2,5-bis(p-toluidino)terephthalic acid

Short-term toxicity to fish

Administrative data

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: The study was conducted according to relevant guidelines and compliant to GLP. The results are plausible and well documented.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

yes

Remarks:

Because of a thermostat defect, on the last study day the temperature dropped to 20.1 °C which is below the defined lowest value of 22 +- 1°C

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

yes

Remarks:

Because of a thermostat defect, on the last study day the temperature dropped to 20.1 °C which is below the defined lowest value of 22 +- 1°C

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2,5-bis(p-toluidino)terephthalic acid

EC Number:

233-651-6

EC Name:

2,5-bis(p-toluidino)terephthalic acid

Cas Number:

10291-28-8

Molecular formula:

C22H20N2O4

IUPAC Name:

2,5-bis[(4-methylphenyl)amino]benzene-1,4-dicarboxylic acid

Details on test material:

Trade name: Toluidinosäure TR 100
Synonyms: Terephthalic acid, 2,5-di-p-toluidino
Chemical name: 1,4-Benzenedicarboxylic acid, 2,5-bis[(4- methylphenyl)amino]-
Sample received: 1994-04-08
Storage: in fume cupboard at approx. 20 °C in the dark
Solubility in water: low

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Representative sample of total water amount 10L: 20mL
Appearance of sample : yellow solutions
Temperature of water samples during the toxicity testing 22°C+-1°C

Test solutions

Vehicle:

yes

Details on test solutions:

The test substance was weighed into a beaker. The test substance was made to a paste using 0.095 mL Tween 80/L as solubilizing agent. Water for dilution was added and the mixture homogenized using an Ultra-Turrax and poured into the chamber under stirring with a glass rod. The chamber contents were then stirred for approximately 30 minutes with a KPG-stirrer and a glass rod. The tested concentration was present as a yellow solution, with particulate matters at the botton of the chamber.
The given concentration (100 mg/L) is based on the amount of substance introduced into the chamber and not on the dissolved portion.
In order to determine the substance concentration, water samples were taken from the middle of the test chamber by pipetting approx. 20 mL into a screw-neck glass bottle of each first and third preparation of the concentration after 0 and 24 hours, respectively.

Test organisms

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Species: zebra fish Brachydanio rerio (HAMILTON-BUCHANAN)
Breeding station : Corporate Toxicology (Hoechst AG)
Date of hatching: 1993-09-03
Delivery date: 1993-11-01
The fish were kept for at least 14 days before the start of the study in water for dilution under the following conditions:
Temperature 22 ± 1 °C
Oxygen content: >= 80 % of the saturation value
Duration of light period : 12 hours daily
Population density : < =1 g fish/L water
Feeding : twice daily ad libitum
Feed : Tetra Min, Tetra Werke, Melle (FRG)
The body length of 7 representative fish from the batch was measured:
Batch no.: 11/93/H4/L7, Date: 1994-10-05, n= 7, Variation range of body length: 2.8 - 3.5 cm, Mean = 3.1 cm, SD = 0.31

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Post exposure observation period:

24 hours

Test conditions

Hardness:

The total hardness of the dilution water was determined weekly and was approx. 2.3 mmol Ca2+ + Mg2+ / L during the study.

Test temperature:

Concentration group and control groups: 20.1 - 21.8 °C: Because of a thermostat defect, on the last study day the temperature dropped to 20.1 °C which is below the defined lowest value of 22 +- 1°C. However, as the behavioral pattern did not deviate from the norm in any of the study groups, an impairment of the quality and integrity of the study can therefore be excluded. There were no further unforeseen circumstances.

pH:

Concentration group: 7.8 - 8.3
Controls: 7.9 - 8.5

Dissolved oxygen:

[mg/L]
Concentration group: 7.8 - 9.6
Controls: 8.0 - 9.6

Salinity:

not applicable

Nominal and measured concentrations:

Nominal concentration: 100 mg/L
Mean measured concentrations:
First preparation, 0h: 63.7 mg/L
First preparation, 24h: 51.8 mg/L
Third preparation, 0h: 60.5 mg/L
Third preparation, 24h: 52.2 mg/L
The values determined were between 51.8 and 63.7 % of the theoretical value. The reported concentration based on the analytical mean concentration is 57.05 mg/L. The low content of the test substance is caused by low water solubility.

