Amines, N-C10-16-alkyltrimethylenedi-, reaction products with chloroacetic acid

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2007-06-28 to 2007-07-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 150, 300, and 600 and 1200 µg a.s. per litre (test start), 150, 300, and 600 µg/L (at 48 h and at test end)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution was continuosly dispensed and diluted with dilution water using the respective equipment to deliver a series of five concentrations at the same time to the corresponding vessels.
- Controls: dilution water only

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Strain: Walbaum
- Source: NRW Landesanstalt für Fischerei (governmental fisheries agency), Albaum, Germany on March 22, 2007 and further bred in the test facility.
- Length at study initiation: 5 ± 1cm
- Feeding during test: Acc. to the guidelines, the fish were not fed during the test.

ACCLIMATION
Not reported, the fish were bred in the test facility.

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

13.5-13.8°C

pH:

7.7–8.3

Dissolved oxygen:

92–101% saturation

Nominal and measured concentrations:

Nominal: 93.5, 188, 375, 750, 1500, 3000, and 6000 μg test item/L, representing 18.8, 37.5, 75, 150, 300, 600, and 1200 μg a.i./L
Mean measured concentrations were in the range of 43 % to 57 % of nominal, independent of the concentration. For the evaluation of the effect concentrations the means of the measured four highest concentrations were used. The means of the treatments were calculated to be 84.9, 136.3, 258.6, and 672.5 μg a.i./L (56.6 %, 45.4 %, 43.1 %, and 56.1 % of nominal).

Details on test conditions:

TEST SYSTEM
- Test vessel: glass aquaria, 42 × 28 × 28 cm, approx. 25 L test solution
- Flow-through rate 2.5 L/h (daily turnover: 5 vol)
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Cu-free tap water, Fraunhofer IME
- Alkalinity: 0.7–1.1 mmol/L
- Total Hardness: 0.7–1.0 mmol/L
- Conductivity: 161.3–183.7
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16:8 h (L:D)
- Light intensity: Not reported

EFFECT PARAMETERS MEASURED: mortality (daily)

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

207.4 µg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL: 157.5–273.2 µg a.i./L

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

351 µg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL: not determined due to mathematical reasons

Details on results:

- Behavioural abnormalities: At concentrations up to and including 84.9 μg a.s./L, no abnormal condition or behavior was observed. At 136.3 μg a.s./L, there was no effect during the first three days of the test. On the last day, the surviving fish exhibited abnormal behavior characterized by slow or uncoordinated swimming at the water surface and/or dark discoloration. At 258.6 μg a.s./L, similar effects already started during the first day. At the highest concentration, toxic effects started immediately.
- Other biological observations: At concentrations up to and including 84.9 μg a.s./L, no mortality was observed. At 136.3 μg a.s./L, there was no effect during the first three days of the test. On the last day, three of ten fish died. At 258.6 μg a.s./L, mortality started on the second day. At the highest concentration all fish died within 24 h.
- Mortality of control: None of the introduced control animals died.

Reported statistics and error estimates:

All statistical calculations were based on mean measured concentrations of the active substance. Calculations were performed with the computer software ToxRat Professional version 2.09 (release 08.11.2006) by ToxRat® Solutions GmbH. When the test results showed a concentration-response relationship, the data were analysed by regression to determine the EC50 including the 95% confidence interval as well as the EC10 using Probit-analysis assuming log-normal distribution of the values.

Sublethal observations / clinical signs:

Table A7.4.1.1-27:Measured concentrations and mean of the four “lead components” C-12 PDA, N-C12 Gly, N’-C12 Gly, and N’-C12 diGly of the test item.

Nominal concentration			Measured concentrations of the test item [µg a.s./L]
Total a.s. [µg/L]	Sum of lead components		Test start	48 h	96 h	Mean
150	98.9	µg/L	66.3	41.0	60.5	56.0
		% nom.	67.0	41.5	61.2	56.6
300	197.8	µg/L	119.1	91.5	59.0	89.9
		% nom.	60.2	46.3	29.8	45.4
600	395.5	µg/L	185.2	198.1	128.1	170.5
		% nom.	46.8	50.1	32.4	43.1
1200	791.0	µg/L	443.3	n.a.	n.a.	443.3
		% nom.	56.0			56.0
	Extrapolated to total a.s.
150	150	µg/L				84.9
		% nom.				56.6
300	300	µg/L				136.3
		% nom.				45.4
600	600	µg/L				258.6
		% nom.				43.1
1200	1200	µg/L				672.5
		% nom.				56.1

 

Table A7.4.1.1-28:Cumulative mortality and clinical signs of intoxication during the test period of 96 h (n per vessel = 10). a.s. = active substance; m.m. = mean measured; Concentrations are given as nominal and mean measured concentrations.

