3,5,5-trimethylcyclohex-2-enone

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test procedure comparable to standard method and in accordance with general accepted scientific standards; detailed documentation of test procedure and test conditions

Principles of method if other than guideline:

Method: other

GLP compliance:

no

Analytical monitoring:

yes

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISMS
- fish were breeded and cultured at the U.S. EPA Environmental Research Laboratory -Dulutk
- Adults were held at 25 °C in flowing water and a photoperiod of 16 h light
- fish were fed frozen adult brine shrimp (Artemesia sp.)
- 27-74 day old fish were used for the test

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

- 50.0 mg/l CaCO3

Test temperature:

- 24.5 °C

pH:

- 7.5

Dissolved oxygen:

- 7.1 mg/l

Nominal and measured concentrations:

- Nominal: 46.5, 71.5, 110, 170, 261 mg/l               
- Measured: 43.8, 72.0, 114, 186, 275 mg/l 

Details on test conditions:

TEST SYSTEM
- Test type: flow through
- Number of replicates: 2
- Fish per replicate: 5-25
- Fish exposed in Lake Superior water
- Test temperature: 24.5 °C
- Adjustment of pH: adjusted to lake water using NaOH
- Photoperiod: 16 h light
- Alkalinity: 40.5 mg/l CaCO3
- 5 test substance concentrations in the range of 46.5 to 213  mg/l tested (plus control)

EFFECT PARAMETERS
- number of fish recorded every 24 h;  
- observations of fish behaviour and body morphology at regular intervals; 

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

228 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

217 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

behaviour

Details on results:

- Affected fish lost schooling behavior, were hypoactive and underreactive  to external stimuli and were darkly colored. 
- They had also spinal column deformities and lost equilibrium prior to death.
- mean lenght at the end of the test: 15.5 mm; 
- mean weight at the end of the test: 0.052 g 

Reported statistics and error estimates:

No confidence limits estimated

Sublethal observations / clinical signs:

Measured concentrations (mg/L):

Time	Control	A	B	C	D	E
0 h	5.0	32.6	54.1	89.8	147	213
24 h	5.0	43.0	73.1	115	184	279
48 h	5.0	43.6	77.0	127	211	317
72 h	5.0	54.5	81.0	123	194	282
96 h	5.0	45.4	75.0	115	196	284
Average	5.0	42.0	72.0	114	186	275

Conclusions:

The LC50 (96 hours) was determined as 228 mg/L indicating that the test item does not pose a significant hazard to fish.

Executive summary:

The acute effects of the substance on Pimephales promelas were investigated in a 96h study in accordance with general accepted scientific standards. The following endpoints were derived

 

EC50 (96h): 217 mg/L

LC50 (96h): 228 mg/L

 

The study was assessed as "reliable without restrictions".

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

Method: other: Acute Toxicity Test based on a procedure of the Committee on Methods for Toxicity Tests with Aquatic Organisms

GLP compliance:

not specified

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

STOCK AND TEST SOLUTION AND THEIR PREPARATION
- Vehicle, solvent: not used
- Other procedures: diluter cycled for >= 24 h in advance

Test organisms (species):

Cyprinodon variegatus

Details on test organisms:

TEST ORGANISMS
- Supplier: inhouse (Bionomics Marine Research Laboratory)
- Age/size/weight/loading: < 20 days
- Pretreatment: in natural, flowing seawater at 30 +/- 1 °C

Test type:

flow-through

Water media type:

saltwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

- not reported

pH:

- not reported

Nominal and measured concentrations:

Control, 30, 59, 119, 238, 476 mg/L (nominal)
Analytical concentrations were 149-187 % of nominal with test concentrations of 47-608 mg/L. Results refer to mean measured concentration.

Details on test conditions:

EST SYSTEM
- Concentrations: 5 levels, 47-608 mg/l
- Dosing rate: 1 l per cycle
- Renewal of test solution: 4-7 cycles per hour
- Exposure vessel type: glass aquariums 30 cm wide, 30 cm high, 91 cm  long, containing approximately 41 l solution
- Number of replicates, fish per replicate: not reported
- Adjustment of pH: no
- Intensity of irradiation: approximately 1100 lux incident to water  surface
- Photoperiod: 16 hours light, 8 hours dark

DILUTION WATER
- Source: natural sea water, Big Lagoon (estuary of Gulf of Mexico),  filtered
- Salinity: daily measurement (few exceptions)
- pH: daily measurement (few exceptions)
- Oxygen content: daily measurement (few exceptions)

TEST PARAMETER: 
- mortality

 STATISTICAL ANALYSIS: 
- conversion of concentrations to log and of  percentages mortality to probit
- 96-h LC50s and their 95% confidence limits

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

140 mg/L

Nominal / measured:

not specified

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 90-217 mg/L

Sublethal observations / clinical signs:

95 % confidence limits of LC50: 90-217 mg/l

Conclusions:

The LC50 (96 hours) was determined as 140 mg/L indicating that the test item does not pose a significant hazard to saltwater fish.

Executive summary:

The acute toxicity of the substance on Cyprinodon variegatus was evaluated in a 96 -h test according to a procedure of the Committee on Methods for Toxicity Tests with Aquatic Organisms (USA). The following endpoints were determined:

LC50 (96h): 140 mg/L; 95% CL: 90 -217 mg/L

The study was assessed as "reliable with restrictions".

