N,N-bis(2,3-epoxypropyl)aniline

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:

26 November 2012 to 10 January 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study has been performed according to OECD and/or EC guidelines and according to GLP principles

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

2008

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

ISO 6341 (Water quality - Determination of the Inhibition of the Mobility of Daphnia magna Straus (Cladocera, Crustacea))

Version / remarks:

1996

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000.

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline

EC Number:

275-662-9

EC Name:

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline

Cas Number:

71604-74-5

Molecular formula:

C15H19NO4

IUPAC Name:

3-(oxiran-2-ylmethoxy)-N,N-bis(oxiran-2-ylmethyl)aniline

Test material form:

liquid: viscous

Details on test material:

Description: Clear slightly yellow very viscous liquid
Purity/Composition: 100% Mono constituent substance
Test substance storage: In refrigerator (2-8°C) in the dark
Stability under storage conditions: Stable

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Purity/composition correction factor required: No
Volatile: Yes, vapour pressure: <13 Pa at 293K
Stability in water: Unknown
Solubility in water: No

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Singular samples for possible analysis were taken from all concentrations and the control according to the schedule below.

Frequency at t=0 h and t=48 h
Volume 2 ml from the approximate centre of the test vessels
Storage Samples were stored in a freezer until analysis.

At the end of the exposure period, the replicates were pooled at each concentration before sampling.

Additionally, singular reserve samples of 2 ml were taken for possible analysis. If not used, these samples were stored in a freezer for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.

Test solutions

Vehicle:

no

Details on test solutions:

The batch of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline tested was a clear slightly yellow very viscous liquid with a purity of 100% Mono constituent substance and not completely soluble in test medium at the loading rate initially prepared.

A final test was performed based on the results obtained in a preceding combined limit/range-finding test. Preparation of test solutions started with a loading rate of 100 mg/l applying a 1-day period of magnetic stirring followed by a one-hour settlement period. The aqueous Water Soluble Fraction (WSF) was siphoned off and was used as the highest concentration. The lower test concentrations were prepared by subsequent dilutions of the WSF prepared at 100 mg/l. The final test solutions were all clear and colourless. Note that due to volatile nature of the test substance the preparation of test solutions was performed in closed vessels.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions.
Source: In-house laboratory culture with a known history.
Reason for selection: This system has been selected as an internationally accepted invertebrate species.
Validity of batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.
Characteristics: For the test selection of young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Remarks on exposure duration:

none

Post exposure observation period:

no

Test conditions

Hardness:

180 mg/l expressed as CaCO3

Test temperature:

18.5 to 19.7°C

pH:

8.0- 8.2

Dissolved oxygen:

8.9 - 9.4

Salinity:

not measured

Nominal and measured concentrations:

Nominal: 0.10, 0.32,1.0, 3.2, 10, 32 and 100% of the WSF prepared at 100 mg/l.

Measured: Analysis of the samples taken at the start of the test from 10, 32 and 100% of the WSF prepared at 100 mg/l showed that actual concentrations were at the level of nominal, namely 9.8, 32 and 98 mg/l. During the exposure period the measured concentration remained constant (86-88% of initial). Therefore, the effect parameters were expressed in terms of analytically confirmed nominal concentrations.

Details on test conditions:

Test duration: 48 hours
Test type: Static
Test vessels: 50 ml, all-glass, without headspace and air tight closed
Medium: Adjusted ISO medium
Number of daphnids: 20 per concentration
Loading: 5 per vessel containing approximately 50 ml of test solution
Light: 16 hours photoperiod daily
Feeding: No feeding
Aeration: No aeration of the test solutions
Introduction of daphnids: Within 45 minutes after preparation of the highest test solution.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: -

Effect concentrations 2

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

22 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 19-27

Effect concentrations 3

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

18 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 10-32

Details on results:

- Behavioural abnormalities: no
- Observations on body length and weight: no
- Mortality of control: no
- Other adverse effects control: no
- Abnormal responses: no

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50/LC50: 0.38 mg/l with a 95% confidence interval between 0.34 and 0.45 mg/l.

Reported statistics and error estimates:

Calculation of EC50:

The 24-hours EC50-value was calculated from the probits of the percentages of affected daphnids and the logarithms of the corresponding test substance concentrations using the maximum likelihood estimation method (Finney, D.J., 1971: Probit analysis, Cambridge University Press, Cambridge, U.K., 3rd edition).

The 48-hours EC50 could not be determined using the maximum likelihood estimation method with the probits of the percentages of immobile daphnids as function of the logarithms of the corresponding concentrations (Finney, D.J., 1971: Probit analysis, Cambridge University Press, Cambridge, U.K., 3rd edition). This was because there was no concentration between the highest concentration (A) at which 0% immobility and the lowest concentration (B) at which 100% immobility occurred. Instead, the EC50 was calculated as (AB)½, with A and B being limits of the 95% confidence interval.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the present study m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline did not induce acute immobilisation of Daphnia magna at an analytically confirmed nominal concentration of 10 mg/l after 48 hours of exposure (NOEC).

The 48h-EC50 was 18 mg/l based on analytically confirmed nominal concentrations (95% confidence interval between 10 and 32 mg/l).

Executive summary:

The study procedures described in this report were based on the OECD guideline No. 202, 2004. In addition, the procedures were designed to meet the test methods of theCommissionRegulation (EC) No440/2008,Part C.2, 2008, the ISO International Standard 6341, 1996 and the OECD series on testing and assessment number 23, 2000.

 

The batch of m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline tested was a clear slightly yellow very viscous liquid with a purity of 100% Mono constituent substance and not completely soluble in test medium at the loading rate initially prepared.

 

A final test was performed based on the results obtained in a preceding combined limit/range-finding test. Preparation of test solutions started with a loading rate of 100 mg/l applying a 1-day period of magnetic stirring followed by a one-hour settlement period. The aqueous Water Soluble Fraction (WSF) was siphoned off and was used as the highest concentration. The lower test concentrations were prepared by subsequent dilutions of the WSF prepared at 100 mg/l. The final test solutions were all clear and colourless.

 

Twenty daphnia per group (5 per vessel, 4 vessels) were exposed to a control and solutions containing 0.10, 0.32, 1.0, 3.2, 10, 32 and 100% of the WSF prepared at 100 mg/l. The total test period was 48 hours and samples for analysis of actual exposure concentrations were taken at the start and the end of the test period. Due to volatile nature of the test substance, the study was performed in air tight closed vessels with headspace reduced to minimum.

 

The effect parameters were expressed in terms of analytically confirmed nominal concentrations.

 

The study met the acceptability criteria prescribed by the protocol and was considered valid.

 

m-(2,3-epoxypropoxy)-N,N-bis(2,3-epoxypropyl)aniline did not induce acute immobilisation ofDaphnia magnaat an analytically confirmed nominal concentration of 10 mg/l after 48 hours of exposure (NOEC).

 

The 48h-EC₅₀was 18 mg/l based on analytically confirmed nominal concentrations (95% confidence interval between 10 and 100 mg/l).