Phenol, 4,4′-(1-methylethylidene)bis-, polymer with 2-(chloromethyl)oxirane and N1,N1-diethyl-1,3-propanediamine

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:

23 March 2018 - 18 September 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

13 April 2004

Deviations:

no

Test guideline 2

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

Version / remarks:

2000

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

DEPDA adduct

Cas Number:

123739-10-6

IUPAC Name:

DEPDA adduct

Test material form:

solid

Details on test material:

- Physical appearance: dark amber solid
- Storage conditions: at room temperature

Specific details on test material used for the study:

- No correction factor for purity was required and therefore not applied.
- The test material is irritant/corrosive.
- Stability at higher temperatures: yes, overnight at maximum 45°C or for maximum 1 hour at maximum 80°C.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

For determination of test concentrations in the final test, single samples for possible analysis were taken from additional vessels without organisms from all test concentrations and the control according to the schedule below.
Frequency: at t=0 h and t=48 h
Volume: 20 mL from the approximate centre of the test vessels
Storage: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the test facility

Test solutions

Vehicle:

yes

Remarks:

Tetrahydrofuraan

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION:
Stock solutions were prepared in tetrahydrofuran (THF, VWR International, Batch 1820907) at nominal concentrations ranging between 3.0 and 30 mg/mL. Preparation of the stock solutions started with the highest stock solution of 30 mg/mL, applying 10 minutes of ultrasonic treatment to ensure maximum dissolution of the test item in medium. The lower stock solutions were prepared separately by proportional dilution of the highest stock solution in THF. For the preparation of each test concentration, 100 µL of a stock solution was added to 1 L of test medium resulting in test concentrations that were a factor of 10,000 lower than the stock concentrations.
- Evidence of undissolved material: stock solutions appeared to be light yellow to brown with increasing concentrations, and that small undissolved particles were observed in the highest stock solution. Test solutions were clear and colorless at the end of the preparation procedure.
- Controls: Blank-control (test medium without test item or other additives) and solvent-control (test medium without test item but with 0.1 mL THF/L)

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Straus, 1820
- Source: in-house laboratory culture with a known history. At least third generation, obtained by a cyclical parthenogenesis under specified breeding conditions.
- Age of parental stock: 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.
- Age at study initiation: < 24 hours, from parental daphnids of more than two weeks old.
- Feeding during test: no

ACCLIMATION
- Acclimation period: no

BREEDING:
- Method: each batch was started with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium (M7) in an all-glass culture vessel. Cultures were max. 4 weeks old. After 7 days of cultivation half of the medium was renewed twice a week.
- Feeding: daily, a suspension of fresh water algae.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

180 mg/L

Test temperature:

20°C

pH:

At t=0 h: 8.4-8.7
At t=48 h: 8.9-9.0

Dissolved oxygen:

At t=0 h: 7.9-8.0 mg/L
At t=48 h: 8.1 mg/L

Nominal and measured concentrations:

Nominal concentration: 0, 0 (THF), 0.30, 0.54, 0.96, 1.7 and 3.0 mg/L
Measured concentrations at both t=0 and t=48 h were below the limit of quantification for 0.30 and 0.54 mg/L and at t=48 only for 0.96 mg/L. Measured concentrations for 1.7 and 3.0 at t=48 were 1.62 and 2.54 mg/L respectively (for details see table 1).

Details on test conditions:

TEST SYSTEM
- Test vessel: 60 mL, all-glass
- Fill volume: 50 mL
- Aeration: no
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Standard medium used: yes, according to OECD 202
- Source/preparation of dilution water: tap water purified by reverse osmosis
- Culture medium different from test medium: yes, M7 was used as culture medium

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: daily photoperiod of 16 hours

EFFECT PARAMETERS MEASURED: mobility at 24 and 48 hours.
- Additional measurements: pH and dissolved oxygen at the beginning and at the end of the test, in the limit test concentration and the control. Temperature of the medium: continuously in a temperature control vessel.