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

Method: other: those recommended by the American Public Health Association (APHA et al. 1980) and the Environmental Protection Agency ( U.S. EPA 1974)

GLP compliance:

not specified

Analytical monitoring:

yes

Details on sampling:

MONITORING OF TEST SUBSTANCE CONCENTRATION:  
- sampling twice weekly, HPLC analysis

Vehicle:

no

Details on test solutions:

STOCK AND TEST SOLUTION AND THEIR PREPARATION
- Dispersion: Diluter
- Vehicle, solvent: none

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISMS -
- Age/size/weight/loading: Initial age 6-8 weeks

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

- Hardness: 35 (24-70) mg/l CaCO3

Test temperature:

- Test temperature: 24.5 +/- 1 °C

pH:

- pH: 7.4 (7.1-7.6)

Dissolved oxygen:

- Oxygen content: 7.9 (6.2-8.4) mg/l

Nominal and measured concentrations:

- Concentrations: 73-412 mg/l

Details on test conditions:

DILUTION WATER
- Source: wells at Western Fish Toxicology Station, Corvallis, Oregon
- Alkalinity: 28 (20-47) mg/l CaCO3

TEST SYSTEM
- Renewal of test solution: 88 volume additions in 24 hr
- Exposure vessel type: 1 l glass beakers, 900 ml solution, 10.5 cm deep
- Number of replicates, fish per replicate: 2 replicates, 10 fish per  chamber
- Intensity of irradiation: approximately 270 lux on surface
- Photoperiod: 16 h light / 8 h dark

STATISTICAL METHODS: 
- trimmed Spearman-Karber method

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

255 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 229-283

Sublethal observations / clinical signs:

95 % confidence interval (LC50): 229-283 mg/l

Executive summary:

Pimephales promelas (6 -8 weeks old) was exposed to the test item in a 96-h flow-through test. The study was conducted according to accepted scientific guidance (U.S.). The following endpoints were determined:

 

LC50 (96h): 255 mg/L; 95% CL: 229 -293 mg/L

 

The study was assessed as "reliable with restrictions".

Reference 4

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

Method: other: those recommended by the American Public Health Association (APHA et al. 1980) and the Environmental Protection Agency ( U.S. EPA 1974)

GLP compliance:

not specified

Analytical monitoring:

yes

Details on sampling:

MONITORING OF TEST SUBSTANCE CONCENTRATION:  
- sampling twice weekly, HPLC analysis

Details on test solutions:

STOCK AND TEST SOLUTION AND THEIR PREPARATION
- Dispersion: Diluter
- Vehicle, solvent: none

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISMS -
- Age/size/weight/loading: Initial age 3 weeks

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

- Hardness: 35 (24-70) mg/l CaCO3

Test temperature:

- Test temperature: 24.5 +/- 1 °C

pH:

- pH: 7.4 (7.1-7.6)

Dissolved oxygen:

- Oxygen content: 7.9 (6.2-8.4) mg/l

Nominal and measured concentrations:

- Concentrations: 59-394 mg/l

Details on test conditions:

DILUTION WATER
- Source: wells at Western Fish Toxicology Station, Corvallis, Oregon
- Alkalinity: 28 (20-47) mg/l CaCO3

TEST SYSTEM
- Renewal of test solution: 88 volume additions in 24 hr
- Exposure vessel type: 1 l glass beakers, 900 ml solution, 10.5 cm deep
- Number of replicates, fish per replicate: 2 replicates, 10 fish per  chamber
- Intensity of irradiation: approximately 270 lux on surface
- Photoperiod: 16 h light / 8 h dark

STATISTICAL METHODS: 
- trimmed Spearman-Karber method

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

145 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 132-159

Sublethal observations / clinical signs:

RS-Freetext:
95 % confidence interval (LC50): 132-159 mg/l

Executive summary:

 

Pimephales promelas (3 weeks old) was exposed to the test item in a 96-h flow-through test. The study was conducted according to accepted scientific guidance (U.S.). The following endpoints were determined:

 

LC50 (96h): 145 mg/L; 95% CL: 132 -159 mg/L

 

The study was assessed as "reliable with restrictions".

Description of key information

The acute toxicity of the test item on one freshwater species (Pimephales promelas) and on one marine species (Cyprinidon variegatus) was assessed over a period of 96 h according to accepted scientific standards.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

228 mg/L

Marine water fish

Marine water fish

Effect concentration:

140 mg/L

Additional information

Among the existing valid studies, the two above mentioned studies were found to show the highest toxicity. The 96 -h EC50 values were 228 mg/L for the freshwater species Pimephales promelas (Geiger et al., 1990) and 140 mg/L for the marine species Cyprinidon variegatus (Ward et al., 1981).

Supporting information:

Pimephales promelas in different early life stages were exposed to the test item in a 96-h flow-through test. The study was conducted according to accepted scientific guidance (U.S.). The following endpoints were determined:

LC50 (96h): 145 mg/L; 95% CL: 132 -159 mg/L (Fish 3 weeks old) (Cairns, 1982)

LC50 (96h): 255 mg/L; 95% CL: 229 -293 mg/L (Fish 6-8 weeks old) (Cairns, 1982)