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Long-term toxicity to fish

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Reference
Endpoint:
long-term toxicity to fish, other
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not specified.
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
US EPA standardised method used. GLP not specified. Results taken from TSCA fiche OTS0546071.
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: EPA method 1000.0 (3rd ed.; EPA/600/4-91/002)
Deviations:
no
Principles of method if other than guideline:
Chronic Toxicity
The procedure used in the test followed EPA method 1000.0 (3rd ed.; EPA/600/4-91/002). The test had a total of 4 replicates of 10 organisms per concentration (0, 2.5, 5, 10, 20, and 40 mg/L). Cadmium chloride was tested as a reference toxicant. The test was performed using the target temperature of 25+1°C and a 16-hour light/8-hour dark photoperiod. Synthetic moderately hard water served as laboratory dilution water for the test. The water was stored at 20°C under gentle aeration until needed, up to 14 days. Stocks of the test substance were prepared each day in moderately hard synthetic freshwater, and these stocks were used to dose the test solutions. Larval fish <24 hours old were randomly loaded into test chambers. During testing, fish were fed a suspension of brine shrimp daily. The test solutions were renewed daily.
Mortality observations were recorded daily. Temperature, pH, dissolved oxygen, and conductivity were measured in the new and old test solutions daily, and at test termination. Growth of the larvae were measured at the termination of the test.
The results of the toxicity tests were analyzed using the ToxCalc statistical software package and followed US EPA guidance.
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material:

Analytical monitoring:
no
Details on sampling:
- Concentrations: A rangefinding toxicity tests used the concentration series of 100, 50, 25, 12.5 and 6.25 mg/L oxydianiline and a dilution water control. The definitive toxicity test used the concentration series of 40, 20, 10, 5 and 2.5 mg/L oxydianiline and a dilution water control.
- Sampling method: None
- Sample storage conditions before analysis: The test solutions were renewed each day by siphoning 80 percent of the old test solutions from the test vessels and replacing with freshly made test solutions, made at the time.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stocks of the test substance were prepared each day in moderately hard synthetic freshwater, and these stocks were used to dose the test solutions.
- Eluate: Synthetic moderately hard water (US EPA 1993) served as laboratory dilution water for the tests. Batches of this water were made by passing deionized water through activated carbon, adding reagent grade chemicals, and aerating overnight. The water was stored at 20°C under gentle aeration until needed up to, 14-days.
- Controls: Cadmium Chloride
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Not applicable
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): Not applicable
- Evidence of undissolved material (e.g. precipitate, surface film, etc): Not specified
- Conductivity: 254 - 384 µS/cm
Test organisms (species):
Pimephales promelas
Details on test organisms:
Pimephales promelas (fathead minnow) were obtained from EA's Culture Facility. Brood organisms were maintained in recirculating dechlorinated tap water at 20°C in 10-gallon aquaria. For the chronic toxicity tests, eggs produced from the brood system were removed from the brood aquaria and placed into dechlorinated tap water at 25°C for the incubation period. Newly hatched larvae, less than 24 hours old, from lots FH 2-9/25-26 were used in the chronic definitive toxicity testing..
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
7 d
Post exposure observation period:
None
Hardness:
Not measured
Test temperature:
Chronic Definitive: 24.0-25.7 (°C)
pH:
Chronic Definitive: 7.4-8.4
Dissolved oxygen:
Chronic Definitive: 6.0-8.1 mg/L
Salinity:
Not applicable.
Nominal and measured concentrations:
Chronic definitive toxicity test: 40, 20, 10, 5 and 2.5 mg/L test substance and a dilution water control.
Details on test conditions:
For the P. promelas chronic toxicity tests, larval fish <24 hours old were randomly loaded into the test chambers. The rangefinding test had a total of 2 replicates of 10 organisms per concentration and control. The definitive test had a total of 4 replicates of 10 organisms per concentration and control. The test chambers were 1-liter plastic beakers with 250 ml of test solution. During testing, organisms in each test chamber were fed daily a suspension of brine shrimp Artemia nauplii. The uneaten and dead brine shrimp were removed from the test vessels daily. The test solutions were renewed each day by siphoning 80 percent of the old test solutions from the test vessels and replacing with freshly made test solutions.
Reference substance (positive control):
yes
Remarks:
Cadmium chloride
Key result
Duration:
7 d
Dose descriptor:
NOEC
Effect conc.:
5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
7 d
Dose descriptor:
LOEC
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
7 d
Dose descriptor:
other: IC25
Effect conc.:
14.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: (confidence limits, 12.0-31.2 mg/L)
Details on results:
Results of the definitive P. promelas definitive test were similar to the range fnding (see results of the acute toxicity test). At test termination, there was 15 percent survival in 40 mg/L test substance concentration and 55 percent survival in the 20 mg/L test substance concentration. Both concentrations were sigificantly different (p=0.05) from the control which had 98 percent survival. There was a minimum of 84 percent survival in the 2.5 to 10 mg/L test substance concentrations. Mean growth in the 10 mg/L test substance concentration (0.524 mg/organism) was significantly different (p = 0.05) from the control which had 0.659 mg/organism. Mean growth in the 2.5 mg/L and 5 mg/L test substance concentrations was 0.606 and 0.627 mg/organism, respectively. Using growth as the most sensitive test endpoint, the NOEC was 5 mg/L test substance , the LOEC was 10 mg/L test substance. The IC25 was 14.1 mg/L test substance .
Results with reference substance (positive control):
Results with cadmium chloride: IC25= 22.1 mg/l CdCl2 ; acceptable control chart limits of 11.2 - 35.2 mg/l.
Reported statistics and error estimates:
The results of the toxicity tests were analyzed using the ToxCalc statistical software package (Version 5.0, Tidepool Scientific Software) and followed US EPA guidance (US EPA 1994).

