Reaction products of disodium 3,5-diamino-2-[(2-sulfonatophenyl)diazenyl]benzoate and  diazotised sodium 2-[(4-aminophenyl)sulfonyl]ethyl sulfate

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Study initiation date - 30 January 2008; Experiment start date - 30 January 2008; Experiment end date - 20 February 2008; Study completion date - 20 May 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Identity: FAT 40810/A
Batch: WP 6/02
Purity: approx. 75 %
Appearance: Solid, dark brownish powder
Expiration date: 12 December 2010
Storage: At room temperature at about 20 °C

Analytical monitoring:

yes

Details on sampling:

Samples from stock solution and test solutions were analysed to determine the actual test item concentrations in comparison to the nominally applied concentrations. Samples of the stock solution S1 and the fresh test solutions were taken on days 0, 7 and 14 from the measuring flasks. Samples of the aged test solutions were taken on days 2, 9 and 16 after the content of the replicates of a given concentration was mixed. Samples were taken from all test solutions.

Vehicle:

no

Details on test solutions:

Medium M4 was used to dilute the test item and to keep the daphnids during the period of the test. The medium consists of trace elements, vitamins and macro-nutrients. The required amounts of Medium M4 were prepared not longer than 4 weeks before they were used. During storage Medium M4 was aerated. Before use the physical-chemical characteristics of the Medium M4 were determined as required by the test guideline. Preparation of Test Solutions: After stirring S1 for 5 min at room temperature, the test solutions were prepared by diluting corresponding volumes of S1 with Medium M4. The test item coloured the stock solution and it appeared red. In the test solutions the intensity of the colour varied with the concentration levei from red in the highest concentration to light orange in the lowest concentration.

Test organisms (species):

Daphnia magna

Details on test organisms:

Material of stock vessel: glass
Amount of medium per stock vessel: 1.8 L
Depth of medium in the stock vessel: approximately 15 cm
Number of daphnids kept as stock per culture vessel (batch size): approximately 20
Separation of adult and young daphnids: 1 to 4 times per week
Medium: Elendt Medium M4
Renewal of medium: batchwise — approximately 2 times per week
Temperature: 20 ± 2 °C
Photoperiod (light/dark) :16/8 h
Light intensity: 50 - 1000 Iux
Aeration: none
Food in the cultures: Algae (Desmodesmus subspicatus, Pseudokirchneriella subcapitata), TetraMin, yeast
Feeding frequency: 3 times per week
One day before starting the test all young daphnids were removed from the selected holding vessels which contained adult daphnids. At least 6 hours before placing young daphnids into the test vessels the newborn daphnids were separated from parental daphnids which were kept in the selected holding vessels.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

246.5 - 250.0 mg/L CaCO3

Test temperature:

19.6 °C (mean value of 14 measurements; 19.0 — 20.6 °C), manual measurement;
19.55 °C (mean value of 506 measurements; 17.4 — 22.2 °C), online measurement

pH:

7.6 - 7.8

Dissolved oxygen:

96.0 - 96.8 % (8.8 - 8.9 mg/L)

Nominal and measured concentrations:

Concentrations of FAT 40810/A in the blank medium were found to be below the limit of quantification. The concentration of FAT 40810/A found in the test water samples ranged from 6.1 to 6.7 mg/L for the samples at a nominal concentration of 6.3 mg/L (mean recovery 103 %), from 49.6 to 54.5 mg/L for the samples at nominal concentration of 50 mg/L (mean recovery 105 %) and from 102.2 to 108.8 mg/L for the samples at nominal concentration of 100 mg/L (mean recovery 106 %). The following nominal concentrations were used in this study: control (0 mg/L), 6.3, 12.5, 25.0, 50 and 100 mg/L

Details on test conditions:

Test organism: Daphnia magna (Straus), clone 5
Age of parental daphnids: 19 - 22 d (offspring used was not the first brood of these parental daphnids)
Age of the daphnids at the beginning of the test: 6 — 24 h
Number of test concentrations: 5 plus a control
Test concentrations: 6.3; 12.5; 25.0; 50.0; and 100.0 mg/L plus control
Number of replicates per test concentration: 12 at start of exposure
Number of organisms per test vessel: 1 per replicate at start of exposure
Test units: 100 mL glass beaker, covered by glass lids
Amount of test solution per test vessel: 70 - 90 mL
Test medium: Elendt Medium M4
Application of test item: test item dissolved in Elendt medium M4 applied once at the beginning of the test and then 3 times per week (renewal of test solutions)
Test duration (exposure): 21d
Aeration of test vessels: none
Adjustment of pH in the test solutions: none
Renewal of the test solution during the test period: semi-static conditions; test solutions renewal three times per week
Feeding during exposure: three times per week with fresh algae (Desmodesmus subspicatus) suspension (0.1 — 0.2 mg C/daphnia *d)
Temperature: 19.6 °C (mean value of 14 measurements; 19.0 — 20.6 °C), manual measurement;
19.55 °C (mean value of 506 measurements; 17.4 — 22.2 °C), online measurement
Light regimen: 16 h light / 8 h dark
Light intensity: 458 Ix at the beginning of the test (mean value of 5 measurements; 408 - 594 Ix)