Test substance concentration (µg/L)	Test duration
	8 h	24 h	48 h	72 h	96h
Control	0	0	0	0	0
18.8 n.	0	0	0	0	0
37.5 n.	0	0	0	0	0
75 n.	0	0	0	0	0
150 n. (84.9 m.m.)	0	0	0	0	0
300 n. (136.3 m.m.)	0	0	0	0	3^msd
600 n. (258.6 m.m.)	0	0^md	1^mpd	3^mpsbd	6^msbd
1200 n. (672.5 m.m.)	4^upb	10	10	10	10
Temperature [°C]	SeeTable A7.4.1.1-24
pH	SeeTable A7.4.1.1-26
Oxygen [mg/l]	SeeTable A7.4.1.1-25
More than one surviving fish showed the following clinical signs of intoxication: m = slow movements u = uncoordinated swimming p = unbalanced position s = swimming at the water surface b = lying at the bottom of the test vessel d = dark colour

 

Table A7.4.1.1-29:Effect data,LC values after 48 h and 96 h [µg a.s./L] LC = Lethal concentration; CL = Confidence limits; n.d. = not determined due to mathematical reasons; effect concentrations given as mean measured concentrations.

	48 h [µg/L]	96 h [µg/L]
LC0	84.9	84.9
LC10	253.3	109.5
LC50(95% CL)	351.0 (n.d.)	207.4 (157.5–273.2)

 

 

Validity criteria fulfilled:

yes

Conclusions:

The 96 h LC50 of Amines, N-C10-C16-alkyltrimethylenedi-, reaction products with chloroacetic acid in rainbow trout (Oncorhynchus mykiss) was 207.4 µg a.s./L (CL: 157.5–273.2 µg a.s./L).

Executive summary:

The influence of Amines, N-C10-C16-alkyltrimethylenedi-, reaction products with chloroacetic acid (20% a.i.) on acute toxicity to rainbow trout (Oncorhynchus mykiss) was investigated in accordance with the OECD Guideline 203 and EU method C.1 (92/69/EEC).

The fish were placed in water containing the test item at nominal concentrations of 93.5, 188, 375, 750, 1500, 3000, and 6000 μg test item per litre, representing 18.8, 37.5, 75, 150, 300, 600, and 1200 μg active substance (a.s.) per litre. The test was conducted under flow-through conditions for 96 hours. Effects on survival were determined after 24, 48, 72, and 96 hours. Samples of test solutions were taken at test start, after 48 hours and at test end.

In test media the mean measured concentrations of the test item were in the range of 43 % to 57 % of nominal, independent of the concentration. For the evaluation of the effect concentrations the means of the measured four highest concentrations were used. At nominal concentrations up to and including 84.9μg a.s./L, neither mortality nor abnormal condition or behavior was observed. At higher concentrations, there was a clear concentration and time-effect dependency, starting with 30 % mortality at 136.3μg a.s./L on day four and ending with 100 % mortality at 672.5μg a.s./L on the first day. Clinical signs of intoxication were apparent only in surviving or moribund fish at concentrations causing partial or total mortality.

The 48 h LC50 was 351.0 µg a.s./L, the 96 h LC50 was 207.4 µg a.s./L (CL: 157.5–273.2 µg a.s./L).

Since the validity criteria are fulfilled - except maintenance of test concentrations within 20 % initial, which is however addressed by consideration of measured concentrations - the study is considered to be valid.

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1994-12-05 to 1994-12-09

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: stock solution preparation: 50 mg of the test item was dissolved in 1 L of dilution water, 8.4, 15, 27, 48, 84 and 150 ml were taken and diluted with dilution water to 1.5 litre to obtain test concentrations
- Differential loading: no
- Controls: dilution water only

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Source: commercial hatchery M.B. Ruysbroek B.V., Maassluis, The Netherlands
- Length at study initiation: 2.6 ± 0.2 (SD) cm
- Weight at study initiation: 0.13 ± 0.02 (SD) g
- Method of breeding: not reported
- Maintenance of the brood fish: not reported

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

198 mg CaCO3/L

Test temperature:

24.7°C - 25.5°C

pH:

7.7 - 8.2

Dissolved oxygen:

5.9 - 8.4 mg/L

Nominal and measured concentrations:

Nominal test concentrations: 0 (control), 0.056, 0.10, 0.18, 0.32, 0.56 and and 1.0 mg a.i./L

Details on test conditions:

TEST SYSTEM
- Test vessel: 2 litre all-glass beakers
- Fill volume: 1.5 L
- Aeration: yes
- Renewal rate of test solution: daily
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: synthetic medium prepared by addition of several salts to ground water from a locality near Linschoten (the Netherlands)
- Total organic carbon: 2.1 mg/L
- Culture medium different from test medium: no
- Intervals of water quality measurement: in control and treatment groups at 0 h, 24 h, 48 h, 72 h in new and old media and at 96 h in old media.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 6 h light-8 h dark regime with transition periods of ca. 30 minutes
- Light intensity: not reported

EFFECT PARAMETERS MEASURED: mortality and condition: at 4 h, 24 h, 48 h, 72 h and 96 h

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

0.18 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence interval: 0.12 - 0.18 mg a.i./L

