Amidation products of C16-18 (even numbered), C18 unsaturated fatty acids esters with 1,1'-iminodipropan-2-ol

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

Jun. 15,2016 to Jul. 15,2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Version / remarks:

Guidelines for Testing of Chemicals, Procedure 207, Earthwonn, Acute Toxicity Tests. Paris:OECD, 1984.

Deviations:

yes

Remarks:

See "Any other information" for details

Qualifier:

according to guideline

Guideline:

EU Method C.8 (Toxicity for Earthworms: Artificial Soil Test)

Version / remarks:

EC Directive 2001/59, C.8. Toxicity for earthworms artificial soil test. 0. J. L142, 2008:533-537.

Deviations:

yes

Remarks:

See "Any other information" for details

Qualifier:

according to guideline

Guideline:

other: HJ/T 153-2004

Version / remarks:

HJ/T 153-2004, the guidelines for the testing of chemicals. Beijing: State environmental protection administration of China, 2004.

Deviations:

yes

Remarks:

See "Any other information" for details

Qualifier:

according to guideline

Guideline:

other: GB/T 21809-2008

Version / remarks:

GB/T 21809-2008, Chemicals- Earthworm, Acute Toxicity Tests [S]. Beijing: Standardization Administration Committee of the PR China, 2008.

Deviations:

yes

Remarks:

See "Any other information" for details

Qualifier:

according to guideline

Guideline:

other: The Guidelines for the Testing of Chemicals-Effects on Biotic Systems

Version / remarks:

Chemical Registration Center of MEP. The Guidelines for the Testing of Chemicals-Effects on Biotic Systems. The second edition. Beijing: China Environmental Science Press. 2013: 58-62.

Deviations:

not specified

GLP compliance:

yes

Specific details on test material used for the study:

Lot No.: RC-1045

Analytical monitoring:

not specified

Details on sampling:

Not specified

Vehicle:

no

Details on preparation and application of test substrate:

Preparation of the Test SubstrateThe test substance was directly added into the soil. A total amount of 10 g mixture of the test substance and quartz sand was prepared firstly and then mixed with 550 g dry weight of soil for each test container.The medium was thoroughly mixed and deionised water was added finally to give an overall moisture content of about 35 per cent of the dry weight. Moisture content was determined by weighing and then drying a small sample at 105°C and re-weighing.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

Test OrganismEisenia foetida was adopted as the test species. The worms were obtained from an earthworm nursery of Nanjing city in China.Name: Nanjing Earthworm NurseryAddress: Pudong community, Changlu Street, Dachang District, NanjingAcclimation of the Test WormsThe earthworms newly obtained in the lab were released from quarantine after 14 days and then this batch was pre-cultured in soil with cow dung for 104 days (Batch No.: E20 160318E). Before the test, the worms were acclimated in test soil for another 24 h. The worms were then washed quickly with distilled water and surplus water absorbed on filter paper before use. Worms for limit test were adult (two months old with clitellum) with an individual weight of 378 mg- 428 mg.

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

yes

Total exposure duration:

14 d

Post exposure observation period:

No post exposure observation period specified in the study report.

Test temperature:

test temperature was 19.8 °C to 20.3 °C

pH:

pH from 6.13 to 6.36

Moisture:

humidity of 79% to 81%

Details on test conditions:

Range-finding TestA pre-test was performed with test substance concentrations of 100, 500 and 1000 mg/kg artificial soil (dry weight) to determine the range of concentrations to be tested in the study.Simultaneously one blank control was established under the same conditions.One test batch for each concentration and control with 10 individuals each was used. The test worms (10 worms each) were firstly placed on the surface of test substrate (dry weight: 560 g each; Moisture content: 35% dry soil weight). After having been sealed with plastic films, containers were put into the chambers.The test duration was 14 days, and the test temperature was maintained at 20.1 °C to 20.7 °C with the humidity of 79% to 81% and continuous light (intensity of 400 lux to 800 lux).Limit TestBased on the results of the range-finding test, a Limit Test was undertaken with test substance concentration of 1000 mg/kg (dry weight of soil) and a blank control. Four test batches for each concentration (1000 mg/kg (dry weight of soil)) and control, each with l 0 individuals were used.The test worms (10 worms each) were firstly placed on the surface of test substrate (dry weight:560 g each; Moisture content: 35% dry soil weight). After having been sealed with plastic films, containers were put into the chambers.The test duration was 14 days, and the test temperature was 19.8 °C to 20.3 °C with the humidity of 79% to 81% and continuous light (intensity of 400 lux to 800 lux).ObservationsOn the 7th day of exposure, the mortality was assessed by emptying test medium onto a stainless plate, sorting worms from the medium and testing their reaction to a mechanical stimulus at the front end. Worms were classified as dead when they do not respond to a gentle mechanical stimulus to the front end. Due to rapid decomposition under test conditions, missing earthworms were also considered dead. At the completion of this assessment, soils were replaced in the test container and the surviving earthworms were replaced on the same test substrate surface for the 14th day assessment.In addition, the number and type of behavioural and other pathological abnormalities (e.g. lethargic or rigid worms) were determined during each mortality assessment on 7th d and 14th d.The pH value was determined according to "Determination of pH in Soil" (NY/T 1377-2007).The moisture content was determined by weighing and then drying a small sample at 105 °C and re-weighing (HJ 613-2011).The worms average weights were weighed on Day 0 prior to assigning to the each test containers of both control and treatment groups. The worms were washed prior to weighing (with deionised water) and the excess water removed by placing the worms briefly on filter paper. The average weights of the surviving worms were weighted with the same methods on day 14.

Nominal and measured concentrations:

Nominal concentration: 1000 mg/kg (dry weight of soil)

Reference substance (positive control):

yes

Remarks:

2-Chloracetamide, CAS: 79-07-2; purity: 99.96%;

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

LC0

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Details on results:

Test ConditionsDuring this test, artificial soils was used with the moisture content of the test medium controlled in range of 27 % to 35 % with pH from 6.13 to 6.36. The test conditions were maintained as follows: temperature of 19.8 °C to 20.3 °C, humidity of 79% to 81% and continuous light with intensity of 400 lux to 800 lux. No unusual circumstances that might have affected the integrity and quality of the study were observed. Mortality was not observed in the control at the end of the study. Thus, the test system was considered to be valid.Earthworm MortalityNo dead worms were observed in treatments and control. The results of the Range-finding test and the Limit Test both showed that the 7 d and 14 d-LC50 to Eisenia foetida in artificial soil were greater than 1000 mg/kg soil dry weight.Behavioural EffectsAt the beginning of the administration, test worms soon disappeared into the test soil medium.During the test, all earthworms remained in the soil.

Results with reference substance (positive control):

Reference controlThe 7-day and 14-day LC50 values of a reference substance (2-chloracetamide) were determined 3 times a year to assure that the laboratory test conditions were adequate and had not changed significantly. The 7-day LC50 and 14-day LC50 for reference substance were 35.1 mg/kg and 25.1 mg/kg, respectively.

Reported statistics and error estimates:

The LC50 and 95% confidence limits can be used to calculate by Trimmed Spearman-Karber Method (Version 1.5, USEPA). In this test, no statistical method was adopted due to the low toxicity for the test substance to the test earthworm.

pH and Moisture Content of the Artificial Soil

Concentration applied to soil (mg/kg (dry soil))	pH		Moisture content^1(%)
	0 d	14 d	0 d	14 d
0	6.30	6.17	35	30
0	6.18	6.24	25	28
0	6.31	6.13	35	27
0	6.27	6.27	35	29
1000	6.23	6.31	35	29
1000	6.32	6.36	35	30
1000	6.19	6.28	35	28
1000	6.14	6.19	35	28
Min	6.14	6.13	35	27
Max	6.32	6.36	35	30

¹Note: Moisture content is then determined by drying a small sample at 105°C and re-weighing.

 

The Average Weights of the Worms at Start and End of Exposure

Concentration applied to soil (mg/kg (dry soil))	0 d average weights (mg)	14 d average weights (mg)
0	413	348
0	378	306
0	428	366
0	396	349
1000	389	335
1000	425	368
1000	398	335
1000	403	353
Min	378	306
Max	428	368

 

