2,4-bis[N'-(4-methylphenyl)ureido]toluene

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 November 1992 - 23 March 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study has been performed according to OECD and/or EC guidelines and according to GLP principles. However, the analytical concentrations were not measured for all test levels, particularly they were not measured for the purported NOEC level. In addition, there was significant variability in the concentrations, with a mid-level concentration higher than the high concentration at the start of the test, and the high level concentration increasing over the course of the test, whilst the other concentrations decreased. Limited information available to verify the composition of the used test substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

(1984)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2,4-bis[N'-(4-methylphenyl)ureido]toluene

EC Number:

411-790-5

EC Name:

2,4-bis[N'-(4-methylphenyl)ureido]toluene

Cas Number:

54390-87-3

Molecular formula:

C23H24N4O2, C24H34N4O2

IUPAC Name:

3-(2-methyl-3-{[(4-methylphenyl)carbamoyl]amino}phenyl)-1-(4-methylphenyl)urea; 3-(2-methyl-5-{[(4-methylphenyl)carbamoyl]amino}phenyl)-1-(4-methylphenyl)urea; 3-{2-methyl-3-[(octylcarbamoyl)amino]phenyl}-1-(4-methylphenyl)urea

Details on test material:

- Name of test material (as cited in study report): KY-MA
- Substance type: light yellow solid
- Physical state: solid
- Stability under test conditions: Stable in water for at least 48 hours
- Storage condition of test material: RT in the dark

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At t = 0 and t = 48h, duplicate samples of 10 mL were taken from the vehicle control and the solutions containing 10%, 32% and 100% of filtrate. aliquot from test and control vessels at t = 0h and t = 48 h. Simultaneously, reserve samples of 25 mL were taken from all test solutions.
- Sample storage conditions before analysis: -20 degrees C

Test solutions

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A supersaturated solution of 1000 mg/L nominal was prepared by stirring for 48h; Cremophor RH40 was added as dispersant. After stirring, the saturated solution was filtered and the filtrate was diluted further with ISO-medium with the dispersant Cremophor RH40 to concentrations of 10, 18, 32, 56 and 100% of the filtrate
- Controls: blank, Cremophor RH40 100 mg/L
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Cremophor RH40
- Concentration of vehicle in test medium (stock solution and final test solution): 100 mg/L
- Evidence of undissolved material (e.g. precipitate, surface film, etc): After filtering, clear without substance deposits

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Source: in-house
- Age at study initiation (mean and range, SD): < 24 hours
- Method of breeding: new batch w/ <3d old animals; 4 wk max age; renewal every 7d (half medium); daily feeding with freshwater algae

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

250 mg/L CaC03

Test temperature:

19.5-20.5 degrees C

pH:

7.8-8.1

Dissolved oxygen:

8.0-9.8 mg/L

Nominal and measured concentrations:

Blank and vehicle controls: nominal = 0 mg/L; measured = 0 mg/L
Test Substance:
Nominal = 10% of filtrate (100 mg/L); measured t0 = 0.24 mg/L, t48 = 0.13 mg/L; mean concentration = 0.19 mg/L
Nominal = 18%' of filtrate (180 mg/L); not measured; estimated mean concentration = 0.34 mg/L
Nominal = 32% of filtrate (320 mg/L); measured t0 = 0.76 mg/L, t48 = 0.48 mg/L; mean concentration = 0.62 mg/L
Nominal = 56% of filtrate (560 mg/L; not measured; estimated mean concentration = 0.70 mg/L
Nominal = 100% (1000mg/L); measured t0 = 0.69 mg/L, t48 = 0.90 mg/L; mean concentration = 0.80 mg/L

Details on test conditions:

TEST SYSTEM
- Material, size, headspace, fill volume: glass, 100 mL
- Aeration: None
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Milli-Q ultrapure water (reverse osmosis purified tap water)

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Photoperiod: 16 hours

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : imobilization (including mortality) at 24 and 48 hours

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 0.34 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: low solubility of test stubstance; significant precipitation in stock solution (pre-filtering)
- Effect concentrations exceeding solubility of substance in test medium: tests performed at above solubility; all effect concentrations were above the solubility of the test substance.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
-48h EC50: >0.32 and < 0.56 mg/L

Any other information on results incl. tables

Table: Acute immobilization of daphnia after 24 and 48 hours

Measured test concentration in mg/L (%-filtrate)	replicate	Number exposed	Number immobile at 24h	Number immobile at 48h
0	A	10	0	0
	B	10	0	0
0 + Cremophor	A	10	0	0
	B	10	0	0
0.19 (10%)	A	10	0	0
	B	10	0	0
0.34 (18%)	A	10	0	0
	B	10	0	0
0.62 (32%)	A	10	0	8
	B	10	0	6
0.70 (56%)	A	10	0	6
	B	10	0	5
0.80 (100%)	A	10	0	5
	B	10	0	4

Concentrations of 0.62 mg/L (32% filtrate) showed greater than 50% immobility, while the next lowest measured concentration (0.19 mg/L) (10% filtrate) showed no effect. A concentration between these two was not measured, though this concentration was estimated to be 0.34 mg/L, also showing no immobilization. Based on this, the EC50 value is assumed to be higher than 0.34 mg/L, which is higher than the maximum solubility of KY-MA in water (0.006 mg/L). There was no correlation between amount of filtrate and level of immobilization. The 32% filtrate showed the highest levels of immobilization at 48 hours, and had the highest start concentrations. Finally, the concentrations decreased between t0 and t48 in the 10% and 32% filtrate concentrations, but increased in the 100% concentration.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The 48h-EC50 value is higher than 0.34 mg/L, which is higher than the maximum solubility of KY-MA in water (0.006 mg/L).

Executive summary:

An Acute Immobilization Study was performed with Daphnia magna according to OECD 202. The test was performed using filtrates of a supersaturated solution of KY-MA (at 1000 mg/L). Three of the filtrate levels (10%, 32% and 100%) were then analyzed for actual KY-MA concentration.

Concentrations of 0.62 mg/L (32% filtrate) showed greater than 50% immobility, while the next lowest measured concentration (0.19 mg/L) (10% filtrate) showed no effect. A concentration between these two was not measured, though this concentration was estimated to be 0.34 mg/L, also showing no immobilization. Based on this, the 48h-EC50 value is assumed to be higher than 0.34 mg/L, which is higher than the maximum solubility of KY-MA in water (0.006 mg/L).

There was no correlation between amount of filtrate and level of immobilization. The 32% filtrate showed the highest levels of immobilization at 48 hours, and had the highest start concentrations. Finally, the concentrations decreased between t0 and t48 in the 10% and 32% filtrate concentrations, but increased in the 100% concentration.