Reaction mass of 4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one and 4-(2,6,6-trimethylcyclohex-1-ene-1-yl)-but-3-ene-2-one

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:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Experimental test result performed according to the OECD test guideline.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

no

Analytical monitoring:

no

Details on sampling:

The test solution was prepared by dissolving 50 mg & 75 mg of the test substance in 10 liters deionized water with continuous stirring for achieving test concentrations of 5 mg/L and 7.5 mg/L, respectively.

Vehicle:

no

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

- Test species/strain : Zebra Fish (Danio rerio)
- Age of the animals : 3-4 months
- Weight/length and physical condition at the initiation of the study:
- Average weight: 0.116 g (approx.)
- Average length: 2.3 cm
- Physical conditions: No abnormalities
- Housing : The fishes were kept in a static tank in tap water, under natural conditions for 15 days along with proper feed and aeration.
- Aeration : Aeration in test vessels was provided 1 day before the start of experiment
- Diet : Standard brand feed, once daily
- Duration of housing : 15 days
- Photoperiod : 12 hours light and 12 hours darkness
- Oxygen content : 7.4

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

29°C

pH:

6-7 pH

Dissolved oxygen:

7.4 mg/l

Nominal and measured concentrations:

On the basis of the range finding study result, test chemical concentration used for the study were 0, 5 and 7.5 mg/l.

Details on test conditions:

The test vessels used were all glass bowl aquaria of 12” each

Test concentrations:
Test concentrations selected for the experiment are based upon the available data and hence the range finding study was planned using two test substance concentrations i.e., 5 mg/L and 7.5 mg/L.

Range finding test:
Range finding test was conducted with the two test concentrations, viz., 5 mg/L and 7.5 mg/L. But at 7.5 mg/L one mortality was observed within 24 hrs. Therefore, the experiment was continued for both the concentration, i.e., 5 mg/L and 7.5 mg/L in which no mortality was found. Observations (mortality, visible symptoms, pH, Temperature, dissolved oxygen content) were recorded after 24 hours, 48 hours, 72 hours and 96 hours of the start of the experiment.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

7.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 7.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Sublethal observations / clinical signs:

Mortality and symptoms:

Nominal conc. (mg/l)	Number of fish	Mortality (cumulated) after
		24h	48h	72h	96h
0 (control)	8	0	0	0	0
5	8	0	0	0	0
7.5	8	1	0	0	0

 

Nominal conc. (mg/l)	Number of fish	Symptoms after
		24h	48h	72h	96h
0 (control)	8	NS8	NS8	NS8	NS8
5	8	NS8	NS8	NS8	NS8
7.5	8	NS7	NS7	NS7	NS7

 

Explanation of symptoms:

A: apathy

N: narcotic-like state

C: swimming near water surface

D: swimming at the bottom of bowl

M: Slow movements compared to control

NS: No abnormal symptoms

Number behind symbol for symptom= number of affected fish

pH and oxygen content:

Nominal conc. (mg/l)	pH readings after
	24h	48h	72h	96h
0 (control)	7.2	6.5	6.2	6.3
5	6.5	6.3	5.9	5.7
7.5	6.1	5.9	5.7	5.4

Nominal conc. (mg/l)	Oxygen content (mg/l) after
	24h	48h	72h	96h
0 (control)	7.6	6.7	6.5	6.1
5	6.5	6.3	6.2	6.0
7.5	6.7	6.3	6.3	6.1

 

Test temperature:

Nominal conc. (mg/l)	Temperature (°C) after
	24h	48h	72h	96h
0 (control)	29	30	31	31
5	29	30	31	31
7.5	29	30	31	31

Validity criteria fulfilled:

yes

Conclusions:

Based on the effect of test chemical on mortality of the test organism Danio rerio (Zebra Fish), the LC0 and LC50 value was determined to be 7.5 and > 7.5 mg/l, respectively.

