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Long-term toxicity to aquatic invertebrates

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Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
according to
Guideline:
other: Dutch proposed standards NEN 6501 and 6502
Principles of method if other than guideline:
Chronic toxicity and reproduction test to Daphnia magna.
Three chronic toxicity tests were carried out. In the first two tests mortality was the main criterion and reproduction was only qualitatively recorded. In the third test reproduction was the main criterion and was quantitatively measured.
GLP compliance:
no
Analytical monitoring:
no
Details on sampling:
not applicable
Vehicle:
no
Details on test solutions:
Three chronic toxicity tests were carried out:
- Method:
First test: Six concentrations of the compound: 100, 180, 320, 560, 1000 and 1800 mg/L were tested.
Second test: Five concentrations of the compound: 10, 18, 32, 56 and 100 mg/L were tested.
Third test: Five concentrations of the compound: 5.6, 10, 18, 32 and 56 mg/L were tested.
The highest concentrations were prepared by adding the required amount of melamine to DSW water and stirring this mixture until the compound was completely dissolved. The lower concentrations were prepared by diluting the highest one with DSW water.
- Controls: blank DSW water
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): not applicable
- Concentration of vehicle in test medium (stock solution and final test solution): not applicable
- Evidence of undissolved material (e.g. precipitate, surface film, etc): not stated
- Composition of the synthetic medium used in the tests (DSW water):
Na+: 1.19 mmol/L; K+: 0.20 mmol/L; Ca++: 1.36 mmol/L; Mg++: 0.73 mmol/L; Cl-: 2.72 mmol/L; SO4 --: 0.73 mmol/L; HCO3-: 1.39 mmol/L
This medium is prepared by adding several salts to ground water from the locality "Linschoten". The ground water contains several other elements in low concentrations (<< 1 mg/L). Media prepared from this water have proven to be very good for culturing several animals. The pH of the medium, after aeration, is 8.3 - 8.5. Before using this medium in toxicity tests with daphnids it is filtered by active carbon and filter paper, the latter in order to remove any carbon particles.
Test organisms (species):
Daphnia magna
Details on test organisms:
Valid for all of the three tests:

TEST ORGANISM
- Common name: Daphnia magna
- Source: laboratory-cultured
- Feeding during test
- Food type: algal cells
- Amount and Frequency: The daphnids were fed with 10exp8 algal cells per day per 40 daphnids, this supply being gradually raised to 1.5 x 10exp9 cells for daphnids of 10 days or older.
- Age at study initiation: less than 24 hrs old

ACCLIMATION
- Acclimation period: no data
- Acclimation conditions (same as test or not): no data
- Type and amount of food: no data
- Feeding frequency: no data
- Health during acclimation (any mortality observed): no data

Valid for the third of the three tests where reproduction was the main criterion and was quantitatively measured
- Age of parental stock (mean and range, SD): 40 Daphnids (less than 24 hrs old) were exposed for 7 days per concentration (Control, 5.6, 10, 18, 32 and 56 mg/L). At day 7 25 egg-bearing daphnids were taken from the 40 (or less) daphnids still surviving. With these animals the test was continued and any new born young were counted and removed at each replacement day. The test continued for 3 weeks.
Therefore the age of the parental stock was less than 24 hrs old when introduced to the test media and 7 to 8 days when progenity was started to be counted.



Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Remarks on exposure duration:
Three tests were conducted. The exposure varied from 7 to 21 days.
Post exposure observation period:
no data
Hardness:
no data
Test temperature:
20°C
pH:
Melamine did not influence the pH of the test solutions in the concentrations used.
pH of the DSW water used for preparation of the test media: 8.3 to 8.5.
Dissolved oxygen:
All test solutions were almost saturated with oxygen during the tests.
Salinity:
Not applicable.
Nominal and measured concentrations:
Three chronic toxicity tests with the following nominal concentrations were carried out:
First test: Six concentrations of the compound: 100, 180, 320, 560, 1000 and 1800 mg/L were tested.
Second test: Five concentrations of the compound: 10, 18, 32, 56 and 100 mg/L were tested.
Third test: Five concentrations of the compound: 5.6, 10, 18, 32 and 56 mg/L were tested.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: disposal plastic tanks
- Aeration: From the fourth day onwards the test solutions were aerated
- Renewal of test solution: 3 times a week: each Monday, Wednesday and Friday
- No. of organisms per vessel: 40 for the first two tests and during the first week of the third test; At day 7 of the third test 25 egg-bearing daphnids were taken from the 40 (or less) daphnids still surviving. With these animals the test was continued and any new born young were counted and removed at each replacement day (Monday, Wednesday and Friday). The test continued for 3 weeks.
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): not applicable
- Biomass loading rate: no data