Details on test conditions:

The study was conducted in a semistatic system. The fish were transferred to new test chambers every 24 hours. The test chambers, which were calibrated to 10 liters, were made of glass (length 30 cm, width 22 cm, height 24 cm) and stood in a water bath made of Hostalit Z with a Plexiglas viewing panel. The temperature of the water bath was regulated by a thermostat to 22 + 1 °C. The chambers were illuminated from above from 6.00 a.m. to 6.00 p.m. The light intensity directly over the chambers was approximately 700 lux.
The test chamber was not aerated during the course of the study.

Water for dilution:
Reconstituted water, composition according to ISO/DIS 7346/1 was used as water for dilution.
Preparation was carried out in a unit consisting of two Hostalen lined steel vessels with a capacity of 1000 litres, each.
The dilution water was prepared as described and was aerated until oxygen saturation. The pH was measured before the use of each study batch and was 8.3 - 8.5.

Study conduct :
After the first test batches had been prepared and water parameters recorded, 7 fish were assigned to each test and control chamber. The fish received no feed for the entire study period. Inspection of the fish in the first prepared test batch of each concentration took place after 3, 6 and 24 hours, in all further prepared test batcnes after 3 - 6 and 24 hours, and involved recording the lethality and visible changes in appearance and behavior. Dead fish were removed from the chambers. Fish were considered dead when there was a lack of opercular movement and no response to slight mechanical stimulus. The water parameters were measured and recorded before study start and after 0 and 24 hours. This procedure was repeated for all fresh prepared test batches over a total of 96 hours.

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

>= 57.05 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: limit concentration of 100 mg/L (= nominal concentration) could not be reached due to low solubility of test material

Effect concentrations 2

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

> 57.05 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: limit concentration of 100 mg/L (= nominal concentration) could not be reached due to low solubility of test material

Effect concentrations 3

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 57.05 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: limit concentration of 100 mg/L (= nominal concentration) could not be reached due to low solubility of test material

Effect concentrations 4

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

> 57.05 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: limit concentration of 100 mg/L (= nominal concentration) could not be reached due to low solubility of test material

Effect concentrations 5

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 57.05 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: limit concentration of 100 mg/L (= nominal concentration) could not be reached due to low solubility of test material

Details on results:

In this 96-hour acute toxicity study of the submission substance in zebra fish (Brachydanio rerio) no lethality occurred at 100 mg/L nominal concentration (57.05 mg/L measured concentration).
Neither in the control nor in the solvent control occurred any mortalities.
The LC50 after 96 hours therefore exceeds these values and the solubility limit of the test substance in the test water.

The fish showed changes in appearance and behavior (dark colored integument, increased fright reaction, hypoactivity), swimming behavior (swimming posture tail-heavy, swimming at the bottom) and respiration rate (projecting opercula) during the first 72 hours of exposure. At the end of the study no symptoms were observed (fourth day and post observation day (fifth day).

Results with reference substance (positive control):

not applicalble

Reported statistics and error estimates:

not applicable

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

In a 72 h acute toxicity test on Brachydanio rerio with 24 h postobservation period, the toxixity of the submission substance to fish was determined in a semistatic system. Due to the poor solubility, a limit test at nominal 100 mg/L submission substance concentration with the vehicle Tween 80 had been performed. The mean measured concentration had been 57.5 mg/L. There were no mortalities at this concentration and in the control groups. The NOEC (24h, 48h, 72h, 96h) is therefore >= 57.5 mg/L, the LC50 (24h, 48h, 72h, 96h) > 57.5 mg/L.

Executive summary:

This 96-hour acute toxicity study of Toluidinosäure TR 100 in zebra fish (Brachydanio rerio) was conducted in a semistatic system, because in a static system the substance was not analytically detectable to a sufficient degree over 96 hours. No lethality occurred at 100 mg/L (nominal concentration) and 57.05 mg/L (measured concentration). The LC50 after 96 hours therefore exceeds these values and the solubility limit in the test water. To get a better suspension of the test substance in the test water, TWEEN 80 was used as emulsifying agent. The concentrations tested were 100 mg/L, a negative control (0 mg/L) and a control group with the emulsifying agent TWEEN 80, using the concentration 0.095 mL/L. The tested concentration was present as clear yellow solution with particulate matters at the bottom of the chamber. The substance concentration was determined in the 100 mg/L group. The values determined were between 51.8 and 63.7 % of the theoretical value. The low concentration of the test substance was caused by low water solubility.