Executive summary:

An acute toxicity test was conducted for 96 hrs for assessing the effect of test chemical on Zebra fish (Danio rerio). The test was performed in accordance to OECD guideline No. 203 “Fish Acute Toxicity Test”. Zebra fish (Danio rerio) of average length of 2.3 cm and average weight of 0.116 g was used as a test organism for the study. Test fishes were kept in a static tank in tap water under natural conditions for 15 days along with proper feed and aeration. During the housing period, test fishes were fed once daily with standard brand fed. The test conditions during the housing of the test organisms were oxygen content of 7.4 mg/l, pH 6 -7, water temperature 29°C and under a photoperiod of 12:12 hr light: dark conditions, respectively. Test chemical solution was prepared by dissolving 50 mg & 75 mg of the test substance in 10 liters deionized water with continuous stirring for achieving test concentrations of 5 mg/L and 7.5 mg/L, respectively. Range finding test was conducted with these two test concentrations, i.e., 5 mg/L and 7.5 mg/L. Since, at 7.5 mg/L one mortality was observed within 24 hrs. Therefore, the final experiment was continued for both these test chemical concentration, in which no mortality was found. Observations (mortality, visible symptoms, pH, Temperature, dissolved oxygen content) were recorded after 24 hours, 48 hours, 72 hours and 96 hours of the start of the experiment. Test fishes were exposed to test chemical in a glass bowl aquaria. The test vessels were placed in a room at a temperature of 29°C, pH 6 -7 under a photoperiod of 12:12 hr light: dark conditions, respectively. Aeration in test vessels was provided 1 day before the start of the experiment. Mortality in the control vessel was observed to be 0%. On the basis of effect of test chemical on mortality of the test organism, the 96 hr LC0 and median lethal concentration [LC50 (96 h)] for test chemical on Danio rerio (Zebra Fish) was determined to be 7.5 and > 7.5 mg/l, respectively. Thus, test chemical can be considered as toxic to aquatic fishes. Since, the test chemical is readily biodegradable in water, chemical was considered as non-toxic to fish and hence, considered to be 'not classified' as per the CLP classification criteria.

Description of key information

An acute toxicity test was conducted for 96 hrs for assessing the effect of test chemical on Zebra fish (Danio rerio) (Experimental study report, 2013). The test was performed in accordance to OECD guideline No. 203 “Fish Acute Toxicity Test”. Zebra fish (Danio rerio) of average length of 2.3 cm and average weight of 0.116 g was used as a test organism for the study. Test fishes were kept in a static tank in tap water under natural conditions for 15 days along with proper feed and aeration. During the housing period, test fishes were fed once daily with standard brand fed. The test conditions during the housing of the test organisms were oxygen content of 7.4 mg/l, pH 6 -7, water temperature 29°C and under a photoperiod of 12:12 hr light: dark conditions, respectively. Test chemical solution was prepared by dissolving 50 mg & 75 mg of the test substance in 10 liters deionized water with continuous stirring for achieving test concentrations of 5 mg/L and 7.5 mg/L, respectively. Range finding test was conducted with these two test concentrations, i.e., 5 mg/L and 7.5 mg/L. Since, at 7.5 mg/L one mortality was observed within 24 hrs. Therefore, the final experiment was continued for both these test chemical concentration, in which no mortality was found. Observations (mortality, visible symptoms, pH, Temperature, dissolved oxygen content) were recorded after 24 hours, 48 hours, 72 hours and 96 hours of the start of the experiment. Test fishes were exposed to test chemical in a glass bowl aquaria. The test vessels were placed in a room at a temperature of 29°C, pH 6 -7 under a photoperiod of 12:12 hr light: dark conditions, respectively. Aeration in test vessels was provided 1 day before the start of the experiment. Mortality in the control vessel was observed to be 0%. On the basis of effect of test chemical on mortality of the test organism, the 96 hr LC0 and median lethal concentration [LC50 (96 h)] for test chemical on Danio rerio (Zebra Fish) was determined to be 7.5 and > 7.5 mg/l, respectively. Thus, test chemical can be considered as toxic to aquatic fishes. Since, the test chemical is readily biodegradable in water, chemical was considered as non-toxic to fish and hence, considered to be 'not classified' as per the CLP classification criteria.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

7.5 mg/L

Additional information

Experimental study of the test chemical and various supporting weight of evidence studies for its structurally and functionally similar read across chemical were reviewed for short term toxicity to aquatic fish end point which are summarized as below:

 

