N,N'-bis(5,5-dimethyl-1,3,2-dioxaphosphinane-2-oxide-2-yl)ethane-1,2-diamine

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:

June 3, 2013 to June 5, 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

OECD Guidelines for Testing of Chemicals, No.202, April 13, 2004, “Daphnia sp., Acute Immobilisation Test”

Deviations:

not specified

Qualifier:

according to guideline

Guideline:

other: Japanese Standard Test Guidelines

Version / remarks:

“Acute swimming immobilization test in Daphnia magna” stipulated in the "Test Methods of the New Chemical Substances etc." (March 31, 2011, Yakushokuhatsu 0331 No.7, Heisei 23.03.29 Seikyoku No.5, Kanpokihatsu No.110331009, partly amended by Yakushokuhatsu 0402 No.1, Heisei 24.03.28 Seikyoku No.2, Kanpokihatsu No. 120402001 on April 2, 2012)

Deviations:

not specified

GLP compliance:

yes

Specific details on test material used for the study:

No further details specified in the study report.

Analytical monitoring:

yes

Details on sampling:

Frequency of measurement: At the start and end of exposure.
Sampling methods: Separately sampled from the preparation vessel at the start of exposure. An aliquot was sampled from the middle depth of each test vessel and mixed at the end of exposure.
Sampling amount: Approximately 10 mL was sampled for all test groups.

Vehicle:

no

Details on test solutions:

Required amounts of the provided substance and the dilution water were mixed to make a 100 mg/L solution. Then the test solution was prepared by stirring for 48 hours in the thermostatic bath that was set to be 20 °C, and then was distributed into test vessels.

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: Daphnia magna (Clone A)
Reason for selection of the species: Use of the species is recommended in the test guidelines.
Supply source Sheffield University in England (Sheffield S10 2UQ, United Kingdom)
Acquisition date: July 9, 1990
Acclimatization: Juveniles born from subcultured adults at Kurume Laboratory were used. Adults from the group (lot) with 100 % of the survival rate which were cultured in the same water quality (dechlorinated tap water) at the same water temperature (20 ± 1 °C) and lighting cycle (16 hours light/8 hours dark) as the test conditions for 17days were used to produce juveniles. Throughout the subculture period, Chlorella vulgaris was given to daphnids once daily for 0.1 to 0.2 mgC (organic carbon content)/daphnids.
Selection of the juveniles: Juveniles of 24 hours or younger of age were used.
Group allocation: Random sampling

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

No post exposure observation period required for the type of study.

Hardness:

Not specified

Test temperature:

Water temperature of the test solution was 19.8 - 20.1 °C.

pH:

the pH was 7.7 - 7.8

Dissolved oxygen:

The dissolved oxygen concentration was 8.6 mg/L

Salinity:

Not applicable

Conductivity:

Not specified

Nominal and measured concentrations:

100 mg/L nominal (as a limit test at the upper concentration limit for used test method)

Details on test conditions:

Test equipment and apparatus
Test vessel: 100 mL glass beaker
Cover: A transparent cover made of plastic
Thermostatic bath: Plastic aquarium, heating/cooling unit (Sato Kogei Co., Ltd., HCA 250)

Test conditions
Exposure system: Static water system (without exchange of the test water)
Exposure period: 48 hours
Test concentration: 100 mg/L (as a limit test at the upper concentration limit for used test method). This test concentration was determined based on the result of the preliminary study.
Control group: Dilution water without the test substance which was treated by the same preparation method for the test solution.
Number of the replicates: 4 replicates/test group
Number of test organisms: 20 organisms/test group (5 organisms/test vessel)
Volume of the test solution: 400 mL/test group (100 mL/test vessel)
Water temperature: 20 ± 1 °C
Dissolved oxygen concentration: More than 3 mg/L (without aeration)
pH adjustment: None
Lighting: Room light, 16-hour light/8-hour dark cycle
Feeding: Food was not given.

Observation and measurement
Condition of the test organisms
Swimming immobilization and abnormality of activity and appearances of test organisms were observed at 24 and 48 hours after the start of exposure. The individual was regarded to be immobilized if it was not able to swim within 15 seconds after gentle agitation of the test vessel.

Condition of the test solution
Condition of the test solution was observed at the start and end of exposure.

Quality of the test solution
Measurement items: Concentration of dissolved oxygen concentration, pH, and water temperature
Frequency of measurement: At the start and at the end of exposure.
Measurement methods: The test solution separately sampled from the preparation vessel was measured at the start of exposure, and one test vessel per each test group was measured at the end of exposure.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Swimming immobilization
No swimming immobilization of test organisms was observed during the exposure period in the test concentration groups. In addition, the swimming immobilization of the control groups was 0% and the validity criteria (not more than 10%) was fulfilled.

