Dimethyl acetylsuccinate

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:

3 May 2010 to XXXXXXX

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study with GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Dimethyl acetylsuccinate

EC Number:

233-897-4

EC Name:

Dimethyl acetylsuccinate

Cas Number:

10420-33-4

Molecular formula:

C8H12O5

IUPAC Name:

dimethyl 2-acetylsuccinate

Details on test material:

Batch manufactured and supplied by: DSM Fine Chemicals, Austria Nfg GmbH & Co KG, St.-Peter-Straße 25, P.O. Box 933, 4021 Linz / Austria
Test Item: Dimethylacetylsuccinate
Common Name: DMAS
CAS No.: 10420-33-4
Chemical Name (IUPAC): (+/-)Acetylsuccinicaciddimethylester
Physical Appearance: Solid
Purity as per Certificate of Analysis: 97.8%
Batch No.: LEDM5A1003
Manufactured Date: Dec. 2009
Expiry Date: not applicable
Retest Date: Dec. 2011
Recommended Storage Condition: Ambient (+18 to +36º)
Note: Date of receipt of test item at Test Facility: 29.03.2010
Test item code by Test Facility: D011-05

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: nominal concentrations: 15, 24, 38, 61 and 100 mg/L
- Sampling method: The test solutions of all the five test concentrations along with the negative control were analysed at the start of test, at approximately 24 hours and at the end of the test for the active ingredient concentration of the test item by the analytical R&D department of Advinus Therapeutics Private Limited.

Test solutions

Vehicle:

not specified

Details on test solutions:

Test Item Stock:
One thousand milligrams (1000 mg) of the test item were mixed with a small volume of the test medium and the volume was made up to 100 mL with the test medium using a volumetric flask (10 mg/mL).
Stock of the test item was prepared and used within approximately 20 minutes after preparation.

Test Solutions:
G1 (Negative control): 150 mL each test medium in 6 test containers
G2 (15 mg/L): 3 mL stock solution was made up to the mark in a 2 L volumetric flask using test medium. From this, 150 mL each was dispensed into 6 test containers
G3 (24 mg/L): 4.8 mL stock solution was made up to the mark in a 2 L volumetric flask using test medium. From this, 150 mL each was dispensed into 6 test containers
G4 (38 mg/L): 7.6 mL stock solution was made up to the mark in a 2 L volumetric flask using test medium. From this, 150 mL each was dispensed into 6 test containers
G5 (61 mg/L): 12.2 mL stock solution was made up to the mark in a 2 L volumetric flask using test medium. From this, 150 mL each was dispensed into 6 test containers
G6 (100 mg/L): 20 mL stock solution was made up to the mark in a 2 L volumetric flask using test medium. From this, 150 mL each was dispensed into 6 test containers
G7 (0.32 mg/L) (Positive control): 0.32 mL dilution of the positive control was mixed with 1 L of test medium. From this, 150 mL each was dispensed into 4 test containers.

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Controls: positive and negative control
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Source: Department of Safety Assessment, Advinus Therapeutics Private Limited, Bangalore, India
- Age at study initiation: less than 24 hours
- Method of breeding: Daphnia were maintained in the test medium. The test medium in which the Daphnia were maintained was changed at least twice a week and they were fed with the unicellular green alga, Chlorella vulgaris. One day prior to the start of the treatment, about 100 Daphnids were incubated in approximately 2 L of test medium for both the range finding as well as the definitive test. Next day, the young ones hatched which were less than 24-hour old were collected and used in the range finding and the definitive tests.
- Feeding during test: no

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

no

Test conditions

Hardness:

Total hardness as CaCO3: 2.36 m moles/L

Test temperature:

The test solutions were incubated under dark conditions with the temperature of the growth chamber at 20.3 to 20.4 ºC during the treatment period.

pH:

The pH of the negative control ranged between 7.74 and 7.79 during the treatment period. The pH of the test solutions ranged between 7.66 and 7.76 during the treatment period.
For details please see: Key. Bopanna 2010. DMAS. Daphnia magna, acute immobilisation test Tables and Figures.pdf

Dissolved oxygen:

The dissolved oxygen concentration of the negative control ranged between 6.2 and 7.3 mg/L during the treatment period. The dissolved oxygen concentration of the test solutions ranged between 6.0 and 7.3 mg/L during the treatment period. The average dissolved oxygen concentration of the test solutions was 6.7 ± 0.6 mg/L.
For details please see: Key. Bopanna 2010. DMAS. Daphnia magna, acute immobilisation test Tables and Figures.pdf

Salinity:

Not applicable

Nominal and measured concentrations:

Nominal concentrations: 15, 24, 38, 61 and 100 mg/L.
Measured: 0h: 15.464, 23.864, 36.351, 56.894 and 91.558 mg/L
24 h: 9.215, 15.296, 25.978, 43.237 and 76.418 mg/L
48 h: 6.139, 11.397, 20.423, 36.546 and 70.105 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL glass beakers
- Aeration: no
- No. of Daphnia: Ten Daphnia per group divided into two replicates of five Daphnia each in the range finding test. Twenty Daphnia per group divided into four replicates of five Daphnia each in the definitive test.
- No. of vessels per concentration: definitive test: 4; range finding test: 2
- No. of vessels per control: definitive test: 4; range finding test: 2
- Biomass loading rate: At least 2 mL of test solution to each Daphnia

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: no
For details please see: Key. Bopanna 2010. DMAS. Daphnia magna, acute immobilisation test Tables and Figures.pdf


OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: The test was conducted in the dark
- Light intensity: not applicable

EFFECT PARAMETERS MEASURED:
Immobility of Daphnia at 24 +/- 1 and 48 +/- 1 hours after the start of the treatment was recorded.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: ca. 1.6
- Range finding study: No immobility of the Daphnia was observed in the negative control either at 24 or at 48 hours post exposure. There was no immobilization at 24 hours in any of the tested concentration. The per cent immobilization for the test concentrations 0.1, 1, 10 and 100 mg/L at 48 hours was 0, 0, 0 and 30, respectively.
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7)

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 79.36 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

27.58 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 3

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

45.56 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: not reported
- Other biological observations: not reported
- Mortality of control: no mortality and no immobilization of daphnia was observed
- Other adverse effects control: not reported
- Abnormal responses: no
- Any observations that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

Positive control:
- Immobilisation: 0.32 mg/L Potassium dichromate (K2Cr2O7) gave 40 and 80 % immobilization of Daphnia in the positive control at 24 and 48 hours post exposure, respectively.

Reported statistics and error estimates:

The EC50 was not calculated as the % immobilization of Daphnia was not more than 50 % in any of the concentrations tested during the test period.

Any other information on results incl. tables

For more details please see: Key. Bopanna 2010. DMAS. Daphnia magna, acute immobilisation test Tables and Figures.pdf

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The assessment of toxic effects of Dimethylacetylsuccinate on Daphnia magna by 48 hour acute immobilization test at 15, 24, 38, 61 and
100 mg/L concentrations resulted in 0, 0, 0, 5 and 25 % immobilization, respectively. There was 80 % immobility in the case of positive control (0.32 mg/L Potassium dichromate) under similar test conditions and there was no immobilization observed in the negative control during the test period.

The NOEC and LOEC were observed at the test item concentrations of 38 and 61 mg/L, respectively.

Executive summary:

The acute immobilization effect of the test item Dimethylacetylsuccinate was studied onDaphnia magna. TheDaphnia magnaless than
24 hours old were exposed to nominal concentrations of 15, 24, 38, 61 and 100 mg/L of Dimethylacetylsuccinate along with negative and positive control in reconstituted water at 20 ± 1^oC. The number ofDaphniaimmobilized was recorded at 24 and 48 hours post exposure.

At 48 hours post-exposure, no immobilization ofDaphniawas observed in the negative control while the positive control recorded
80 % immobilization. Dimethylacetylsuccinate at concentrations of 15, 24, 38 and 61 mg/L did not immobilize anyDaphniawhereas, at 100 mg/L, 25 % immobilization was observed.

As the test item was not stable in the test medium, the results are expressed as mean measured concentrations.

On the basis of the observations made during this test, the NOEC (which is the highest tested concentration of test item at which no effects onDaphniawere observed = No Observed Effect Concentration) was 27.58 mg/L and LOEC (Lowest Observed Effect Concentration) was
45.56 mg/L.