long chain alkyl thio carbamide metal complex

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 July to 16 September 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD series on testing and assessment No. 23.: guidance document on aquatic toxicity testing of difficult substances and mixtures.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
no data

OTHER PROPERTIES (if relevant for this endpoint)
- Results of test for ready biodegradability: no data

Analytical monitoring:

yes

Details on sampling:

Additional flasks containing the freshly prepared control and test media were sampled (4 x 100 ml) immediately and at 72 hours, acidified with nitric acid and stored at about 4ºC until analysis.

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: a concentrated stock (94.6 g a.i./litre) was prepared by dissolving 2.7173 g of test substance in 25 ml of tetrahydrofuran (THF). Four intermediate stock solutions (4.05, 8.89, 19.7, 43.1 g a.i./litre) were prepared in THF from the concentrated stock. A 2.0 ml aliquot of each stock was applied to the inner surfaces of 2-litre glass bottles and the solvent removed by evaporation using compressed air. The bottles were filled with 2 litres of sterile culture medium and stirred in the dark at about 40ºC for 48 hours, then left at room temperature for at least 24 hours.
- Eluate: about 1 litre of each water accommodated fraction (WAF) was removed, samples were subjected to chemical analysis of molybdenum content and the nominal concentrations of active ingredient were calculated.
- Controls: yes
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): tetrahydrofuran
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): not applicable, evaporated off
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no data

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Common name: Selenastrum capricornutum
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Culture Collection of Algae and Protozoa, Institute of Freshwater Ecology, Cumbria, UK
- Age of inoculum (at test initiation): 3 days (in log phase)
- Method of cultivation: in liquid medium with shaking under continuous illumination.

ACCLIMATION
- Acclimation period: 6 days
- Culturing media and conditions (same as test or not): yes
- Any deformed or abnormal cells observed: no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

none

Hardness:

no data

Test temperature:

22.1-23.1ºC

pH:

7.7-8.1 at 0 h; 7.4-7.7 at 72 h

Dissolved oxygen:

no data

Salinity:

not applicable

Nominal and measured concentrations:

Since the test substance is not readily soluble in water, it was coated onto the inside walls of the vessels and culture medium added. Chemical analysis of the molybdenum content of the medium was carried out to determine the water accommodated fractions (WAFs) of active ingredient. Concentrations deposited on the vessel walls were 4.05, 8.89, 19.7, 43.1 and 94.6 mg a.i./litre (nominal).

Based on the measured levels of molybdenum in the WAFs at 0 and 72 h (0.0124, 0.0451, 0.0547, 0.127 and 0.517 mg Mo/litre), the actual concentrations of active ingredient were calculated to be 0.135, 0.492, 0.597, 1.38 and 5.64 mg/litre; however, since the molybdenum content of the water soluble portion of the test material is not known, these values should be treated with caution.

Details on test conditions:

TEST SYSTEM
- Test vessel: flasks
- Type (delete if not applicable): loose cottonwool plugs
- Material, size, headspace, fill volume: glass, size: 250 ml, headspace: 150 ml, fill volume: 100 ml
- Aeration: by shaking
- Initial cells density: 1.0 X 10(4)/ml
- Control end cells density: 157 X 10(4)/ml
- No. of organisms per vessel: 1 X 10(6); (1 X 10(4)/ml)
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes, OECD medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ultrapure water (filtered, dechlorinated tap water, softened and treated by reverse osmosis, before microfiltration and purification)
- Culture medium different from test medium: no
- Intervals of water quality measurement: water quality measurements and chemical analysis performed at start of the study

OTHER TEST CONDITIONS
- Sterile test conditions: yes (test substance not sterilised)
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: about 8240 lux from 4 X 30 W "cool white" 1 metre fluorescent tubes

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): 24, 48 and 72 hours
- Determination of cell concentrations: counting chamber

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
- Justification for using less concentrations than requested by guideline: not applicable

RANGE-FINDING STUDY: yes
- Test concentrations: 0.946, 9.46, 94.6 mg a.i./litre
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

4.05 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

in medium

Basis for effect:

cell number

Remarks:

comparing the percentage inhibition in the test group with that for the control cultures

Remarks on result:

other:

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

9.62 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

in medium

Basis for effect:

other: derived from area under the growth curve

Remarks on result:

other: 95% CL 8.89-19.7 mg/L

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

14 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

in medium

Basis for effect:

growth rate

Remarks on result:

other: 95% CL 8.89-19.7 mg/L

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape: no
- Colour differences: no
- Flocculation: no
- Adherence to test vessels: no data
- Aggregation of algal cells: no data
- Any stimulation of growth found in any treatment: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: not applicable

