Dodecan-1-ol, ethoxylated

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All the test concentrations were analytically determined to check the stability in test medium through out the exposure period using LC-MS/MS
- Sample storage conditions before analysis: test samples were analysed immediately after sampling

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test medium was reconstituted water, as described in the OECD test guideline. The conductivity of the deionised water used in the preparation of the test medium was 0.054 µS/cm during range finding and definitive test.
- Controls: reconstituted water as per OECD 203.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Not applicable
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): Not applicable
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): Not applicable
- Other relevant information: Not applicable

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebrafish
- Source: Pet bonding, Mahalakshmipuram Bangalore
- Length at study initiation (length definition, mean, range and SD): 1.4 to 1.8 cm
- Weight at study initiation (mean and range, SD): 2.3856 (group body weight)


ACCLIMATION
- Acclimation period: 7 days
- Acclimation conditions (same as test or not): same as test, details mentioned in below table
- Feeding frequency during acclimation: 3 times per week
- Health during acclimation (any mortality observed): no mortality


FEEDING DURING TEST
- Food type: Not applicable
- Amount: Not applicable
- Frequency: Not applicable

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

250 mg/L as CaCO3

Test temperature:

22.1 to 22.3°C

pH:

7.65 to 7.78

Dissolved oxygen:

86 to 95 % air saturation

Salinity:

Not applicable

Conductivity:

0.054 µS/cm

Nominal and measured concentrations:

0.5, 1.0, 2.0, 4.0, 8.0 and 16 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: aquarium
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: glass, 20L
- Aeration: NO
- Type of flow-through (e.g. peristaltic or proportional diluter): no applicable
- Renewal rate of test solution (frequency/flow rate): Not applicable
- No. of organisms per vessel: 10 per vessel
- No. of vessels per concentration (replicates): 10/ concentration
- No. of vessels per control (replicates): no replicates
- No. of vessels per vehicle control (replicates): no replicates
- Biomass loading rate: 0.1193 g/L

OTHER TEST CONDITIONS
- Photoperiod: 16 h photoperiod daily


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Mortality was observed at 2±0.5 h, 5 ± 1 h, 24 ± 2, 30 ± 2, 48 ± 2, 54 ± 2, 72 ± 2, 78 ± 2 and 96 ± 2

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study :
The range finding test was carried out with test concentrations of 0.001, 0.01, 0.1, 1, 10 and 100 mg/L of the test medium along with a negative control in a semi-static mode (renewal of test solution at every 24 h interval). Five fish were used for each test solution with single replicate per test concentration.
- Test concentrations: 0.5 (G2), 1.0 (G3), 2.0 (G4), 4.0 (G5), 8.0 (G6) and 16.0 (G7) mg/L
- Results used to determine the conditions for the definitive study: Based on the mortaily observed after exposure period the concentrations and method was decided

Reference substance (positive control):

not required

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

2.427 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: fiducial limits at 95% ranging from 1.6769 to 3.5123

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

3.76 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

3.354 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: fiducial limits at 95% ranging from 0.396 to 6.226

Details on results:

- Behavioural abnormalities: No behavioural abnormalities were identified in treated groups and control groups
- Mortality of control: No Mortality

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

The LC50 value with 95% fiducial limits, and the equation for the dose-response curve was determined by Probit analysis by the method of Finney (1978) using developed and validated computer program.



The observed LC50, NOEC and LOEC value for the test item were expressed in mg/L.

Sublethal observations / clinical signs:

Acclimatization Conditions

Fish were acclimatized at the following conditions during both range finding test and definitive test:

 

Parameter	Range finding test	Definitive test
Duration of acclimatization	7 days	7 days
Light intensity (lux)	675	639
Photoperiodicity	16 h photoperiod daily	16 h photoperiod daily
Temperature of test medium	22.1 to 22.3°C	22.1 to 22.3°C
Dissolved Oxygen concentration of test medium (% saturation)	87 to 97	88 to 97
Hardness (mg/L as CaCO3)	250	250
TOC (mg/L)	0.8	0.8
Feeding	3 times per week	3 times per week

