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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-01-24 - 2017-03-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2017

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
OECD Guidelines for Testing of Chemicals No. 202. Daphnia sp., Acute Immobilisation Test. Adopted: 13 April 2004
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
LUBW Landesanstalt für Umwelt, Messungen und Naturschutz Baden-Württemberg, Postfach 10 01 63, 76231 Karlsruhe

Test material

Constituent 1
Chemical structure
Reference substance name:
3-[(aminoiminomethyl)thio]propanesulphonic acid
EC Number:
244-520-8
EC Name:
3-[(aminoiminomethyl)thio]propanesulphonic acid
Cas Number:
21668-81-5
Molecular formula:
C4H10N2O3S2
IUPAC Name:
3-(carbamimidoylsulfanyl)propane-1-sulfonic acid
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: ambient (≤ + 30 °C), dark, dry

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Sampling method: Analytical samples were taken from all test item concentrations and control at test start and after 48 hours from aged solutions. For each sampling also a retain sample was taken.
- Sample storage conditions before analysis: All samples were stored deep frozen until they were transferred to the analytical laboratory.

Test solutions

Vehicle:
no
Remarks:
Test medium was used.
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The necessary amount of test item for preparing the stock solution was weighed on a weighing scoop and transferred to a volumetric flask. Test medium was added up to the bench mark and the solution was homogenised by shaking. Afterwards the solution was clear and transparent. Lower test solutions were prepared by dilution of the appropriate solution with test medium. 50 mL of the prepared solutions were transferred to each test vessel.
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)):
100, 10, 1, 0.1, 0 mg/L (GLP range finding test)
100 (stock), 45.5, 20.7, 9.39, 4.27, 0 mg/L (main test)
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Strain/clone: Daphnia magna Straus, Clone V
- Justification for species other than prescribed by test guideline: n/a
- Source: The animals are continuously bred in the laboratory and were originally purchased in a healthy condition from the Federal Environment Agency in Berlin/Germany. Daphnia magna was bred as single culture (1 daphnid per 100 mL) in Elendt M4 medium. The pH-value of the aerated water was within a range of 6.0 – 9.0. The dissolved oxygen was above 60 % saturation and the total hardness 140 - 250 mg/L (as CaCO3), corresponding to 7.8 - 14°dH. The animals were fed with single cell green algae (Desmodesmus subspicatus, formerly Scenedesmus subspicatus) at least three times a week.
The daphnids were reared at a temperature of 20 ± 2 °C in a climatic chamber with 16 hours of illumination and 8 hours of darkness. The medium was changed three times per week. A pipette was used to separate the young daphnids from the adults.
Freshly hatched daphnids less than 24 hours old were used for the test.
- Feeding during test: none
- Food type: single cell green algae (Desmodesmus subspicatus, formerly Scenedesmus subspicatus)
- Frequency: at least three times a week

ACCLIMATION
- Acclimation period: n/a; the animals are continuously bred in the laboratory

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES:
A pipette was used to separate the young daphnids from the adults. Freshly hatched daphnids less than 24 hours old were used for the test.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
none

Test conditions

Hardness:
12°dH (214 mg/L as CaCO3)
Test temperature:
19.1 – 20.5 °C
pH:
7.64 – 7.99
Dissolved oxygen:
≥ 8.6 mg/L
Nominal and measured concentrations:
100, 45.5, 20.7, 9.39, 4.27, 0 mg/l (main test, nominal)
106, 44.9, 20.0, 8.78, 3.31, n.d. mg/l (main test, fresh medium, measured)
113, 46.0, 21.5, 8.85, 3.64, n.d. mg/l (main test, 48h aged medium, measured)
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass vessel (100 mL)
- Type (delete if not applicable): The test units were covered with a glass plate (thus reducing evaporation).
- Material, size, headspace, fill volume: 50 ml fill volume
- Aeration: none during test
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours photoperiod /8 hours darkness daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
After 24 h and 48 h the immobilised daphnids were counted. All daphnids not able to swim within 15 seconds after gentle agitation of the test vessel were considered to be immobilised. If present, behavioural changes of daphnids were recorded at 24 and 48 hours after starting the test.
The test temperature and the pH-value as well as the oxygen concentration of the test solutions were measured at all concentrations at t = 0 h fresh, t = 24 h and t = 48 h from aged test solutions in one separate replicate per test item concentration without test organisms.

VEHICLE CONTROL PERFORMED: yes (test medium)

RANGE-FINDING STUDY
- Test concentrations: In the GLP range finding test the following nominal concentrations were tested: 100, 10.0, 1.00 and 0.100 mg/L and control.
- Results used to determine the conditions for the definitive study:
Table: Results of the range finding test, 24 h values
Control 0.100 1.00 10.0 100
mg/L
immobilised daphnids after 24 h
Group 1 0 0 0 0 2
Group 2 0 0 0 0 3
Group 3 0 0 0 0 1
Group 4 0 1 0 0 1
Σ 0 1 0 0 7
% 0 5 0 0 35

Table Results of the range finding test, 48 h values
Control 0.100 1.00 10.0 100
mg/L
immobilised daphnids after 48h
Group 1 0 0 0 0 2
Group 2 0 1 0 0 3
Group 3 0 0 0 0 1
Group 4 0 1 0 1 0
Σ 0 2 0 1 6
% 0 10 0 5 30
Reference substance (positive control):
yes
Remarks:
potassium dichromate (1.0 mg/L, 2.0 mg/L)

