Benzoic acid, 2-[[(2'-cyano[1,1'-biphenyl]-4-yl)methyl]amino]-3-nitro-, methyl ester

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August 19, 2004 - September 29, 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Name: CAN3
Batch No.: February 2004
Active Ingredient(s) l Purity: > 99% 2-[N-(2'-cianobiphenyl-4-il)-methylanlino]-3-methyhritrobenzoate
Certificate of Analysis: not available
Aggregate State at RT: solid (according to IBACON personnel)
Colour: yellow
Solubility: in water: 0.1 µg/mL
Stability: (i) technical product: see expiry date; (ii) in water: stable under test conditions
Expiry Date: February 2009
Storage: in original container, at room temperature, in the dark

Analytical monitoring:

no

Details on sampling:

- Concentrations: 10, 18, 32, 56 and 100 mg test item/L and a control. In the control, test water was used without addition of the test item.
- Sampling method: The test item was not well soluble in test water. To avoid physical effects of undissolved test item on Daphnia no concentrations above the solubility limit of the test item in test water were tested. Therefore, a stock suspension of 100 mg test item/L was prepared by suspending 80 mg test item in 800 mL test water for preparing the test concentrations. The stock suspension was stirred for 24 hours in the dark to
dissolve as much test item as possible. Then, the undissolved test item was separated by filtration (0.45 µm cellulose nitrate filter) and the required test concentrations were prepared. The test concentrations were prepared just before introduction of the Daphnia (= start ofthe test).

Vehicle:

yes

Details on test solutions:

Measurement of pH, dissolved Oxygen and Water Temperature:
The pH-values and dissolved oxygen concentrations were determined at test start and end in the test media of all test
concentrations and the control. The water temperature was measured in one control beaker at the start and the end of the test.

Behaviour of the Test Item in Test Water:
The behaviour of the test item in test water was determined at the start of the test, after 24 and 48 hours test duration in all test concentrations.

Test organisms (species):

Daphnia magna

Details on test organisms:

Species:
Daphnia magna (Straus), clone 5

Age at Test Start:
6.75 to 22.5 hours old

Sex:
female

Origin:
supplied 1997 by the Umweltbundesamt, Institut fur Wasser-, Boden- und Lufthygiene, Berlin, Germany

Breeding Conditions:
The Daphnia were bred in the laboratories of IBACON under similar temperature and light conditions as in the test, and in
reconstituted water of a similar quality regarding to pH, components of the main ions and total hardness as the test water used in the test. The test organisms were not first brood progeny. The Daphnia of the stock culture were fed at least each working day with green algae (Desmodesmus subspicatus) freshly grown in the laboratories of IBACON.

Toxic Standard:
For the evaluation of the quality of the Daphnia clone and the experimental conditions the substance potassium dichromate
p.A. is tested at least twice a year to demonstrate satisfactory test conditions.
Result of the latest reference test:
48-hour EC 50: 2.12 mg potassium dichromate/L
(confidence limits: 1.02-4.40 mg/L)
Study completion date: February 19, 2004

Acclimatisation:
for 6.75 hours under test conditions.

Test type:

static

Water media type:

other: Reconstituted Water

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

The immobility or mortality of the Daphnia was determined by visual controls after 24 and 48 hours. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile.

Hardness:

Water Hardness: 2.5 mmol/L (= 250.0 mg/L) as CaCO3

Test temperature:

20-21 °C

pH:

7.7- 8.1

Dissolved oxygen:

Oxygen concentrations were determined at test start and end in the test media of all test concentrations and the control. At the end of the test the dissolved oxygen concentration in the test media should be ≥ 2 mg O2/L in the control and test
vessels.

Salinity:

In deionised water analytical grade salts were added to following nominal concentrations:
CaCl2 x 2H2O 2.0 mmol/L (= 294.0 mg/L)
MgSO4 x 7H2O 0.5 mmol/L (= 123.0 mg/L)
NaHCO3 0.75 mmol/L (= 65.0 mg/L)
KCl 0.075 mmol/L (= 5.8 mg/L)
Water Hardness 2.5 mmol/L (= 250.0 mg/L) as CaCO3
Alkalinity 0.8 mmol/L
Ratio of Ca: Mg = 4 : 1 (based on molarity)
Na : K = 10 : 1 (based on molarity)

Conductivity:

conductivity < 5 µS/cm

Nominal and measured concentrations:

Nominal concentrations: 10, 18, 32,56 and 100 mg test iterm/L

Details on test conditions:

Type and Size: glass beakers of 100 mL volume with 80 mL test medium
Identification: Each test unit was uniquely identified with study number, treatment and replicate number.

Test Conditions
Test Environment: controlled environment room
Water Temperature: 20-21 °C
pH-Value of Test Water: 7.7- 8.1
Light Regime: 16 h light : 8 h dark
Light Intensity: 410- 750 lux

Introduction of Individuals: 20 Daphnia per control and test concentration, divided into 4 groups of 5 animals, each group in 80 mL test medium.
Feeding: The test organisms were not fed during the test.
Exposure Time: 48 hours.
Test Procedure: The test item is considered as stable in test water, so a static test system was chosen.

Reference substance (positive control):

yes

Remarks:

For the evaluation of the quality of the Daphnia clone and the experimental conditions the substance potassium dichromate p.A. is tested at least twice a year to demonstrate satisfactory test conditions.

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: After 48 hours in the control and at all test concentrations no mortality or immobilization of the test animals was observed.

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Remarks on result:

other: After 48 hours in the control and at all test concentrations no mortality or immobilization of the test animals was observed.

Details on results:

Validity Criteria of the Study
Control:
The experiment is valid because the mortality of Daphnia magna in the control did not exceed 10 % and no Daphnia were trapped at the water surface.

Dissolved Oxygen Concentration:
At the end of the test the dissolved oxygen concentration in the test media was ≥ 2 mg O2/L in the control and test vessels.

Signs of intoxication after 24 hours:
After 24 hours in the control and at all test concentrations no mortality or immobilization of the test animals was observed.

Signs of intoxication after 48 hours:
After 48 hours in the control and at all test concentrations no mortality or immobilization of the test animals was observed.

pH-Values: 7.7- 8.1
Dissolved Oxygen Concentrations: at least 5.7 mg/L or higher
Water Temperature: 20-21 °C
Behaviour of the Test Item: the test water was coloured by the test item at all test concentrations during the entire study.

Validity criteria fulfilled:

yes

Conclusions:

After 48 hours in the control and at all test concentrations no mortality or immobilization of the test animals was observed.

Executive summary:

The EC 0, 50 and 100 could not be determined, because there was no effect of the test item at all concentrations. The EC 50 is estimated to be higher than 100 mg test item/L. The 48-hour NOEC of CAN3 to Daphnia magna was determined to be at least nominai 100 mg test item/L. The NOEC might even be higher than this concentration, but concentrations in excess of nominal 100 mg test item/L have not been tested. The 48-hour LOEC were clearly higher than nominal 100 mg test item/L. This value could not be quantified due to the absence of toxicity of CAN3 at all tested concentrations.

Description of key information

The EC 0, 50 and 100 could not be determined, because there was no effect of the test item at all concentrations. The EC 50 is estimated to be higher than 100 mg test item/L. The 48-hour NOEC of CAN3 to Daphnia magna was determined to be at least nominai 100 mg test item/L. The NOEC might even be higher than this concentration, but concentrations in excess of nominal 100 mg test item/L have not been tested. The 48-hour LOEC were clearly higher than nominal 100 mg test item/L. This value could not be quantified due to the absence of toxicity of CAN3 at all tested concentrations.