Test Conditions

Parameter	Range finding test	Definitive test
Conditions of testing	Semi-static with renewal of test medium at every 24 h interval	Static
Duration of the test	96 ±1 h	96 ±1 h
Photoperiodicity	16 h photoperiod daily	16 h photoperiod daily
Temperature of test medium	22.1 to 22.6°C	22.1 to 22.6°C
Dissolved oxygen concentration of test medium (% saturation)	86 to 96	86 to 95
pH of test medium	7.68 to 7.74	7.65 to 7.78
Hardness (mg/L as CaCO3)	250	250
TOC (mg/L)	0.8	0.8
Number of fish per group	5	10
Total length of test fish (cm)	1.4 to 1.8	1.4 to 1.8
Group body weight of test fish (g)	2.4359	2.3856
Loading of fish (g/L)	0.1218	0.1193
Feeding	None	None

Acclimatization Test conditions:

Day	Temperature (ºC)	Dissolved oxygen concentration (% saturation)
Day1	22.2	97
Day2	22.1	96
Day 3	22.2	95
Day 4	22.2	93
Day 5	22.2	90
Day 6	22.3	89
Day 7	22.3	87

TABLE 1.            Range Finding Test -pH, Temperature and Dissolved Oxygen Saturation of Test Concentrations

Group	Treatment (mg/L)	Day 1		Day 2		Day 3		Day 4/Day 5
		Start	End	Start	End	Start	End	Start	End
		Temperature (°C)
G1	Negative control	22.3	22.2	22.3	22.2	22.4	22.4	22.3	22.5
G2	0.001	22.4	22.3	22.4	22.1	22.5	22.5	22.3	22.4
G3	0.01	22.5	22.4	22.5	22.2	22.6	22.6	22.2	22.6
G4	0.1	22.3	22.5	22.6	22.2	22.4	22.4	22.3	22.3
G5	1	22.2	22.3	22.4	22.4	22.5	22.2	22.2	22.4
G6	10	22.2	22.2	*	*	*	*	*	*
G7	100	22.3	22.3	*	*	*	*	*	*
		pH
G1	Negative control	7.69	7.71	7.69	7.7	7.71	7.72	7.68	7.69
G2	0.001	7.69	7.72	7.71	7.72	7.68	7.70	7.69	7.71
G3	0.01	7.71	7.72	7.72	7.73	7.69	7.71	7.70	7.72
G4	0.1	7.68	7.71	7.68	7.70	7.71	7.72	7.71	7.73
G5	1	7.69	7.72	7.69	7.71	7.69	7.71	7.72	7.74
G6	10	7.71	7.69	*	*	*	*	*	*
G7	100	7.68	7.72	*	*	*	*	*	*
		Dissolved Oxygen Saturation (%)
G1	Negative control	95	94	93	92	92	90	89	88
G2	0.001	96	93	92	92	91	90	88	87
G3	0.01	95	93	92	92	91	90	89	88
G4	0.1	96	95	93	92	90	89	87	86
G5	1	94	93	92	90	89	88	87	86
G6	10	95	93	*	*	*	*	*	*
G7	100	94	92	*	*	*	*	*	*

Note: Day refers to treatment days    * No data since all fish dead

TABLE 1.            Range Finding Test – Total Length of Individual Fish, Group Body Weight and Loading of Fish

Total length of fish (cm)	1.8	1.7	1.6	1.5	1.5	1.4	1.5	1.4	1.7	1.5
Group body weight (g)	2.4359
Volume of test solution	10 L
No. of fish per group	5
Loading of fish (g/L)	0.1218

TABLE 1.            Range Finding Test – Summary of Toxic Signs and Mortality         

Group

Treatment (mg/L)

Observation of toxic sign and mortality

Cumulative mortality at 96 h

2 h

5 h

24 h

30 h

48 h

54 h

72 h

78 h

96 h

No.

%

G1

Negative control

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

0

0

G2

0.001

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

0

0

G3

0.01

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

0

0

G4

0.1

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

0

0

G5

1

N (5)

N (5)

N (5)

N (5)

N (5)

N (5)

N(4) 1(A)

N(4) M(1)

N(4)

1

20

G6

10

N(2) M(3)

N(1) M(1)

M(1)

*

*

*

*

*

*

5

100

G7

100

N(1) M(4)

M(1)

*

*

*

*

*

*

*

5

100

Total number of fish in each group at the start of test:5; Numbers in parentheses denote the number of fish; N-Normal,
M: Mortality, A: Abnormal horizontal orientation.