Observation results of symptom
During the exposure period, symptom such as swimming immobilization, abnormal behavior or appearance was not observed in test organisms of the test concentration groups. No individual which showed abnormal behavior or appearance (discoloration, trapping at surface of water, etc.,) was observed in the control group during the exposure period and the validity criteria (not more than 10%) was fulfilled.

Observation and measurement results of test solution
Condition of the test solution
The test solution was colorless and transparent at the start of the exposure and the condition was the same at the end of exposure.

Water quality of the test solution
The dissolved oxygen concentration was 8.6 mg/L, the pH was 7.7 - 7.8 and water temperature of the test solution was 19.8 - 20.1 °C. The dissolved oxygen concentration fulfilled the validity criteria (3 mg/L and above at the end of exposure).

Test substance concentration in the test solution
The measured concentration of the test substance in test solution were 100 mg/L at the start of exposure (104 % of the nominal concentration) and 99 mg/L at the end of exposure (99 % of the nominal concentration). These concentrations were maintained within ± 20 % of the nominal concentration.

Results with reference substance (positive control):

Reference substance: potassium dichromate (special grade,
Lot No. HLH7646, Wako Pure Chemical Industries, Ltd.)
Test period: from April 17, 2013 to April 19, 2013
48-hour EC50: 0.28 mg/L
This value was within the determined range of the background data in Kurume Laboratory, which is a range of “mean ± 2 x standard deviation” [mean ± standard deviation: 0.24 ± 0.06 mg/L (n = 83)].

Reported statistics and error estimates:

Not specified

Immobility

Nominal concentration (mg/L)		Immobility (%)
		24 hours		48 hours
		Replicate	Test level	Replicate	Test level
Control	A	0	0	0	0
	B	0		0
	C	0		0
	D	0		0
100	A	0	0	0	0
	B	0		0
	C	0		0
	D	0		0

 

Observed abnormal response

Nominal concentration (mg/L)	Observed abnormal response
	24 hours		48 hours
	Immobilization	Other symptom	Immobilization	Other symptom
Control	-	-	-	-
100	-	-	-	-

 

Condition of test solutions

Nominal concentration (mg/L)	Dissolved oxygen concentration (mg/L)		pH		Temperature (°C)
	At the start	At the end	At the start	At the end	At the start	At the end
Control	8.6	8.6	7.8	7.7	19.8	20.1
100	8.6	8.6	7.8	7.7	19.8	20.1

 

Measured concentrations of test item in test solutions

Nominal concentration (mg/L)	Measured concentration (mg/L) (Percentage of measured concentration versus nominal concentration %)
	At the start	At the end	Geometric mean
Control	n.d.	n.d.
100	100 (104)	99 (99)	100 (101)

n.d.: <20.0 mg/L

Validity criteria fulfilled:

yes

Conclusions:

Both 24 and 48-hour EC50 of the test substance to Daphnia magna were > 100 mg/L.

Executive summary:

Test organism: Daphnia magna

Dilution water: Dechlorinated tap water

Test groups: 100 mg/L group and a control group

Preparation of test solutions: Prepared by dissolving the provided substance in the dilution water and then stirring for 48hours.

Exposure system: Static water system

Exposure period: 48 hours

Number of replicates: 4 replicates per test group

Number of test organisms: 20 organisms per test group (5 organisms per test vessel)

Volume of the test solution: 400 mL per test group (100 mL per test vessel)

Water temperature: 19.8 – 20.1 °C

Lighting: Room light, 16-hour light/8-hour dark cycle

Feeding: None

Aeration: None

Analysis of the test substance in test solutions: HPLC method (at the start and end of exposure)

 

The test was carried out as the limit test to confirm the presence or absence of the effect of the test substance to the test organism at the upper concentration limit (100 mg/L) specified in used test method. As a result, there is no effect to the test organisms and therefore it is considered that the test substance causes no acute effect to the test organisms at concentrations close to its solubility.

The test substance concentration in the test solution was maintained ±20% of the nominal concentration, and the environmental conditions were within the proper range for the test.

Therefore, it was considered that this study was in accordance with this test method.

 

Both 24 and 48-hour EC50 of the test substance to Daphnia magna were > 100 mg/L.

Description of key information

48-hour EC50 of the test substance to Daphnia magna: > 100 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

The test was carried out as the limit test to confirm the presence or absence of the effect of the test substance to the test organism at the upper concentration limit (100 mg/L) specified in used test method. As a result, there is no effect to the test organisms and therefore it is considered that the test substance causes no acute effect to the test organisms at concentrations close to its solubility.

Both 24 and 48-hour EC50 of the test substance to Daphnia magna were > 100 mg/L.