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
DSW water was used for the test:
Na+: 1.19 mmol/L; K+: 0.20 mmol/L; Ca++: 1.36 mmol/L; Mg++: 0.73 mmol/L; Cl-: 2.72 mmol/L; SO4 --: 0.73 mmol/L; HCO3-: 1.39 mmol/L
This medium is prepared by adding several salts to ground water from the locality "Linschoten". The ground water contains several other elements in
low concentrations (<< 1 mg/L). Media prepared from this water have proven to be very good for culturing several animals. The pH of the medium,
after aeration, is 8.3 - 8.5. Before using this medium in toxicity tests with daphnids it is filtered by active carbon and filter paper, the latter in order to
remove any carbon particles.
- Culture medium different from test medium: not stated.
Reference substance (positive control):
no
Duration:
7 d
Dose descriptor:
LC50
Effect conc.:
> 32 - < 56 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
21 d
Dose descriptor:
LC50
Effect conc.:
> 32 - < 56 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
> 32 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
18 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- The LC50 values after chronic exposure could not be calculated exactly, since there were no concentrations with partial mortality. From the results given in the tables it appears that the LC50 7 d is about the same as the LC50 21 d and will be > 32 mg melamine / L and < 56 mg melamine /L.

- At 32 mg melamine / L there may be a small effect on mortality.

- No effect on reproduction was found. All surviving animals reproduced normally, compared to the blanks.

- At 18 mg melamine / L no effect on mortality or reproduction was found.
Results with reference substance (positive control):
no data
Reported statistics and error estimates:
no data

Table1, Percentage survival of daphnids in the reproduction test:

conc. melamine mg/L              % survival
    1st part of the test        2nd part of the test
 4 d  7 d  7 d  14 d  21 d
0  100  99  100  100  96
5.6  100  99  100  100  100
10  99  99  100  98  94
 18  100  100  100  100  100
 32  99  93  100  98  99
 56  99  0  -  -  -

Table 2, Percentage reproduction of daphnids after chronic exposure:

conc. melamine mg/L cumulative number of young per female % reproduction average % reproduction
0 113 100 100
0 114
5.6 109 96 96
5.6 110 96
10 127 111 108
10 119 104
18 108 95 97
18 113 99
32 105 92 96
32 114 100
Validity criteria fulfilled:
not specified
Conclusions:
The chronic toxicity and the reproduction toxicity of melamine to Daphnia magna is low.
Executive summary:

The compound melamine (2,4,6-triamino-triazine) was tested for acute and chronic toxicity to the water flea Daphnia magna. Together with the observation of the test organisms, pH values and oxygen levels were measured. The pH values of the test solutions and the blanks were approximately the same. The test solutions were almost saturated with oxygen in all tests.

The NOEC 21d for reproduction and mortality was found to be 18 mg/L.

The LC50 7d and 21d were estimated to be >32 mg/L and < 56 mg/L.

Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
Please refer to the analogue justification attached to section 13
Reason / purpose:
read-across source
Duration:
21 d
Dose descriptor:
LC50
Effect conc.:
> 32 - < 56 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
18 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction

Description of key information

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater invertebrates:
18 mg/L

Additional information

Data on the chronic toxicity of the target substance diphosphoric acid, compound with 1,3,5-triamine (1:2) to aquatic invertebrates are not available. The evaluation of the chronic toxicity to aquatic invertebrates is based on a read across approach with available data on the source substance melamine (CAS 108-78-7). The read across approach is in accordance with Regulation (EC) No. 1907/2006, Annex XI, 1.5. Further justification is given in the analogue justification in IUCLID section 13.

The acute and chronic toxicity of melamine (CAS 108-78-7) to Daphnia magna was tested in the available study. The study comprised three different tests conducted with different test concentrations and varying exposure periods. All test were carried out in a semi-static set up, analytic dose verification was not performed.

In a first test, conducted with nominal test substance concentrations of 100, 180, 320, 560, 1000 and 1800 mg/L, the mortality of the Daphnids was recorded over a test period of 7 days. In a second test Daphnids were exposed to nominal concentrations of 10, 18, 32, 56 and 100 mg/L. The animals were exposed for 14 days and mortality was the only effect parameter recorded during the exposure period.

In a third test mortality and reproduction of the surviving Daphnids was recorded during an exposure period of 21 days. The Daphnids were exposed to nominal test substance concentrations of 5.6, 10, 18, 32 and 56 mg/L. The determined NOEC (21 d) for mortality was 18 mg/L (NOEC (21 d) reproduction: >32 mg/L). The LC50 (7d) and LC50 (21 d) were estimated to be >32 mg/L and < 56 mg/L, respectively.