* No data since all dead

TABLE 1.            Definitive test -Acclimatization Conditions

Day	Temperature (ºC)	Dissolved oxygen concentration (% saturation)
Day1	22.3	97
Day2	22.1	95
Day3	22.2	94
Day4	22.2	92
Day5	22.1	90
Day6	22.3	89
Day7	22.3	88

TABLE 1.            Definitive Test – pH

Group	Treatment (mg/L)	pH					Mean	± SD
		Day 1	Day 2	Day 3	Day 4	Day 5
G1	Negative control	7.69	7.71	7.72	7.74	7.75	7.72	0.02
G2	0.5	7.71	7.72	7.74	7.76	7.77	7.74	0.03
G3	1.0	7.68	7.72	7.74	7.76	7.78	7.74	0.04
G4	2.0	7.70	7.71	7.73	7.74	7.76	7.73	0.02
G5	4.0	7.68	7.71	7.73	7.76	7.77	7.73	0.04
G6	8.0	7.65	7.66	7.67	7.69	7.71	7.68	0.02
G7	16.0	7.71	7.72	7.75	7.76	*	7.74	0.02
*: No data since all fish dead

TABLE 1.            Definitive Test –Temperature of Test Solutions

Group		Treatment (mg/L)		Temperature of Test Solutions (°C)						Mean		± SD
			Day 1		Day 2		Day 3		Day 4				Day 5
G1		Negative control		22.4		22.4		22.5		22.5		22.2		22.4		0.1
G2		0.5		22.2		22.5		22.4		22.4		22.1		22.3		0.2
G3		1.0		22.3		22.4		22.5		22.3		22.2		22.3		0.1
G4		2.0		22.4		22.4		22.4		22.4		22.3		22.4		0.0
G5		4.0		22.2		22.5		22.5		22.5		22.4		22.4		0.1
G6		8.0		22.5		22.4		22.4		22.6		22.5		22.5		0.1
G7		16.0		22.3		22.3		22.3		22.3		*		22.3		0.0
*: No data since all fish dead

TABLE 1.            Definitive Test –Dissolved Oxygen Saturation of Test Solutions

Group	Treatment (mg/L)	Dissolved Oxygen Saturation (%)					Mean	± SD
		Day 1	Day 2	Day 3	Day 4	Day 5
G1	Negative control	95	94	92	90	89	92	3
G2	0.5	94	93	90	89	87	91	3
G3	1.0	95	93	91	90	88	91	3
G4	2.0	94	92	91	89	87	91	3
G5	4.0	94	92	90	88	86	90	3
G6	8.0	95	93	90	90	88	91	3
G7	16.0	95	92	91	90	*	92	2
*: No data since all fish dead

TABLE 1.            Definitive Test –Total Length, Body Weight and Loading of Sub Sample Fish

Total length of fish (cm)	1.6	1.8	1.5	1.7	1.6	1.4	1.5	1.6	1.8	1.5
Group body weight (g)	2.3856
Volume of test solution	20 L
No. of fish per group	10
Loading of fish (g/L)	0.1193