COMBINED LIMIT/RANGE-FINDING STUDY: daphnids were exposed to a control and a SS prepared at a loading rate of 100 mg/L (four replicates of five daphnids) and to 1.0 and 10% of the SS (two replicates of five daphnids). At 10% of the SS, 50% and 80% of the daphnids were immobilised after 24 and 48 hours, respectively. At 100% of the SS, 100% of the daphnids were immobilised after 24 hours.
- In the combined limit/range-finding study, the test item was not spiked with THF.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (July 2018)

Results and discussion

Effect concentrations

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.6 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Remarks on result:

other: 95%-CI: 0.52 - 0.70 mg/L

Details on results:

Results of test sample analysis (see table 1): at the end of the test, the measured concentrations remained stable at the two highest concentrations tested (i.e., were 79-82% of initially measured), and decreased below the limit of quantification at 0.96 mg/L. It should be noted that a response was observed in both the controls. The cause is unknown. It was, however, considered negligible and to have no influence on the study. Based on these results, the effect parameters were based on the arithmic mean of measured concentrations at t=24 h and t=48 h (excluding the lowest test concentration).

Results of the final test:
- No immobility was observed in the controls and the lowest test concentration while a dose-related increase of immobility was observed at 0.54 mg/L and higher, reaching ≥ 90% immobility at the three highest test concentrations at the end of the test.
- Abnormal observations: no
- Mortality of control: no
- The experimental conditions remained within the limits prescribed by the study plan and were according to the OECD guideline.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Test concentrations: 0, 0.10, 0.18, 0.32, 0.56, 1.0 and 1.8 mg/L
- 48h-EC50: 0.59 mg/L, 95%-confidence interval: 0.53-0.72 mg/L
- Other: results fell within the historical range of the test facility.

Any other information on results incl. tables

Table 1 Measured concentrations during the final test

Time of sampling [hours]	Nominal concentration [mg/L]	Analyzed concentration [mg/L]	Relative to nominal [%]	Relative to initial [%]
n.a.	Blank
	blank
	blank
0	0	< 1	n.a.
	0-THF	< 1	n.a.
	0.30	< 1	n.a.
	0.54	< 1	n.a.
	0.96	1.38	144
	1.7	1.97	116
	3.0	3.22	107
48	0	< 1	n.a.	n.a.
	0-THF	< 1	n.a.	n.a.
	0.30	< 1	n.a.	n.a.
	0.54	< 1	n.a.	n.a.
	0.96	< 1	n.a.	n.a.
	1.7	1.62	95	82
	3.0	2.54	85	79

n.a. Not applicable.

Table 2 Number of introduced daphnids and incidence of immobility in the final test

Time (h)	Replicate	INCA 460: DEPDA Adduct Nominal conc.(mg/L)
		Blank Control	THF-Control	0.30	0.54 (0.50)	0.96 (0.83)	1.7 (1.8)	3.0 (2.9)
0	A	5	5	5	5	5	5	5
	B	5	5	5	5	5	5	5
	C	5	5	5	5	5	5	5
	D	5	5	5	5	5	5	5
	Total introduced	20	20	20	20	20	20	20
24	A	0	0	0	0	3	3	3
	B	0	0	0	1	4	3	1
	C	0	0	0	0	1	2	3
	D	0	0	0	0	3	5	4
	Total immobilised	0	0	0	1	11	13	11
	Effect %	0	0	0	5	55	65	55
48	A	0	0	0	0	5	4	5
	B	0	0	0	4	5	5	5
	C	0	0	0	1	4	5	5
	D	0	0	0	1	4	5	5
	Total immobilised	0	0	0	6	18	19	20
	Effect %	0	0	0	30	90	95	100

( ) - between brackets, average exposure concentrations (mg/L)

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

1). In the control, no daphnids became immobilised or showed other signs of disease or stress. 2). The oxygen concentration at the end of the test was ≥ 3 mg/L in control and test vessels (i.e., ≥ 8.9 mg/L).

Conclusions:

The 48h-EC50 for Daphnia magna exposed to INCA 460: DEPDA Adduct was determined to be 0.60 mg/L based on average exposure concentrations (95% confidence interval between 0.52 and 0.70 mg/L).