Results with reference substance (positive control):

not applicable

Reported statistics and error estimates:

no data

 Table 1. Growth inhibition

Nominal active ingredient (mg/litre)^a	Replicate	Area under curve (72 hr) x 10^-4	Mean (% inhibition)	Growth rate (0-72 hr) x 10^-2	Mean (% inhibition)
Control	1 2 3 4 5 6	2767 2392 3274 3026 2710 1830	2667	7.040 6.922 7.250 7.117 7.058 6.661	7.008
4.05  WAF	1 2 3	2143 2452 2779	2458 (8)	6.672 6.863 7.023	6.853 (2)
8.89  WAF	1 2 3	1589 1507 1448	1515 (43)	6.334 6.180 6.169	6.228 (11)
19.7  WAF	1 2 3	81.12 49.68 57.12	62.64 (98)	1.526 1.204 1.273	1.334 (81)
43.1  WAF	1 2 3	12.12 1.68 4.56	6.12 (100)	0.3099 0.1697 0.1697	0.216 (97)
94.6  WAF	1 2 3	-1.44 -7.44 -4.44	-4.44 (100)	0.1697 0.1697 0.1697	0.1697 (98)

 ^aThe active ingredient refers to EC434 -650 -5; water accommodated fractions tested

 

Table 2. Cell densities

Nominal active ingredient (mg/litre)^a	Mean cell densities (x 10^4cells/ml)   24 hr           48 hr                  72 hr
Control	4.44	30.8	157
4.05  WAF	3.71	31.4	140
8.89  WAF	3.33	17.9	88.7
19.7  WAF	1.80	2.0	2.63
43.1  WAF	1.17	1.0	1.17
94.6  WAF	0.917	0.833	1.13

 ^aThe active ingredient refers to EC: 434 -650 -5; water accommodated fractions tested

Validity criteria fulfilled:

yes

Conclusions:

In a GLP study conducted according to OECD guideline 201 and OECD guidance on testing and assessment No. 23, EC# 434-650-5 caused a concentration-related inhibition of algal growth when tested at nominal concentrations of up to 94.6 mg a.i./litre for a 72 h incubation period. The NOELR, based on cell number, was 4.05 mg a.i./litre (nominal loading rate).

Executive summary:

In a GLP study conducted according to OECD guideline 201 and OECD guidance on testing and assessment No. 23, EC# 434-650-5 test material was assessed for the potential to inhibit the growth of the freshwater algal species, Selenastrum capricornutum.

 

Since the test material was not very soluble in solvents that would not affect algal growth, various concentrations were dissolved in tetrahydrofuran and coated onto the inner surfaces of glass bottles, the solvent evaporated off, and the bottles filled with water and stirred for 72 h. The water accommodated fractions (WAFs) were removed and analysed for molybdenum (Mo) content, and the concentrations of active ingredient (EC# 434-650-5) were calculated for each WAF.

 

Flasks containing the algal strain, culture medium and nominal concentrations of 0, 4.05, 8.89, 19.7, 43.1 or 94.6 mg a.i./litre of were prepared and incubated with shaking for 72 h under continuous illumination. Three flasks were prepared for each test concentration and six flasks for the control cultures. Cell densities were measured at 0, 24, 48 and 72 h and used to calculate the growth rate and area under the curve for each culture. Based on the measured levels of Mo in the WAFs at 0 and 72 h (0.0124, 0.0451, 0.0547, 0.127 and 0.517 mg Mo/litre), the actual concentrations of active ingredient were calculated to be 0.135, 0.492, 0.597, 1.38 and 5.64 mg/litre; however, since the Mo content of the water-soluble portion of the test material is not known, these values should be treated with caution and have therefore not been used to derive the endpoints.

 

The test material caused a concentration-related inhibition of algal growth. The no-observed-effect loading rate was 4.05 mg a.i./litre, based on cell number, and the E_bL₅₀and E_tL₅₀ were 9.62 and 14.0 mg a.i./litre, respectively, based on area under the growth curve and average specific growth rate.

 

In conclusion, the test material showed evidence of an inhibitory effect on the growth of the algal species, Selenastrum capricornutum, when tested at nominal concentrations of up to 94.6 mg a.i./litre. In view of the structural and chemical similarities, it is considered that the results of this study can be used for read-across to EC# 457-320-2.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reason / purpose for cross-reference:

read-across source

Duration:

72 h

Dose descriptor:

EL50

Remarks:

WAF

Effect conc.:

9.62 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Remarks:

Area under the growth curve

Key result

Duration:

72 h

Dose descriptor:

EL50

Remarks:

WAF

Effect conc.:

14 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Remarks:

Average specific growth rate

Key result

Duration:

72 h

Dose descriptor:

NOELR

Remarks:

WAF

Effect conc.:

4.05 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Validity criteria fulfilled:

yes

Conclusions:

In a GLP study conducted according to OECD guideline 201 and OECD guidance on testing and assessment No. 23, EC# 434-650-5 caused a concentration-related inhibition of algal growth when tested at nominal concentrations of up to 94.6 mg a.i./litre for a 72 h incubation period. The NOELR, based on cell number, was 4.05 mg a.i./litre (nominal loading rate).