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

ca. 50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Details on results:

Mortalities of the parental daphnids in the highest concentration started on day 5, on this date 9 of 12 daphnids died. On day 8, mortalities reached 100 % in the highest concentration. From day 10 on aborted eggs were observed occasionally in all concentration levels, without showing a definite concentration-response pattern. For the parameter fecundity no reduction of the offspring was found up to the concentration where the survival of the daphnids fulfilled the validity criteria for the controls. The parameter length of the parental daphnids showed in all treatments a significant difference compared to the controls.

Results with reference substance (positive control):

not applicable

Reported statistics and error estimates:

The statistical software packages ToxRat Professional 2.09 was used for these calculations.

Validity criteria fulfilled:

yes

Conclusions:

The NOEC for mortality and reproduction in this daphnia reproduction test with FAT40810/A was set to 50 mg/L and the LOEC to 100 mg/L.

Executive summary:

The aim of the study was to determine the chronic toxicity of the test item on the water flea Daphnia magna. To achieve this, a series of concentrations of the test item in aqueous solution was prepared, and the test organisms were exposed to these concentrations, as well as to controls without the test item, for a period of 21 days. During the test period the test solutions were renewed in a static-renewal test system. This study was conducted in accordance with OECD test guideline 221 in a GLP-certified laboratory. The test concentration used was 0, 6.3, 12.5, 25, 50 and 100 mg/L (nominal). ln the highest test item concentration level (100.0 mg/L) all parental daphnids died during the exposure period. In all other test concentrations no significant differences were found in the treatments compared to the controls (Williams' t-test, p≤0.05) with the parameter fecundity. Thus the NOEC in this test was 50 mg/L. Under the conditions of this test, the corresponding LOEC is therefore expected to fall between 50 and 100 mg/L. A calculation of the EC50 for fecundity was not possible since there was no significant inhibition of reproduction at all concentration levels with surviving adults (6.3, 12.5, 25.0 and 50.0 mg/L). A significant difference between all treatments and the control was established (Williams' t-test, p≤0.05) for the length of the daphnids at the end of the test. Due to statistical reasons the standard deviation of the values in the control and the treatments are very small and the ANOVA reacts therefore very sensitive. Thus, the effect on length is interpreted as a slight effect. The results of this test indicate that the reproduction is not influenced by the test item at concentration levels with surviving parental daphnids. However, there were slight effects on the length of the parental daphnids at the end of the test at all treatment levels. Based on the findings of this study, the NOEC for mortality and reproduction in this test was set to 50 mg/L and the LOEC to 100 mg/L.

Description of key information

NOEC for mortality and reproduction: 50 mg/L

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

50 mg/L

Additional information

The aim of the study was to determine the chronic toxicity of the test item on the water flea Daphnia magna. To achieve this, a series of concentrations of the test item in aqueous solution was prepared, and the test organisms were exposed to these concentrations, as well as to controls without the test item, for a period of 21 days. During the test period the test solutions were renewed in a static-renewal test system. This study was conducted in accordance with OECD test guideline 221 in a GLP-certified laboratory. The test concentration used was 0, 6.3, 12.5, 25, 50 and 100 mg/L (nominal). ln the highest test item concentration level (100.0 mg/L) all parental daphnids died during the exposure period. In all other test concentrations no significant differences were found in the treatments compared to the controls (Williams' t-test, p≤0.05) with the parameter fecundity. Thus the NOEC in this test was 50 mg/L. Under the conditions of this test, the corresponding LOEC is therefore expected to fall between 50 and 100 mg/L. A calculation of the EC50 for fecundity was not possible since there was no significant inhibition of reproduction at all concentration levels with surviving adults (6.3, 12.5, 25.0 and 50.0 mg/L). A significant difference between all treatments and the control was established (Williams' t-test, p≤0.05) for the length of the daphnids at the end of the test. Due to statistical reasons the standard deviation of the values in the control and the treatments are very small and the ANOVA reacts therefore very sensitive. Thus the effect on length is interpreted as a slight effect. The results of this test indicate that the reproduction is not influenced by the test item at concentration levels with surviving parental daphnids. However, there were slight effects on the length of the parental daphnids at the end of the test at all treatment levels. Based on the findings of this study, the NOEC for mortality and reproduction in this test was set to 50 mg/L and the LOEC to 100 mg/L.