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
40.3 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % confidence limit of EC50: 26.1 – 75.7 mg/l
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
4.27 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
45.5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Behavioural abnormalities: none stated
- Other adverse effects control: no immobility occurred
- Abnormal responses: none stated
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none stated
- Effect concentrations exceeding solubility of substance in test medium: none
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Relevant effect levels: The results indicate an EC50 (24 h) of the reference item potassium dichromate between 1.0 mg/L and 2.0 mg/L. Since the results are in accordance with the requirements of the OECD guideline 202 and fall within the historical data generated with the reference item at the testing facility, the daphnids were suitable for the determination of the toxicological effects of the test item.
- Limit test: no
- Dose-response test: yes
- Other:
Results of the toxic reference test, started on 14 Feb 2017
24 h 48 h
K2Cr2O7 1.00 2.00 1.00 2.00
[mg/L] immobilised daphnids
Group 1 1 5 5 5
Group 2 1 5 5 5
Group 3 2 5 5 5
Group 4 2 5 5 5
Sum 6 20 20 20
% 30 100 100 100

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The study was conducted under GLP according to OECD guideline 202 on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or deviations from the guidelines, the validity criteria were met. Hence, the results can be considered as reliable to assess the toxicity of UPS towards daphnids.
The toxicity against Daphnia magna was tested in a static test design using several concentrations up to 100 mg/L. According to the results of the test, the EC50 (48 h) was determined to be 40.3 mg/L (nominal). The corresponding NOEC (48 h) was 4.27 mg/L (nominal).
Based on the obtained results, 3-[(amino-iminomethyl)-thio]-1-propanesulphonic acid does not need to be classified as hazardous to the aquatic environment, i.e. short-term hazard, Acute Cat.1, according to the Regulation (EC) No. 1272/2008. A substance must be classified as Aquatic Chronic Category 3, if the 48 hr EC50 (for crustacea) is > 10 to ≤ 100 mg/l and the substance is not rapidly degradable and/or the experimentally determined BCF ≥ 500 (or, if absent, the log Kow ≥ 4). In a OECD 301F study, UPS was determined to be not readily biodegradable, the partition coefficient was -2. So although the log Pow is not ≥ 4, the substance is not rapidly degradable, and should be hence classified as Aquatic Chronic Category 3 according to the Regulation (EC) No. 1272/2008.
Executive summary:

The objectives of this study according to OECD TG 202 under GLP were to determine the immobilisation effect of 3-[(amino-iminomethyl)-thio]-1-propanesulphonic acid (UPS) on the water flea Daphnia magna under worst-case exposure conditions, the no observed effect concentration (NOEC) and the effect median concentration (EC50).

Methods:

Test species:Daphnia magna Straus, Clone V, max. 24 hours old.

Test design: Static dose-response test with twenty organisms per test concentration (4 replicates of 5 animals each) were used. The duration of the test was 48 hours.

Endpoints: Endpoints reported are the EC50and the NOEC after 24 and 48 hours.

Test rates: Following a static range-finding test with nominal test item concentrations of 0.100, 1.00, 10.0 and 100 mg/L and control a static main test with nominal concentrations of 4.27, 9.39, 20.7, 45.5 and 100 mg/L and control was performed.

Test conditions: Temperature, pH-value and oxygen concentration of the test solutions measured after 0 hours fresh, 24 hours and 48 hours are reported. Hardness of the test water was measured on the day of application.

Samples analysed: Analytical samples taken at 0 hours (initial value) and 48 hours from aged test solutions were analysed from control and all test item concentrations.

Statistics: The values for EC50were determined by Probit analysis using linear max. likelihood regression. The NOEC was established based on the highest concentration at which the immobilisation is not higher than the allowed control immobilisation (≤ 10 % immobilisation).

Findings:

Validity criteria: Control immobilisation: The percentage of immobilisation should be ≤ 10 %. In this study the control immobilisation was 0 %.

Oxygen concentration: The dissolved oxygen concentration at the end of the test should be ≥ 3 mg/L in all test units. In this test, the dissolved oxygen concentration at the end of the test was ≥ 8.6 mg/L.

Test conditions: The total hardness (as CaCO3) of the untreated control was determined to be 12°dH (214 mg/L CaCO3); the mean pH-value of the untreated control was determined to be 7.75 ± 0.10 (Std. Dev.), the mean temperature of the control and all test item concentrations was measured to be 19.8 ± 0.5 °C (Std. Dev.) and the mean oxygen concentration was determined to be 9.0 ± 0.3 mg/L (Std. Dev.).

Analytical Results: The initial measured content of active ingredient was between 78 and 106 % of nominal and the aged measured content was between 86 and 113 % of nominal. Therefore the toxicological endpoints were evaluated using nominal concentrations.

Statistical Results: EC50and NOEC-values of daphnids exposed to the test item evaluated using nominal concentrations

 

UPS [mg/L] (nominal)

 

24 h

48h

NOEC

45.5

4.27

EC50

> 1001)

40.32)

95 % confidence limit of EC50

-

26.1 – 75.7

- not applicable

1)Due to an inhibition below 50% the database was weak for probit analysis which hence was not performed

2)Probit analysis using linear max. likelihood regression

 

Conclusions: According to the results of the test, the EC50(48 h) was determined to be 40.3 mg/L (nominal). The corresponding NOEC (48 h) was 4.27 mg/L (nominal).

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