TABLE 1.            Definitive Test – Summary of Toxic Signs and Mortality

Group No.	Test concentration (mg/L)	Parameter		2 h	5 h	24 h	30 h	48 h	54 h	72 h	78 h	96 h
G1	Negative control	Toxic signs		N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)
		Mortality		0	0	0	0	0	0	0	0	0
		Cumulative mortality	No.	0	0	0	0	0	0	0	0	0
			%	0	0	0	0	0	0	0	0	0
G2	0.5	Toxic signs		N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)	N(10)
		Mortality		0	0	0	0	0	0	0	0	0
		Cumulative mortality	No.	0	0	0	0	0	0	0	0	0
			%	0	0	0	0	0	0	0	0	0
G3	1.0	Toxic signs		N(10)	N(10)	N(10)	N(10)	N(10)	N(9) A(1)	N(9)	N(9)	N(9)
		Mortality		0	0	0	0	0	0	1	0	0
		Cumulative mortality	No.	0	0	0	0	0	0	1	1	1
			%	0	0	0	0	0	0	10	10	10
G4	2.0	Toxic signs		N(8) C(2)	N(8) C(1)	N(6)	N(6)	N(6)	N(5)	N(5)	N(5)	N(5)
		Mortality		0	1	3	0	0	1	0	0	0
		Cumulative mortality	No.	0	1	4	4	4	5	5	5	5
			%	0	10	40	40	40	50	50	50	50
G5	4.0	Toxic signs		N(8)	N(5) A(2)	N(5)	N(5)	N(4) A(1)	N(3) A(2)	N(3)	N(3)	N(3)
		Mortality		2	1	2	0	0	0	2	0	0
		Cumulative mortality	No.	2	3	5	5	5	5	7	7	7
			%	20	30	50	50	50	50	70	70	70
G6	8.0	Toxic signs		N(5)	N(4)	N(4)	N(3) A(1)	N(3)	N(1)	N(1)	N(1)	N(1)
		Mortality		5	1	0	0	1	2	0	0	0
		Cumulative mortality	No.	5	6	6	6	7	9	9	9	9
			%	50	60	60	60	70	90	90	90	90
G7	16.0	Toxic signs		N(4) A(1)	N(2)	N(2)	N(1) A(1)	N(1)	*	*	*	*
		Mortality		5	3	0	0	1	1	*	*	*
		Cumulative mortality	No.	5	8	8	8	9	10	10	10	10
			%	50	80	80	80	90	100	100	100	100

Total number of fish in each group at the start of test: 10; Numbers in parentheses denote thenumber of fish

N – Normal; M- Mortality; A-Abnormal horizontal orientation; C-Loss of buoyancy control

*: No data since all fish

dead

Validity criteria fulfilled:

yes

Remarks:

• No mortality in control at the end of the test. • The DO saturation of the test solutions ranged from 86 to 95 %. • The concentration of the test item in all tested concentrations was between 98.13 to 118.90 % of the nominal concentrations

Conclusions:

The short term toxicity to fish was conducted by following the OECD 203 guideline, on zebrafish, after the exposure tenure of 96 hours the median lethal concentration was determined to be 2.427 mg/L with fiducial limits at 95% ranging from 1.6769 to 3.5123 based on nominal concentrations.

Executive summary:

The short term toxicty to fish was studied on fresh water fish danio rerio by following OECD 203 test guidelines. Juvenile fishes, of average size 1.46 cm with loading rate of 0.1193 g/l was taken and exposed to the test chemical for the period of 96 hours and mortality was observed at regular intervals of 2±0.5 h, 5 ± 1 h, 24 ± 2, 30 ± 2, 48 ± 2, 54 ± 2, 72 ± 2, 78 ± 2 and 96 ± 2 h. Before initiation of study, test organisms were acclimitised to experimental conditions for the period of at least for 9 days (including 7 days acclimatization) before using it for test, no mortality was observed during the acclimatization period. The test medium was reconstituted water, as described in the OECD test guideline. The conductivity of the deionised water used in the preparation of the test medium was 0.054 µS/cm during range finding and definitive test. The test medium was aerated until the dissolved oxygen concentration equated the air-saturation value. The test medium so prepared was set aside without further aeration for a minimum period of at least two days before use and used within 7 days after preparation for the treatment. The range finding test was carried outas a non-GLP testwith test concentrations of0.001, 0.01, 0.1, 1, 10 and 100mg/L of the test medium along with a negative control in a semi-static mode (renewal of test solution at every 24 h interval). Five fish were used for each test solution with single replicate per test concentration. Based on the outcomes the definitive test was carried out using nominal test concentrations0.5, 1.0, 2.0, 4.0, 8.0 and 16mg/L along with control in a static mode. Ten fish were used for each test solution with single replicate per test concentration. To prepare test concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L,10, 20, 40, 80, 160 and 320 mg test item was added to test aquarium containing 20 L of test medium separately.For negative control test aquarium containing 20 L of test medium was used.Test vessels were randomly positioned in the test area and shielded from unwanted disturbance. The concentration analysis in all test concentrations showed that the mean percent recovery with nominal concentration was in the range of 106.85 to 118.90 %(RSD was 2.55 to 8.48 %)at the start and 98.13 to 106.47 %(RSD was 1.75 to 9.59 %)at the end of the test (96 hour) indicating that the results were within the acceptable limit (80 to 120 % of the nominal concentration with an RSD of<20%). In the range finding test, there was no mortality of fish in the negative control and at the tested concentrations of 0.001, 0.01 and 0.1 mg/L up to 96 h. There was 20, 100 and100 %mortality of fish at the tested concentrations of 1, 10 and 100 mg/L at 96 h exposure, respectively. In the definitive test, fish starved at least for 24 h were exposed to test chemical at nominal concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L along with a negative control under static mode. Fish were observed for toxic signs and mortality for 96 h. Each group had single replicate with ten fish. There was no mortality of fish in the negative control and at the tested concentration of 0.5 mg/L at 96 h exposure.There was 0, 40, 50, 60 and 80 % mortality at 24 hours; 0, 40, 50, 70 and 90 % mortality at 48 hours; and 10, 50, 70, 90 and 100 % mortality at 72 and 96 hours at the tested concentrations of1.0, 2.0, 4.0, 8.0 and 16.0 mg/L.On the basis of observation made during the test, the NOEC (No Observed Effect Concentration) was 0.5 mg/L based on nominal concentrations. On the basis of observation made during the test, the LOEC (Low Observed Effect Concentration) was 1.0 mg/L based on nominal concentrations. The LC50 value for test chemical at 24 hours was 3.76 mg/L based on nominal concentrations. The LC50 value for test chemical at 48 hours was 3.354 mg/L with fiducial limits at 95% ranging from 0.396 to 6.226 based on nominal concentrations. The LC50 value for test chemical at 72 and 96 hours was 2.427 mg/L with fiducial limits at 95% ranging from 1.6769 to 3.5123 based on nominal concentrations. As the test chemical is readily biodegradable, it cannot be classified as per CLP classification criteria.

Description of key information

The short term toxicty to fish was studied on fresh water fish danio rerio by following OECD 203 test guidelines. Juvenile fishes, of average size 1.46 cm with loading rate of 0.1193 g/l was taken and exposed to the test chemical for the period of 96 hours and mortality was observed at regular intervals of 2±0.5 h, 5 ± 1 h, 24 ± 2, 30 ± 2, 48 ± 2, 54 ± 2, 72 ± 2, 78 ± 2 and 96 ± 2 h. Before initiation of study, test organisms were acclimitised to experimental conditions for the period of at least for 9 days (including 7 days acclimatization) before using it for test, no mortality was observed during the acclimatization period. The test medium was reconstituted water, as described in the OECD test guideline. The conductivity of the deionised water used in the preparation of the test medium was 0.054 µS/cm during range finding and definitive test. The test medium was aerated until the dissolved oxygen concentration equated the air-saturation value. The test medium so prepared was set aside without further aeration for a minimum period of at least two days before use and used within 7 days after preparation for the treatment. The range finding test was carried outas a non-GLP testwith test concentrations of0.001, 0.01, 0.1, 1, 10 and 100mg/L of the test medium along with a negative control in a semi-static mode (renewal of test solution at every 24 h interval). Five fish were used for each test solution with single replicate per test concentration. Based on the outcomes the definitive test was carried out using nominal test concentrations0.5, 1.0, 2.0, 4.0, 8.0 and 16mg/L along with control in a static mode. Ten fish were used for each test solution with single replicate per test concentration. To prepare test concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L,10, 20, 40, 80, 160 and 320 mg test item was added to test aquarium containing 20 L of test medium separately.For negative control test aquarium containing 20 L of test medium was used.Test vessels were randomly positioned in the test area and shielded from unwanted disturbance. The concentration analysis in all test concentrations showed that the mean percent recovery with nominal concentration was in the range of 106.85 to 118.90 %(RSD was 2.55 to 8.48 %)at the start and 98.13 to 106.47 %(RSD was 1.75 to 9.59 %)at the end of the test (96 hour) indicating that the results were within the acceptable limit (80 to 120 % of the nominal concentration with an RSD of<20%). In the range finding test, there was no mortality of fish in the negative control and at the tested concentrations of 0.001, 0.01 and 0.1 mg/L up to 96 h. There was 20, 100 and100 %mortality of fish at the tested concentrations of 1, 10 and 100 mg/L at 96 h exposure, respectively. In the definitive test, fish starved at least for 24 h were exposed to test chemical at nominal concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L along with a negative control under static mode. Fish were observed for toxic signs and mortality for 96 h. Each group had single replicate with ten fish. There was no mortality of fish in the negative control and at the tested concentration of 0.5 mg/L at 96 h exposure.There was 0, 40, 50, 60 and 80 % mortality at 24 hours; 0, 40, 50, 70 and 90 % mortality at 48 hours; and 10, 50, 70, 90 and 100 % mortality at 72 and 96 hours at the tested concentrations of1.0, 2.0, 4.0, 8.0 and 16.0 mg/L.On the basis of observation made during the test, the NOEC (No Observed Effect Concentration) was 0.5 mg/L based on nominal concentrations. On the basis of observation made during the test, the LOEC (Low Observed Effect Concentration) was 1.0 mg/L based on nominal concentrations. The LC50 value for test chemical at 24 hours was 3.76 mg/L based on nominal concentrations. The LC50 value for test chemical at 48 hours was 3.354 mg/L with fiducial limits at 95% ranging from 0.396 to 6.226 based on nominal concentrations. The LC50 value for test chemical at 72 and 96 hours was 2.427 mg/L with fiducial limits at 95% ranging from 1.6769 to 3.5123 based on nominal concentrations. As the test chemical is readily biodegradable, it cannot be classified as per CLP classification criteria.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

