Nitrosodiphenylamine

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

Study period:

1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test performed according to OECD 203 testing guidelines, GLP compliant. The only deviation to the guidelines is the use of solvent, because the test item was considered insoluble. However, no mortality was recorded in the solvent control group.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

yes

Remarks:

The only deviation is the use of a solvent, because the test item was considered insoluble. However, the solvent control group did not suffer any loss, which is considered to not impact the results. The experiment is therefore deemed as valid.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

0.75 mL of the test fluid was sampled from each of the test water tanks both before the start of testing and after 24 hours, and after the addition of 0.75 mL of acetonitrile, these were mixed well, and analysed by HPLC as analysis samples. During analysis of the test fluids, assay of a standard solution was performed for each specimen assay, and quantitated from the peak surface area ratio.

Vehicle:

yes

Details on test solutions:

400 mg dimethylsulfoxide and a 1000 mg dispersal agent (HC0-30) were added to 400mg of the tested substance, and after being mixed and dissolved, the volume was set at 2000 mL with purified water, and this was treated as the test substance stock solution (200 mg/L concentration). For the adjuvant stock solution, 200mg dimethyl sulfoxide and 500 mg of the dispersal agent (HC0-30) were mixed and thereafter the volume was set to 1000mL using purified water.
For each concentration to be tested,after adding the above test substance stock solution according to the required final concentration, the adjuvant stock solution was added so that the adjuvant concentration was constant among each beaker. The control groups received the dilution water or the solvent containing dilution water only.

Test organisms (species):

Oryzias latipes

Details on test organisms:

- Common Name: Orange Killfish
- Scientific name: Orzias Latipes
- Scale Length: 1.91 cm (1.68-2.13 cm), n=10
- Weight: 0.116 g (0.085-0.145 g), n = 10
- Supplier: Sankyo Aquaculture Center (1-1 Ban-chi, Ichidani-cho, Shinjuku Ward, Tokyo)
- Delivery Date: November 1, 1996
- Acclimation period: November 1, 1996 - February 3, 1996

Bred and domesticated under the same conditions (water quality, temperature, etc.) as the test conditions. Commercial Tetramine® was given for feed. No feeding was performed from 24 hours prior to exposure. Normal specimens that were grossly healthy were used in the testing. The mortality rate for the 7 days prior to the start of exposure was 5% or less.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Post exposure observation period:

None

Hardness:

61 mg equivalent CaCO3

Test temperature:

23.2 - 24 °C (min-max)

pH:

7.4-7.8 (min-max)

Dissolved oxygen:

5.9-8.4 (min-max) , always >60% saturation

Salinity:

12 mg/L sodium

Nominal and measured concentrations:

- Nominal concentrations: 2 ; 3.8 ; 7.4 ; 14 ; 28 mg/L
- Measured concentrations (geometrical means): 2.07 ; 3.99 ; 7.65 ; 15.3 ; 31.6 mg/L
- Deviations from nominal concentrations did not exceed 20% throughout the experiment. Therefore, nominal concentrations were used for statistical analysis

Details on test conditions:

- Exposure Form: Semi-static method with 5-L glass beakers (The entire volume of test fluid was replaced every 24 hours. In addition, the surface of the water was covered with a Teflon shield in order to prevent the volatization of the tested substance from the water).
- Exposure Period: 96 hours
- Test fluid volume: 5.0 L
- Number of replicate: 1 container per concentration group
- No. of organisms: 10 perconcentration group
- Aeration: None
- Test water temperature: 24±1°C
- Illumination: Room light, 16L:8D
- Feeding: No feeding

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

10.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

The 96-hour median lethal concentration was 10.2mg/L. Furthermore, the mortality rates of the control plot and the adjuvant control plot were 0%.

Results with reference substance (positive control):

Not applicable.

Reported statistics and error estimates:

The mortality rate (%) was calculated from the number of dead bodies in each concentration group and the number of test organisms (10), and the median lethal concentration (LC50) was calculated by the binomial method.

Sublethal observations / clinical signs:

Cumulative mortality:

Control: 0%

Solvent control: 0%

2 mg/L: 20%

3.8 mg/L: 0%

7.4 mg/L: 0%

14 mg/L: 100%

28 mg/L: 100%

Validity criteria fulfilled:

yes

Conclusions:

Oryzias latipes were exposed to a graded series of N-nitrosodiphenylamine concentrations for 96 hours. The estimated LC50 was 10.2 mg/L.

Executive summary:

Oryzias latipes were exposed to a graded series of N-nitrosodiphenylamine concentrations (control; solvent control ; 2 ; 3.8 ; 7.4 ; 14 ; 20 mg/L) for 96 hours, according to the OECD 203 guidelines. The only deviation to these guidelines was the use of a solvent to dissolve the test-item, considered as insoluble. However, the control solvent did not cause any mortality. Measured concentrations did not differ from the nominal concentrations by more than 20%. Therefore, nominal concentrations were used for statistical analysis. The estimated LC50 after 96 hours exposure is 10.2 mg/L

Description of key information

96-h LC50 of fish was estimated to be above 10.2 mg/L by Mitsubishi (1997). In a second study, they found a 7-d LC50 of 8.9 mg/L and a 14-d LC50 of 4.9 mg/L, with a 14-day NOEC of 3.3 mg/L. These data are together consistent. The study of Buccafusco was disregarded because the test-item formed a precipitate within the test system and oxygen concentration dropped to 0.3 mg/L far beneath acceptable limits. LC50 found by Buccafusco was 5 mg/L.
The 96-h LC50 value will be retained for risk assessment since it is compliant with the guidance on information requirements and chemical safety assessment Chapter R10: characterisation of dose [concentration]-response for environment (2008).
Fish LC50 = 10.2 mg/L

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

10.2 mg/L