Acute Toxicity of 2-chloracetamine toEisenia foetida

Concentration applied to soil (mg/kg (day soil))	Number of the test worms	Number of the dead
		7 d	14 d
0	10	0	0
0	10	0	0
0	10	0	0
0	10	0	0
10	10	0	1
10	10	1	2
10	10	1	1
10	10	1	1
20	10	2	3
20	10	1	2
20	10	1	4
20	10	3	5
40	10	6	7
40	10	5	8
40	10	6	8
40	10	5	7
80	10	10	10
80	10	10	10
80	10	9	10
80	10	9	10
100	10	10	10
100	10	10	10
100	10	10	10
100	10	10	10
LC50 (mg/kg (dry soil))		35.1	25.1
95% confidence limit (mg/kg (dry soil))		29.7~41.4	19.8~30.1

 

Number of Dead Earthworms in the Range-finding Test

Concentration applied to soil (mg/kg (dry soil))	Number of the test worms	Number of the dead
		7 d	14 d
0	10	0	0
100	10	0	0
500	10	0	0
1000	10	0	0
LC50 (mg/kg (dry soil))		>1000	>1000

 

Number of Dead Earthworms in the Limit Test

Concentration applied to soil (mg/kg (dry soil))	Number of the test worms	Number of the dead
		7 d	14 d
0	10	0	0
0	10	0	0
0	10	0	0
0	10	0	0
1000	10	0	0
1000	10	0	0
1000	10	0	0
1000	10	0	0
LC50 (mg/kg (dry soil))		>1000	>1000

 

Validity criteria fulfilled:

yes

Conclusions:

The result indicated that the 7 d LC50 and 14 d LC50 of the test substance (MLA-3202) in the artificial soil were both greater than 1000 mg/kg dry weight soil. The highest concentration causing no mortality for 7 d (7-d LC0) and 14 d (14-d LC0) were 1000 mg/kg dry weight soil.

Executive summary:

The test was conducted to determine the acute toxicity of test substance (MLA-3202) to earthworm according to "The guidelines for the testing of chemicals" (HJ/T 153-2004) and "The Guidelines for the Testing of Chemicals-Effects on Biotic Systems" (the 2nd edition. Beijing: China Environment Press. 2013); while with reference to GB/T21809-2008 and Procedure 207 of the 'Guidelines for Testing of Chemicals' of the OECD: "Earthworm, Acute Toxicity Tests" (1984) etc.

 

During this test, artificial soils were used with the moisture content of the test medium controlled in range of 27% to 35% with pH from 6.13 to 6.36. The test conditions were maintained as follows: temperature of 19.8 °C to 20.3 °C, humidity of 79% to 81% and continuous light with intensity of 400 lux to 800 lux. Mortality was not observed in the control at the end of the study.

The 7-day LC50 and 14-day LC50 for reference substance were 35.1 mg/kg and 25.1 mg/kg, respectively. Thus, the test system was considered to be valid.

 

Based on the result of range-finding test and limit test, the 7-day and 14-day LC50 of the test substance (MLA-3202) in the artificial soil were both determined to be greater than 1000 mg/kg dry weight soil. The highest concentration causing no mortality (7-day and 14-day LC0) was 1000 mg/kg dry weight soil.

Description of key information

Key value determined in GLP accredited laboratory study according to "The guidelines for the testing of chemicals" (HJ/T 153-2004) and "The Guidelines for the Testing of Chemicals-Effects on Biotic Systems" (the 2nd edition. Beijing: China Environment Press. 2013); while with reference to GB/T21809-2008 and Procedure 207 of the 'Guidelines for Testing of Chemicals' of the OECD: "Earthworm, Acute Toxicity Tests" (1984) etc.

Key value for chemical safety assessment

Short-term EC50 or LC50 for soil macroorganisms:

1 000 mg/kg soil dw

Additional information

During this test, artificial soils were used with the moisture content of the test medium controlled in range of 27% to 35% with pH from 6.13 to 6.36. The test conditions were maintained as follows: temperature of 19.8 °C to 20.3 °C, humidity of 79% to 81% and continuous light with intensity of 400 lux to 800 lux. Mortality was not observed in the control at the end of the study.

The 7-day LC50 and 14-day LC50 for reference substance were 35.1 mg/kg and 25.1 mg/kg, respectively. Thus, the test system was considered to be valid.

 

Based on the result of range-finding test and limit test, the 7-day and 14-day LC50 of the test substance (MLA-3202) in the artificial soil were both determined to be greater than 1000 mg/kg dry weight soil. The highest concentration causing no mortality (7-day and 14-day LC0) was 1000 mg/kg dry weight soil.