2.427 mg/L

Additional information

Short Term toxicity to fish

The short term toxicty to fish was studied on fresh water fish danio rerio by following OECD 203 test guidelines. Juvenile fishes, of average size 1.46 cm with loading rate of 0.1193 g/l was taken and exposed to the test chemical for the period of 96 hours and mortality was observed at regular intervals of 2±0.5 h, 5 ± 1 h, 24 ± 2, 30 ± 2, 48 ± 2, 54 ± 2, 72 ± 2, 78 ± 2 and 96 ± 2 h. Before initiation of study, test organisms were acclimitised to experimental conditions for the period of at least for 9 days (including 7 days acclimatization) before using it for test, no mortality was observed during the acclimatization period. The test medium was reconstituted water, as described in the OECD test guideline. The conductivity of the deionised water used in the preparation of the test medium was 0.054 µS/cm during range finding and definitive test. The test medium was aerated until the dissolved oxygen concentration equated the air-saturation value. The test medium so prepared was set aside without further aeration for a minimum period of at least two days before use and used within 7 days after preparation for the treatment. The range finding test was carried outas a non-GLP testwith test concentrations of0.001, 0.01, 0.1, 1, 10 and 100mg/L of the test medium along with a negative control in a semi-static mode (renewal of test solution at every 24 h interval). Five fish were used for each test solution with single replicate per test concentration. Based on the outcomes the definitive test was carried out using nominal test concentrations0.5, 1.0, 2.0, 4.0, 8.0 and 16mg/L along with control in a static mode. Ten fish were used for each test solution with single replicate per test concentration. To prepare test concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L,10, 20, 40, 80, 160 and 320 mg test item was added to test aquarium containing 20 L of test medium separately.For negative control test aquarium containing 20 L of test medium was used.Test vessels were randomly positioned in the test area and shielded from unwanted disturbance. The concentration analysis in all test concentrations showed that the mean percent recovery with nominal concentration was in the range of 106.85 to 118.90 %(RSD was 2.55 to 8.48 %)at the start and 98.13 to 106.47 %(RSD was 1.75 to 9.59 %)at the end of the test (96 hour) indicating that the results were within the acceptable limit (80 to 120 % of the nominal concentration with an RSD of<20%). In the range finding test, there was no mortality of fish in the negative control and at the tested concentrations of 0.001, 0.01 and 0.1 mg/L up to 96 h. There was 20, 100 and100 %mortality of fish at the tested concentrations of 1, 10 and 100 mg/L at 96 h exposure, respectively. In the definitive test, fish starved at least for 24 h were exposed to test chemical at nominal concentrations of 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 mg/L along with a negative control under static mode. Fish were observed for toxic signs and mortality for 96 h. Each group had single replicate with ten fish. There was no mortality of fish in the negative control and at the tested concentration of 0.5 mg/L at 96 h exposure.There was 0, 40, 50, 60 and 80 % mortality at 24 hours; 0, 40, 50, 70 and 90 % mortality at 48 hours; and 10, 50, 70, 90 and 100 % mortality at 72 and 96 hours at the tested concentrations of1.0, 2.0, 4.0, 8.0 and 16.0 mg/L.On the basis of observation made during the test, the NOEC (No Observed Effect Concentration) was 0.5 mg/L based on nominal concentrations. On the basis of observation made during the test, the LOEC (Low Observed Effect Concentration) was 1.0 mg/L based on nominal concentrations. The LC50 value for test chemical at 24 hours was 3.76 mg/L based on nominal concentrations. The LC50 value for test chemical at 48 hours was 3.354 mg/L with fiducial limits at 95% ranging from 0.396 to 6.226 based on nominal concentrations. The LC50 value for test chemical at 72 and 96 hours was 2.427 mg/L with fiducial limits at 95% ranging from 1.6769 to 3.5123 based on nominal concentrations. As the test chemical is readily biodegradable, it cannot be classified as per CLP classification criteria.

