Lactic acid, compound with 3-[2-(dimethylamino)ethyl] 1-(2-ethylhexyl) toluene-2,4-dicarbamate (1:1)

Additional toxicological data

Administrative data

Endpoint:

additional toxicological information

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1st October to 10th October 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

No Guideline available however scientific principles of the study are well documented. This study was selected as the key study because the information provided for the hazard endpoint is sufficient for the purpose of classification and labelling and/or risk assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

Materials and methods

Type of study / information:

The Neutral Red Uptake Bioassay was used to assess the toxicity of the test article to Balb/c 3T3 mouse fibroblasts (3T3) (American Type Culture Collection, Manassas, VA). The assay is an adaptation of the procedure described by Borenfreund and Puerner', and Babich, et al. The neutral red uptake bioassay is used to quantitatively measure the toxicity of a test article to Balb/c 3T3 cultures by comparing the neutral red dye (3-amino-7-dimethylamino-2-methyiphenazine hydrochloride) uptake of test article treated Balb/c 3T3 cultures to the neutral red uptake in control (media or solvent treated) Balb/c 3T3 cultures. The concentration of test article causing a reduction in neutral red uptake of 50% (NRU50) was determined.

The purpose of this study was to evaluate the toxicity of the test article, supplied by PPG Industries, Inc., as measured by the test article induced inhibition of neutral red uptake in Balb/c 3T3 cultures. The laboratory phase of this study was conducted from October 1, 2003 to October 10, 2003 at the Institute for In Vitro Sciences, Inc (IIVS). After a preliminary assay, the test article was tested in a valid definitive assay (eight dose levels) to determine the concentration of the test article resulting in the NRU50 end point.

Test guideline

Qualifier:

no guideline available

Principles of method if other than guideline:

n/a

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Lactic acid, compound with 3-[2-(dimethylamino)ethyl] 1-(2-ethylhexyl) toluene-2,4-dicarbamate (1:1)

EC Number:

269-358-5

EC Name:

Lactic acid, compound with 3-[2-(dimethylamino)ethyl] 1-(2-ethylhexyl) toluene-2,4-dicarbamate (1:1)

Cas Number:

68227-46-3

Molecular formula:

C21H35N3O4.C3H6O3

IUPAC Name:

2-hydroxypropanoic acid 2-(dimethylamino)ethyl N-[5-({[(2-ethylhexyl)oxy]carbonyl}amino)-2-methylphenyl]carbamate

Details on test material:

- Name of test material (as cited in study report): WRS-2390
- Physical state: Vicous liquid
- Storage condition of test material: Stored at room temperature and protected from light.

Results and discussion

Any other information on results incl. tables

Definitive Neutral Red Uptake Assay: 

The below tables gives the NRU₅₀ results for the definitive neutral red uptake assay for the test article and the positive control. The NRU₅₀ value of the positive control, SLS, was within the expected range seen historically.

Test Articles	Sponsor designation	NRU50 (µg/ml)		pH (712 mg/mL in water)	pH* (20 mg/mL in CDM)
		Preliminary (10/1/03)	Trial 1 (10/8/03)
03AH00	WRS-2390	5.5	18.2	NCC	8.5
SLS	Positive Control	25.5	36	NA	NCC

* - pH of highest test article concentration prepared in Chemical Dilution Medium

NA - Not Applicable

NCC - Not able to be determined, since a color change was not observed on the pH paper.

The following calculation was performed to determine the estimated LD₅₀ of the test article based on the NRU₅₀ result of the definitive assay. The regression formula is based on the results of testing the reference chemicals with mouse fibroblasts in the BALB/c 3T3 Neutral Red Uptake Cytotoxicity test, published in the Guidance Document on Using in vitro Data to Estimate

in vitro Starting Doses for Acute Toxicity (August 2001, NIH Publication No. 01-4500).

Log (LD₅₀) = 0.435 x log (NRU₅₀ in mmol/L) + 0.625 

NRU50 = 18.2 µg/mL = 0.0182 mg/mL

Formula Weight of WRS-2390 = 483

NRU50 in mmol/L = 0.037681 mmol/L

Estimated log (LD₅₀) = 0.00561 mmol/kg
Estimated LD₅₀ = 1.01 mmol/kg = 489 mg/kg

Applicant's summary and conclusion

Conclusions:

The Estimated LD50 = 1.01 mmol/kg which is equivalent to 489 mg/kg. The study and the conclusions which are drawn from it fulfil the quality criteria (validity, reliability & repeatability).

Executive summary:

In a Neutral red uptake bioassay in balb/c 3T3 mouse fibroblast study (Institute for In Vitro Sciences, Inc. Study number: 03AH00.100068) the test material had an estimated LD50 of 1.01 mmol/kg, which is equivalent to 489 mg/kg.