: plant height

Remarks on result:

other: Port Kenny soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

58 mg/kg soil dw

95% CI:

<= 117

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Port Kenny soil, unleached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

5.3 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Bordeaux soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

45 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Balaklava soil, leached

Species:

Lycopersicon esculentum

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

108 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: biomass

Remarks on result:

other: Port Kenny soil, leached

Details on results:

The growth of the plants was sufficient in all of the control soils to allow for all subsequent toxicity modelling.

Reported statistics and error estimates:

There was no significant increase (p > 0.05) in the measured response at low Ag concentrations, therefore a standard dose log-logistic model was used to fit the data and derive ECx values.
y=c+ (d-c)/(1+(x/e)^b )
EC10 values outside the concentration range tested, i.e., below or above the lowest or highest dose tested, respectively, were considered unreliable due to their large degree of uncertainty (and are thus not added here).

TableA.2: Measured concentrations of silver (Ag) in soils used for the plant toxicity tests (inhibition of root growth and seedling emergence and early growth). Values in parentheses represent the measured concentration as a percentage of the expected nominal concentration

Soil	Nominal concentration (mg/kg)	Average measured concentration (mg/kg)
		Leached	Unleached
Houthalen	5	4.3 (87)	4.3 (85)
	10	8.8 (87)	8.5 (85)
	20	17.0 (83)	18 (90)
	40	32 (81)	37 (92)
	80	62 (77)	80 (100)
	160	107 (67)	163 (102)
	320	185 (58)	320 (100)
Bordeaux	5	4.7 (94)	5.3 (105)
	10	9.4 (94)	9.9 (99)
	20	19 (97)	19 (94)
	40	41 (102)	38 (94)
	80	78 (97)	85 (107)
	160	144 (90)	180 (113)
	320	298 (93)	338 (106)
Inman Valley	25	23 (93)	20 (80)
	50	45 (91)	44 (88)
	100	92 (92)	93 (93)
	200	185 (92)	201 (101)
	400	374 (93)	370 (93)
	800	745 (93)	775 (97)
	1600	1430 (89)	1500 (94)
Charleston	25	23 (91)	24 (97)
	50	47 (93)	43 (86)
	100	80 (80)	82 (82)
	200	161 (80)	169 (85)
	400	368 (92)	364 (91)
	800	758 (95)	664 (83)
	1600	1560 (98)	1540 (96)
Kingaroy	25	26 (106)	26 (102)
	50	53 (106)	53 (106)
	100	97 (97)	86 (86)
	200	198 (99)	211 (106)
Kingaroy	400	396 (99)	398 (100)
	800	800 (100)	813 (102)
	1600	1610 (101)	1620 (101)
Millicent	50	43 (86)	45 (90)
	100	95 (95)	91 (91)
	200	180 (90)	189 (95)
	400	383 (96)	387 (97)
	800	805 (101)	738 (92)
	1600	1460 (91)	1650 (103)
	3200	2900 (90)	3170 (99)
Balaklava	25	23 (93)	23 (91)
	50	45 (90)	44.9 (90)
	100	88 (88)	89.6 (90)
	200	184 (92)	186 (93)
	400	384 (96)	387 (97)
	800	741 (93)	779 (97)
	1600	1530 (96)	1500 (94)
Port Kenny	25	27 (109)	29.3 (117)
	50	52 (104)	52 (105)
	100	80 (80)	103 (103)
	200	194 (97)	204 (102)
	400	342 (86)	373 (93)
	800	760 (95)	760 (95)
	1600	1400 (88)	1420 (89)

Validity criteria fulfilled:

yes

Conclusions:

The most sensitive endpoint in the study was shoot biomass. The EC10 values based on this endpoint ranged from 5.3 (Bordeaux soil) to 108 (Port Kenny soil) mg Ag/kg. The toxicity of Ag to tomato appeared to be controlled by soil pH and organic carbon.

Executive summary:

The toxicity of Ag to plants was tested according to OECD guideline 208 using tomatoes (Lycopersicum esculentum) as the test species. The tests were conducted in eight different soils representing a wide range of soil properties (pH 3.6 – 8.0, organic carbon 0.9 – 12% and clay 1.4 – 60%). Results from leached and unleached soil treatments are reported. The endpoints measured in the seedling emergence and early growth test included emergence, plant height and plant biomass (dry weight). Plant emergence showed very low sensitivity, therefore, the results are not presented. The plant growth endpoints of height and biomass both showed high sensitivity to additions of Ag to the soil. Plant biomass showed the highest sensitivity, however, the results were considerably more variable than the plant height data. The toxicity of Ag to tomato appeared to be controlled by soil pH and organic carbon. The EC10 values based on tomatoes weight were found to range from 5.3 (Bordeaux soil) to 108 (Port Kenny soil) mg Ag/kg. All EC10 values were taken forward to the derivation of the PNEC according to the statistical extrapolation approach. Given the negligible effects of leaching on Ag toxicity to soil organisms, both results for leached and unleached soil treatments were used.

Reference 4

Endpoint:

toxicity to terrestrial plants: short-term

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP-compliant, non-guideline study, published in peer-review literature. Limitations in design and/or reporting but otherwise adequate for assessment.

Principles of method if other than guideline:

Seeds of Phaseolus radiatus (mung bean) and Sorghum bicolor (sorghum) were germinated and then exposed to silver nitrate in artificial soil. The growth of the shoots and roots after 5 days of exposure were measured to the nearest millimeter. Silver nanoparticle tests were run in parallel.

