N-methoxy-1-(2,4,6-trichlorophenyl)propan-2-amine

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Remarks:

The Daphnia Magna reproduction toxicity study was conducted solely to comply with a non-EU national registration requirement, and has been provided here in accordance with REACH, Article 22(1)e.

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 April 2015 to 21 October 2015

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was conducted under a National quality assurance scheme (HJ/T 155-2004) similar to OECD GLP.

Qualifier:

according to guideline

Guideline:

other: HJff 153-2004, The guidelines for the testing of chemicals[S].Beijing: SEP A, 2004.

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: CRC-MEP.The Guidelines for the Testing of Chemicals, Effects on Biotic Systems[M].The 2nd edition. Beijing: China Environment Press. 2013:94-112.

Version / remarks:

2013

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: GB/T 21828-2008, Chemicals-Daphnia magna reproduction test[S]. Beijing: SAC, 2008.

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Version / remarks:

2012

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.20 (Daphnia magna Reproduction Test)

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Ecological Effects Test Guidelines OPPTS 850.1010. Daphnid chronic toxicity test. EPA 712-C-96-120, 1996.

Version / remarks:

1996

Deviations:

no

GLP compliance:

no

Remarks:

The study was conducted under a National quality assurance scheme (HJ/T 155-2004) similar to OECD GLP.

Analytical monitoring:

yes

Details on sampling:

Samples were taken from 0.50 mg/L and 8.00 mg/L test solutions at the beginning (0 d), immediately prior to the first renewal of the test solution (3 d) and weekly thereafter (9, 21 d). 50 mL samples at different concentration were diluted to the 100 mL with acetonitrile and then analysed by the UPLC-PDA method.

Vehicle:

yes

Remarks:

Acetone

Details on test solutions:

The stock solution of 10.0 g/L (adjusted by the purity of 93.2%) was prepared by dissolving 0.1073 g test substance into 10.0 mL acetone.
Stock solution II (I.OD g/L) was prepared by diluting 1.00 mL Stock solution I with acetone to 10.0mL.
Test solutions at different concentrations were prepared by adding the stock solution after the acetone drying at the room temperature and diluting with Elendt M7 and facilitating its dispersion by ultrasonication for 30 min, one control group and solvent control group were tested in parallel.

Test organisms (species):

Daphnia magna

Details on test organisms:

Daphnia magna, not the first brood progeny, were the test species and were 6 to 24 hours old at the start of the test. All organisms used for a particular test originated from cultures established from the same healthy stock of Daphnia. The stock animals were maintained in culture conditions (light, temperature, and medium) similar to those to be used in the test. Lot No: D20150415D.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

221 to 232(mg (CaC03)/L

Test temperature:

20.5 to 21.3°C

pH:

7.01 to 7.66

Dissolved oxygen:

90 to 101%

Nominal and measured concentrations:

nominal concentration of 0.50, 1.00, 2.00, 4.00 and 8.00 mg/L
measured concentration: 0.45 mg/L and 8.15 mg/L (from the lowest and highest nominal concentration)

Details on test conditions:

The concentration of the test substance was demonstrated to be maintained within ±20 percent of the measured initial concentration throughout the test under a 72 h renewal conditions.
When the medium was renewed, a second series of test vessels were prepared and the parent
animals were then be transferred to them by a glass pipette of suitable diameter. All the beakers lids were put on.The volume of medium transferred with the Daphnia was minimized.

Ten animals were individually held at each test concentration and in the control. Treatments were allocated to the test vessels and all subsequent handling of the test vessels was done in a random fashion. The test conditions were maintained as follows:
-Light: 16-hour light and 8-hour dark cycle daily (light intensity: 1000 to 1500 lux);
-Temperature: 20.5°C- 2!.3°C;
-Oxygen concentration: 90% to 101 %; no aerating;
-pH: 7.01 -7.66;
-Feeding: Chiarella vulgaris Beij. was given once every day, 0.1 - 0.2 mg C/Daphnia each time.
-Aeration: None.

In the range-finding test, animals were exposed to concentrations of 1.00 and 10.0 mg/L (nominal concentration). A blank control and solvent control were tested in parallel (dilution medium without test substance).
Based on the results of the range-finding test, nominal concentration of 0.50, 1.00, 2.00, 4.00 and 8.00 mg/L was conducted in the definitive test. Additionally, one blank control and solvent control group were tested in parallel (dilution medium without test substance).

Elendt M7 was used as the control and test media.