Results of the chronic definitive test with fish larvae

 Nominal test concentration (mg/L)  96h - % survival  7d % survival  Mean growth (mg/organism +/- SD)
 Control  98 98   0.659 +/- 0.098
 2.5  98 98  0.606 +/- 0.052 
 5.0  98 98  0.627 +/- 0.027 
 10 84 84  0.524 +/- 0.063* (p<0.05)
 20  60 55  0.453 +/- 0.076
 40  18 15  0.155 +/- 0.139 

96h LC50 = 22.7 mg/L (18.7 - 27.5 mg/L) 95%IC

NOEC = 5 mg/L

LOEC = 10 mg/L

Validity criteria fulfilled:
yes
Remarks:
The result with the reference substance, cadmium chloride is within the range of acceptability
Conclusions:
3,4'-oxydianiline is of moderate concern for chronic toxicity to fish after 7 days of expsure (NOEC = 5 mg/L). The lowest concentration at which a toxicity was observed was 10 mg/L (16% mortality and a significant effect on growth)
Executive summary:

A chronic toxicity test was conducted on Pimephales promelas larvae for 7 days according to US EPA standard. The test was perfomed under semi-static conditions with a renewal of the test concentrations by 80% every day. All validity criteria are fulfilled and the results are expressed as nominal concentrations. As the substance does not degrade rapidly in water, those nominal concentrations are considered to be representative of actual concentrations. The results of the tests lead to a NOEC of 5 mg/L (based on growth of larvae). This result can be used in the dossier and for classification of 4,4'-oxydianiline.

Description of key information

A chronic toxicity test on the analogue 3,4'-oxydianilline was conducted on Pimephales promelas larvae for 7 days according to US EPA standard. The test was perfomed under semi-static conditions with a renewal of the test concentrations by 80% every day. The results of the tests lead to a NOEC of 5 mg/L (based on growth of larvae).

This result can be used in the dossier and for classification of 4,4'-oxydianiline.

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater fish:
5 mg/L

Additional information