In an experimental study from study report (2013),an acute toxicity test was conducted for 96 hrs for assessing the effect of test chemical on Zebra fish (Danio rerio). The test was performed in accordance to OECD guideline No. 203 “Fish Acute Toxicity Test”. Zebra fish (Danio rerio) of average length of 2.3 cm and average weight of 0.116 g was used as a test organism for the study. Test fishes were kept in a static tank in tap water under natural conditions for 15 days along with proper feed and aeration. During the housing period, test fishes were fed once daily with standard brand fed. The test conditions during the housing of the test organisms were oxygen content of 7.4 mg/l, pH 6 -7, water temperature 29°C and under a photoperiod of 12:12 hr light: dark conditions, respectively. Test chemical solution was prepared by dissolving 50 mg & 75 mg of the test substance in 10 liters deionized water with continuous stirring for achieving test concentrations of 5 mg/L and 7.5 mg/L, respectively. Range finding test was conducted with these two test concentrations, i.e., 5 mg/L and 7.5 mg/L. Since, at 7.5 mg/L one mortality was observed within 24 hrs. Therefore, the final experiment was continued for both these test chemical concentration, in which no mortality was found. Observations (mortality, visible symptoms, pH, Temperature, dissolved oxygen content) were recorded after 24 hours, 48 hours, 72 hours and 96 hours of the start of the experiment. Test fishes were exposed to test chemical in a glass bowl aquaria. The test vessels were placed in a room at a temperature of 29°C, pH 6 -7 under a photoperiod of 12:12 hr light: dark conditions, respectively. Aeration in test vessels was provided 1 day before the start of the experiment. Mortality in the control vessel was observed to be 0%. On the basis of effect of test chemical on mortality of the test organism, the 96 hr LC0 and median lethal concentration [LC50 (96 h)] for test chemical on Danio rerio (Zebra Fish) was determined to be 7.5 and > 7.5 mg/l, respectively. Thus, test chemical can be considered as toxic to aquatic fishes. Since, the test chemical is readily biodegradable in water, chemical was considered as non-toxic to fish and hence, considered to be 'not classified' as per the CLP classification criteria.

 

In a supporting weight of evidence study, short term toxicity to fish study was conducted for 96 hr for assessing the effect of the test chemical (Geiger D.L. et. al, 1988). Pimephales promelas (Fathead minnows) of 29 days old with length of 20 mm +/- 1.91 mm and weight of 0.122 +/- 0.035 g was used as a test organism. Stock solution test chemical (42.3 mg/l) were prepared by blending. Test chemical concentrations were verified analytically using gas liquid chromatography. Chemical concentrations used for the study were 0, 1.47, 2.26, 3.47, 5.34 and 8.22 mg/l (nominal conc.) and 0, 1.42, 1.75, 2.54, 4.10 and 6.98 mg/l (measured conc.), respectively. Total 20 fishes were exposed with the test chemical in a 2.0 lit tank for 96 hrs. Renewal rate of the test solution was 18 chamber additions/day and biomass loading rate was 1.22 g/l. Study was performed in a flow through system under test conditions like hardness of 44.0 +/- 0.41 mg/l CaCO3, temperature of 25.5 +/- 0.44 °C, pH 7.8 +/- 0.05 and dissolved oxygen of 6.8 +/- 0.31 mg/l. All test experiments were performed in 1 replicate. After an exposure period of 96 hrs, mortality and symptoms of intoxications was noted. On the basis of the effect of test chemical on growrh rate and behaviour of the test organism Pimephales promelas, the 96 hr EC50 and LC50 value was determined to be 4.71 and 5.09 mg/l, respectively. Thus, chemical can be considered as toxic to fish. Since, the test chemical is readily biodegradable in water, chemical was considered as non-toxic and hence, considered to be 'not classified' as per the CLP classificatiins criteria.

 

Another acute fish toxicity study from the peer reviewed journal (A. M. Api et. al, 2016) was conducted for 96 hr for assessing the effect of the test chemical. The study was performed following the german standard guideline DIN 38412 part 15 under static system. Leuciscus idus (Goldn orfe) was used as a test organism. On the basis of the effect of test chemical on mortality of the test organism Leuciscus idus, the 96 hr LC50 value was determined to be 6.81 mg/l. Thus, test chemical can be considered as toxic to aquatic fish. Since, the test chemical is readily biodegradable in water, test chemical was considered as non-toxic and hence, considered to be non-classified as per the CLP classification criteria.

 

For the test chemical, short term fish toxicity study was conducted for 96 hr for assessing the effect of the test chemical (J-CHECK, 2018). The study was performed in accordance with the OECD Guideline 203 (Fish, Acute Toxicity Test) in a static system. On the basis of the effect of test chemical on mortality of the test organism, the 96 hr LC50 value was determined to be 3.0 mg/l. Thus, test chemical can be considered as toxic to aquatic fish. Since, the test chemical is readily biodegradable in water, test chemical was considered as non-toxic and hence, considered to be non classified as per the CLP classification criteria.

 

On the basis of the overall results, it can be concluded that the test chemicalwas considered as toxic to fish. Since, the test chemical is readily biodegradable in water, chemical was considered as non-toxic to aquatic fishes at environmental concentrations and hence, considered to be'not classified' as per CLP classification criteria.