In an experimental study from peer reviewed journal (Mikio KiKuchi et. al., 1984), an acute toxicity test was conducted for 48 hrs for assessing the effect of test chemical on Oryzias latipes. The test was performed in accordance to the Japanese Industrial Standard (JIS) K0102 (1981) Testing methods for Industrial Wastewater. Adult Oryzias latipes (Japanese medaka) (details regarding average weight and length has been provided in table 2) obtained from a commercial fish farmer in Tokyo was used as a test organism for the study. Test fishes were held in the laboratory for a period of 2 -4 weeks at 20 -23°C prior to use and were fed daily with excess of dry commercial diet pellet. Exact test chemical conc. was not reported, but test concentrations were taken which was increased by geometrical progression of 1.4. Test fishes (total no. of test fishes varies for 4 different runs as mentioned in table 2) were exposed to test chemical in a 3 -5 l glass beaker. The mass/volume ratio was never exceeded of about 1 g fish per liter of water per day. Fishes were not fed during the study. Test solution was renewed after 24 hr. The test vessels were kept in a constant temperature water bath of 21 to 22°C, pH6.7 to 7.1, dissolved oxygen ranges from 8.5 mg/l (at the start of test) to 5 mg/l (at the end of test), hardness of water 25 mg of CaCO3, respectively. Aeration was not provided in test vessels. All tests were carried out in 1 replicate on 4 different runs. Mouth and opercular movement were used to determine the life or death of the test organism. The LC50 were estimated by graphical interpolation of observed survivals greater and less than 50%.On the basis of mortality of the test organism Oryzias latipes (Japanese medaka), the 48 hr LC50 was determined to be 2.4 mg/l.

 

Another short term fish toxicity was conducted for 48 hrs for assessing the effect of test chemical (authoritative database, 2018 and secondary source, 2019). Study was performed using Cyprinus carpio (Commn carp) (larvae) in a static system at 20°C. Test chemical concentration was not verified analytically. On the basis of effect on mortality of the test organism, the 48 hr LC50 was determined to be 3.72 mg/l (95% C. I. – 3.11 to 4.47 mg/l) (nominal concentration).

 

In a supporting weight of evidence study, short term toxicity to fish was conducted for 48 hrs for assessing the effect of test chemical (authoritative database, 2018 and secondary source, 2019). Poecilia reticulata (guppy) was used as a test organism. Study was performed in a static system at 20°C. Test chemical concentration was not verified analytically. On the basis of effect on mortality of the test organism, the 48 hr LC50 was determined to be 8.61 mg/l (95% C. I. – 7.14 to 10.33 mg/l) (nominal concentration).

 

The overall results for the target chemical suggest that, the chemical has tendency to pose toxic effects to freshwater fish in lower concentrations. However, test chemical is readily biodegradable in nature, hence it cannot be classified in to hazardous category, based on CLP classification criteria.