Executive summary:

In a GLP study conducted according to OECD guideline 201 and OECD guidance on testing and assessment No. 23, EC# 434-650-5 test material was assessed for the potential to inhibit the growth of the freshwater algal species,Selenastrum capricornutum.

 

Since the test material was not very soluble in solvents that would not affect algal growth, various concentrations were dissolved in tetrahydrofuran and coated onto the inner surfaces of glass bottles, the solvent evaporated off, and the bottles filled with water and stirred for 72 h. The water accommodated fractions (WAFs) were removed and analysed for molybdenum (Mo) content, and the concentrations of active ingredient (EC# 434-650-5) were calculated for each WAF.

 

Flasks containing the algal strain, culture medium and nominal concentrations of 0, 4.05, 8.89, 19.7, 43.1 or 94.6 mg a.i./litre of were prepared and incubated with shaking for 72 h under continuous illumination. Three flasks were prepared for each test concentration and six flasks for the control cultures. Cell densities were measured at 0, 24, 48 and 72 h and used to calculate the growth rate and area under the curve for each culture. Based on the measured levels of Mo in the WAFs at 0 and 72 h (0.0124, 0.0451, 0.0547, 0.127 and 0.517 mg Mo/litre), the actual concentrations of active ingredient were calculated to be 0.135, 0.492, 0.597, 1.38 and 5.64 mg/litre; however, since the Mo content of the water-soluble portion of the test material is not known, these values should be treated with caution and have therefore not be used to derive C&L.

 

The test material caused a concentration-related inhibition of algal growth. The no-observed-effect loading rate was 4.05 mg a.i./litre, based on cell number, and theE_bL₅₀and E_tL₅₀were 9.62 and 14.0 mg a.i./litre, respectively, based on area under the growth curve and average specific growth rate.

 

In conclusion, the test material showed evidence of an inhibitory effect on the growth of the algal species, Selenastrum capricornutum, when tested at nominal concentrations of up to 94.6 mg a.i./litre. In view of the structural and chemical similarities, it is considered that the results of this study can be used for read-across to EC# 457-320-2.

Description of key information

In an algal growth inhibition study of EC# 434-650-5, the 72-hour NOELR based on cell number was 4.05 mg/litre (nominal loading rate).

Key value for chemical safety assessment

EC50 for freshwater algae:

9.62 mg/L

EC10 or NOEC for freshwater algae:

4.05 mg/L

Additional information

No toxicity to aquatic algae and cyanobacteria studies are available for EC# 457-320-2; however, one study is available for the structurally and chemically similar substance EC# 434-650-5: a 72-hour algal growth inhibition study conducted to GLP and in accordance with OECD Test Guideline 201. Due to the poor solubility of the test material in solvents that would not affect algal growth, test waters were prepared as Water Accommodated Fractions (WAFs) of nominal loading rates of active ingredient (i.e. EC# 434-650-5). WAFs were sampled and analysed for molybdenum (Mo) content, and the concentrations of EC# 434-650-5 calculated.

When this study was performed for the purposes of the ELINCS registration, the source substance was considered to be a mono-constituent and the purity to be 92%, however, after the study completed the purity was determined to be in fact 87%. Therefore, the nominal test concentrations employed (4.27, 9.39, 20.7, 45.5 and 100 mg ai/L) and endpoints (NOELR and EL50) were recalculated in terms of their active ingredient content (4.05, 8.89, 9.46, 19.7, 43.1 and 94.6 mg ai/L). More recent evaluation of the composition of these substances suggests that it is more appropriate to consider them UVCBs. Therefore the differences in purity are likely to reflect the variation in the UVCB composition. This change in purity and, therefore, calculated test concentrations and results are not considered to have affected the integrity or reliability of the data, nor the suitability for read-across. In fact, the altered endpoints are considered to represent a worst case EC50 and NOELR.

Based on the measured levels of Mo at 0 and 72 h (0.0124, 0.0451, 0.0547, 0.127 and 0.517 mg Mo/litre), the actual concentrations of active ingredient were calculated to be 0.135, 0.492, 0.597, 1.38 and 5.64 mg/litre; however, since the Mo content of the water-soluble portion of the test material is not known, these values should be treated with caution and have not been used to derive C&L for this endpoint.

 

The test material caused a concentration-related inhibition of algal growth. The no-observed-effect loading rate was 4.05 mg a.i./litre, based on cell number, and the E_bL₅₀ and E_tL₅₀ were 9.62 and 14.0 mg a.i./litre, respectively, based on area under the growth curve and average specific growth rate.

 

In view of the structural and chemical similarities, it is considered that the results of this study can be used for read-across to EC# 457-320-2.