GLP compliance:

no

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Not applicable

Analytical monitoring:

yes

Details on sampling:

The silver released during exposure was measured by introducing 40 % moisture to the dry soil media containing silver nitrate and keeping the media under standard conditions for 5 days. The samples were then filtered using Whatman No 2 filters and 0.2 µm nylon membranes and the filtrate was analysed for silver ions.

Vehicle:

no

Details on preparation and application of test substrate:

The test substrate was OECD artificial soil.

Species:

other: Phaseolus radiatus

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Purchased from a commercial seed company in Seoul, Korea and sterilised by soaking for 5-10 minutes in 5 % sodium hypochlorite solution before being rinsed thoroughly in deionised water.

Species:

Sorghum bicolor

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

Purchased from a commercial seed company in Seoul, Korea and sterilised by soaking for 5-10 minutes in 5 % sodium hypochlorite solution before being rinsed thoroughly in deionised water.

Test type:

early seedling growth toxicity test

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

5 d

Post exposure observation period:

None

Test temperature:

The glass jars were kept at 25 ± 1°C.

pH:

The pH in the soil media was 6.4 ± 0.2.

Moisture:

Tests were conducted using 0.028 L deionised water (40 % adjusted moisture content).

Details on test conditions:

Tests were conducted in triplicate in 2.7 x 10(-4) L glass jars (i.d. 7 x 10(-2) m), each containing 10 plant seeds just below the soil surface, with 0.07 kg soil dw and 0.028 L deionised water. After 5 days exposure under a lamp, water was added to the soil to create a slurry in order to separate the plants from the soil and the seedling lengths, shoot growth and root growth, were measured to the nearest millimeter.

Nominal and measured concentrations:

The silver nanoparticles were adjusted to nominal concentrations of 0, 100, 200, 300, 400, 500, 600 and 800 mg/kg dry weight soil using deionised water before mixing with the soil.

Reference substance (positive control):

no

Species:

other: Phaseolus radiatus

Duration:

5 d

Dose descriptor:

NOEC

Effect conc.:

> 500 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Shoot length

Species:

other: Phaseolus radiatus

Duration:

5 d

Dose descriptor:

NOEC

Effect conc.:

200 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Root length

Species:

Sorghum bicolor

Duration:

5 d

Dose descriptor:

NOEC

Effect conc.:

> 500 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Shoot length

Species:

Sorghum bicolor

Duration:

5 d

Dose descriptor:

NOEC

Effect conc.:

300 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Root length

Species:

other: Phaseolus radiatus

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

298.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Shoot length

Species:

other: Phaseolus radiatus

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

186.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Root length

Species:

Sorghum bicolor

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

400 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Shoot length

Species:

Sorghum bicolor

Duration:

5 d

Dose descriptor:

EC10

Effect conc.:

301.7 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Basis for effect:

other: Root length

Details on results:

No data reported

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

The Dunnett's test for multiple comparisons was used to determine the NOEC for seedling growth and the EC50 was determined using the Trimmed Spearman-Karber method.

Validity criteria fulfilled:

not applicable

Conclusions:

The toxicity of silver nitrate to Phaseolus radiatus gave a 5 day EC10 for shoot length of 298.5 mg/kg soil dw and a 5 day EC10 of 186.6 mg/kg soil dw for the root length. For Sorghum bicolor, the 5 day EC10 for shoot length was 400 mg/kg soil dw and 301.7 mg/kg soil dw for the root length.

Executive summary:

The study is a non-guideline study, published in peer reviewed literature, with adequate description of methods and conditions and is considered suitable for use for this endpoint. The toxicity of silver nitrate to Phaseolus radiatus gave a 5 day EC10 for shoot length of 298.5 mg/kg soil dw and a 5 day EC10 of 186.6 mg/kg soil dw for the root length. For Sorghum bicolor, the 5 day EC10 for shoot length was 400 mg/kg soil dw and 301.7 mg/kg soil dw for the root length. All EC10 values were taken forward to the derivation of the PNEC according to the statistical extrapolation approach.

Description of key information

The most sensitive effects data is an EC10 of 5.2 mg/kg Ag for shoot height in Lycopersicon esculentum (tomato) from a 21 day exposure. The physico-chemical conditions of the soil were pH 4.6, organic carbon 1.9% and clay 2.5% (Langdon et al. 2013)

Additional information

In total, 53 reliable EC₁₀ or NOEC values for chronic silver toxicity to terrestrial plants were retrieved. These data cover various endpoints (shoot and root elongation, shoot biomass, shoot height) for 4 different plant species (Hordeum vulgare, Lycopersicon esculentum, Phaseolus radiatus and Sorghum bicolor) from 3 different families (Fabaceae, Poaceae and Solanaceae). NOEC and EC₁₀ values vary between 5.2 mg Ag/kg dw for 21-day shoot height of Lycopersicon esculentum (tomato, Langdon et al., 2013) and 422.0 mg Ag/kg dw for 5-day shoot elongation of Hordeum vulgare (barley, Langdon et al., 2013). All NOEC and EC₁₀ values are expressed based on total elemental Ag concentrations in soil. All reliable NOEC and EC₁₀ values were taken forward to the derivation of the PNEC according to the statistical extrapolation approach.