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

0.18 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence interval: 0.18 - 0.26 mg a.i./L

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

0.24 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence interval: 0.18 - 0.32 mg a.i./L

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

0.42 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence interval: 0.32 - 0.56 mg a.i./L

Duration:

4 h

Dose descriptor:

LC50

Effect conc.:

0.75 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence interval: 0.56 - 1.0 mg a.i./L

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.1 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

Details on results
- Behavioural abnormalities: at a concentration of 0.56 mg a.i./l after 4 h: slow swimming, at 0.18 mg/L after 72 h: slow swimming, swimming near bottom, dark colour, after 96 h: disturbed equilibrium
- Mortality of control: no
- Other adverse effects control: no

Reported statistics and error estimates:

The maximum likelihood estimates of the LC50 values were calculated assuming a log-logistic dose-effect relation. Likelihood-ratio confidence intervals were derived from the confidence intervals.

Sublethal observations / clinical signs:

Table 1: Number of surviving fish during the test

Time (h)	Nominal concentrations (mg a.i./L)
	Control	0.056	0.10	0.18	0.32	0.56	1.0
0	10	10	10	10	10	10	10
4	10	10	10	10	10	10	10
24	10	10	10	10	10	10	0
48	10	10	10	10	10	0	0
72	10	10	10	10	0	0	0
96	10	10	10	8	0	0	0

Validity criteria fulfilled:

not specified

Remarks:

No validation criteria were given in the study report. No analytical verification of test concentrations.

Conclusions:

The derived 96 h LC50 value for fresh-water fish species Brachydanio rerio was 0.18 mg a.i./L in a semi-static toxicity test.

Executive summary:

The acute toxicity of Amines, N-C10-C16-alkyltrimethylenedi-, reaction products with chloroacetic acid to the fresh-water zebrafish species Danio rerio was determined as described in the OECD Guideline 203 and according to the OECD principles of Good Laboratory Practice. The test was carried out under semi-static conditions with daily replacement of the test solutions and with 10 fish for the control medium and each concentration. The exposure duration was 96 hours. The nominal concentrations tested were 0, 0.056, 0.10; 0.18, 0.32, 0.56 and 1.0 mg a.i./L. All test solutions were completely clear (visually assessed) throughout the test. The derived 96 h LC50 value was 0.18 mg a.i./L. No chemical analysis was carried out. because a suitable analytical method was not available. Therefore the concentrations quoted in this report refer to the nominal active ingredient of the test substance.

Description of key information

LC50 (96 h)= 207.4 µg a.i./L (mean measured, Oncorhynchus mykiss, OECD Guideline 203 / EU method C.1)

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

207.4 µg/L

Additional information

The influence of Amines, N-C10-C16-alkyltrimethylenedi-, reaction products with chloroacetic acid (20% a.i.) on acute toxicity to rainbow trout (Oncorhynchus mykiss) was investigated in accordance with the OECD Guideline 203 and EU method C.1 (92/69/EEC).

The fish were placed in water containing the test item at nominal concentrations of 93.5, 188, 375, 750, 1500, 3000, and 6000 μg test item per litre, representing 18.8, 37.5, 75, 150, 300, 600, and 1200 μg a.i./L. The test was conducted under flow-through conditions for 96 hours. Effects on survival were determined after 24, 48, 72, and 96 hours. Samples of the four highest test concentrations were taken at test start, after 48 hours and at test end and analysed by HPLC.

In test media the mean measured concentrations of the test item were in the range of 43 % to 57 % of nominal, independent of the concentration. The mean measured concentrations were 84.9, 136.3, 258.6, and 672.5 μg a.i./L. For the evaluation of the effect concentrations the means of the measured four highest concentrations were used. At mean measured concentrations up to and including 84.9 μg a.i./L, neither mortality nor abnormal condition or behavior was observed. At higher concentrations, there was a clear concentration and time-effect dependency, starting with 30 % mortality at 136.3 μg a.i./L on day four and ending with 100 % mortality at 672.5 μg a.i./L on the first day. Clinical signs of intoxication were apparent only in surviving or moribund fish at concentrations causing partial or total mortality. The 48 h LC50 was 351.0 µg a.i./L, the 96 h LC50 was 207.4 µg a.i./L (CL: 157.5–273.2 µg a.i./L).

Since the validity criteria are fulfilled - except maintenance of test concentrations within 20 % initial, which is however addressed by consideration of measured concentrations - the study is considered to be valid.

This result is supported by a second study (OECD 203) with the fresh-water zebrafish species Danio rerio. The test was carried out under semi-static conditions with daily replacement of the test solutions and with 10 fish for the control medium and each concentration. The exposure duration was 96 hours. The nominal concentrations tested were 0, 0.056, 0.10; 0.18, 0.32, 0.56 and 1.0 mg a.i./L. The derived 96 h LC50 value was 0.18 mg a.i./L. No chemical analysis was carried out because a suitable analytical method was not available. Therefore, the concentrations quoted in this report refer to the nominal active ingredient of the test substance.