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

1.43 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Details on results:

During the test, all animals in the solvent control were alive and the number of live offspring produced per parent animal surviving at the end of the test was 69 - 89. The coefficient of variation around the mean number ofliving offspring produced per parent animal in the control(s) was 8.57% (< 25%). At the end of the test period, no adults died in blank control, solvent control, 0.50 mg/L and 1.00 mg/L treatments. 2, JO and JO adults died in 2.00, 4.00 and 8.00 mg/L treatments. Thus the LOEC for survival of adults was 2.00 mg/L (nominal concentration) and the NOEC was 1.00 mg/L (nominal concentration).
According to Dunnett' s test results of the test data (Dunnett' s test, ANOV A, SPSS 17 .0), the significance level of p was less than 0.05. Thus the reproduction rates in the treatments of 2.00 mg/L and the higher concentration treatments showed statistically significant difference from that of the solvent control (p < 0.05). Thus the LOEC for the Daphnia magna reproduction was nominal 2.00 mg/L (nominal concentration) and the NOEC was 1.00 mg/L (nominal concentration).

Reproduction Inhibition Effects iu Range-finding Test:

Nominal Conc. (mg/L)	Initial No. of Adults	21 d
		Mean Fecundity (No. live offspring per adult)	Inhibition Rate (%)
Blank control	10	70	-
Solvent control	10	67	-
1.00	10	60	11.1
10.0	10	0	100
LOEC (Dunnett's Test)		10.0 mg/L (nominal concentration)
NOEC (Dunnett's Test)		10.0 mg/L (nominal concentration)

Validity criteria fulfilled:

yes

Conclusions:

Under valid semi-static test conditions (72 h-renewal), for Daphnia magna adult survival and reproduction the LOEC was nominal 2.00 mg/L (nominal concentration) and the NOEC was 1.00 mg/L (nominal concentration). 21 d EC50 was 1.43 mg/L (based on the nominal concentration), with 95% confidence limit of 1.33 mg/L to 1.54 mg/L (based on the nominal concentration). For survival of adults the LOEC was 2.00 mg/L (nominal concentration), the NOEC was 1.00 mg/L (nominal concentration).

Executive summary:

Under semi-static conditions of 72h renewal, the effect on the reproductive output of Daphnia magna exposed to the test substance (CA5204A) for 21 days was conducted according to "The guidelines for the testing of chemicals-effects on biotic systems" (HJ/T153); "The Guidelines for the Testing of Chemicals" (the 2nd edition, 2013); Procedure 211 of the Daphnia magna, Reproduction Tesf', OECD Guidelines for Testing of Chemicals, 211, adopted on 2 October 2012.

During the test period, the pH values of the control and test media were between 7.01 and 7 .66, the Dissolved Oxygen (DO) values varied from 90% to 101 % of the air saturation at the test temperature, the hardness values varied from 221 to 232 mg (CaC03)/L, and the temperature of the control and test media was maintained in the range of 20.5°C to 21.3°C. All animals in the control were alive and the number of live offspring produced per parent animal surviving at the end of the test was 69 - 89. The coefficient of variation around the number of living offspring produced per parent animal in the control(s) was 8.57%. Therefore, the study met the acceptability criteria and was considered valid.

The concentration of the test substance (CA5204A) in test medium was determined by UPLC-PDA to represent the stability of the test substance in test period. A linear regression equation was obtained with the mean peak squares (A) vs. the concentration of the CA5204A ( c, mg/L): A=259478c+66636, with good linearity of r2 = 0.9969. The results show that linearity for the concentration range of0.10 mg/L to 5.00 mg/L was good. The minimum detection amount of UPLC-PDA was 0.003 mg/L and the minimum detection concentration for water samples was 0.001 mg/L. The analytical results showed that the concentration of the test substance was consistent in the test medium throughout the test period (deviation within 20% of the initial concentration). Thus a semi-static method of 72 h renewal test procedure was reasonable.

The results showed that under valid semi-static test conditions (72 h-renewal), for Daphnia magna reproduction, the lowest observed effect concentration (LOEC) was 2.00 mg/L (nominal concentration) and the no observed effect concentration (NOEC) was 1.00 mg/L (nominal concentration). 21 d EC50 was 1.43 mg/L (based on the nominal concentration), with 95% confidence limit of 1.33 mg/L to 1.54 mg/L (based on the nominal concentration). For survival of adults the LOEC was 2.00 mg/L (nominal concentration), the NOEC was 1.00 mg/L (nominal concentration).

Description of key information

Under valid semi-static test conditions (72 h-renewal), for Daphnia magna adult survival and reproduction the LOEC was nominal 2.00 mg/L (nominal concentration) and the NOEC was 1.00 mg/L (nominal concentration). 21 d EC50 was 1.43 mg/L (based on the nominal concentration), with 95% confidence limit of 1.33 mg/L to 1.54 mg/L (based on the nominal concentration). For survival of adults the LOEC was 2.00 mg/L (nominal concentration), the NOEC was 1.00 mg/L (nominal concentration).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

1 mg/L