m-tolylidene diisocyanate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

maternal body weight measured more frequently

Principles of method if other than guideline:

- Principle of test: Overall method conforms to OECD test guideline 414, except maternal body weight measured more frequently at gestation days 0, 6, 9, 12, 16, and 21.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

m-tolylidene diisocyanate

EC Number:

247-722-4

EC Name:

m-tolylidene diisocyanate

Cas Number:

26471-62-5

Molecular formula:

C9H6N2O2

IUPAC Name:

2,4-diisocyanato-1-methylbenzene, 2,6-diisocyanato-1-methylbenzene

Details on test material:

- Name of test material (as cited in study report): Toluene diisocynate
- Molecular formula (if other than submission substance): Benzene, 2,4- or 2,6-diisocyanateo-1-methyl; CH3C6H3 (NCO)2]
- Substance type: diisocynate
- Physical state: liquid
- Analytical purity: approximately 99%
- Impurities (identity and concentrations): no
- Composition of test material, percentage of components: 80%:20% mixture of the 2,4-TDI and 2,6-TDI
- Isomers composition: 80% 2,4- and 20% 2,6- isomers
- Lot/batch No.: 2,4-TDI and 2,6- TDI (CAS No. 584-84-9 and 91-08-7, respectively)
- Stability under test conditions: Stable, no significant compositional changes occurred while the study progressed and the test material remained approximately 79% 2,4- and 20% 2,6-TDI throughout the study.

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: Stable, no significant compositional changes occurred while the study progressed and the test material remained approximately 79 % 2,4- and 20 % 2,6-TDI throughout the study.

OTHER SPECIFICS:
- Physical state: liquid
- Composition of test material, percentage of components: 80 %:20 % mixture of the 2,4-TDI and 2,6-TDI

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Kingston, NY, USA
- Age at study initiation: ♂ 11 weeks, ♀ 10 weeks
- Weight at study initiation: ♀ gd 0: 220.6 - 240.2
- Fasting period before study: No
- Housing:
Animals during acclimation were housed separated by sex, 2 -3 animals per cage in stainless-steel wire-mesh cages.
During mating, rats were housed 1:1 (one male : one female) in stainless steel wire mesh cages.
Plug-positive females were individually housed for the duration of the study (except during exposure). For exposures, plug-positive females were transferred to stainless steel wire-mesh exposure cage.
- Diet: ad libitum, chow, except during exposures.
- Water: Tap water was available ad libitum except during exposures.
- Acclimation period: Two weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): measured but ranges not given
- Humidity (%): measured but ranges not given
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
From: 1987-03-29
To: 1987-04-20/22

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 4 exposure chambers
- Method of holding animals in test chamber: The animals were exposed to the test material in cages placed in the chamber.
- Temperature and humidity in air chamber: Temperature (22.6 - 23.9 °C) and relative humidity (50.1 - 54.4 %) for all four chambers.
- Air flow rate: approximately 1000 - 1500 L/minute with a t99 (theoretically-derived time required for the chamber to reach 99% of the equilibrium concentration) of ~20 minutes.
- Air change rate: at least 14 air changes/h.

TEST ATMOSPHERE
- Brief description of analytical method used:
Liquid TDI was metered from a syringe pump into a heated glass evaporator. The temperature in the evaporator was monitored and maintained at the lowest level sufficient to vaporise the liquid (46 - 60 °C). The resultant vapour was carried into the chamber (4320 L chambers) by dilution air stream. Each chamber atmosphere was analysed for TDI approximately six times during each 6-hour exposure. Daily nominal concentrations and chamber airflow during the exposure period were calculated for each chamber. Chamber concentrations were monitored via Autostep Portable Monitors. In addition, chamber atmosphere samples were drawn through midget impingers and samples measured via UV.

VEHICLE
- Vehicle: Countercurrent air stream

Analytical verification of doses or concentrations:

yes

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Length of cohabitation: 3 days (maximum)
- Further matings after two unsuccessful attempts: no
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

gestation day 6 - 15

Duration of test:

21 days (days 0-21 of gestation)

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0.02 ppm (nominal)

Remarks:

0.021 ± 0.0018 ppm (analytical), 105 % of target

Doses / concentrations 2

Dose / conc.:

0.1 ppm (nominal)

Remarks:

0.12 ± 0,017 ppm (analytical), 120 % of target

Doses / concentrations 3

Dose / conc.:

0.5 ppm (nominal)

Remarks:

0.48 ± 0.038 ppm (analytical), 69 % of target

No. of animals per sex per dose:

25 Pregnant females rats/group/dose level

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale:
To determine appropriate target concentrations of TDI, a range-finding study was performed following the same study design in the definitive study, except eight inseminated females/group were employed. The concentrations were 0.00, 0.1, 0.50, and 1.0 ppm and the dams were terminated after the last exposure on gd 15. At termination, immediately after the last exposure, the dams were anesthetised with methoxyflurane (Metofan, Pittman Moore) and arterial blood was removed from the abdominal aorta into arterial blood samples containing heparin as an anticoagulant and analysed by Corning 170 Blood Gas analyser, Corning NY. Blood samples were analysed for pO2, pCO2 and pH and bicarbonate calculated using (Corning 170 Blood Gas analyser, Corning NY).
The dams were examined for body weight, gravid uterine weight, liver weight, number of ovarian corpora lutea, number of uterine implantation sites (total early and late absorptions and dead implants and live implants.
At 1.0 ppm: maternal body weights, weight changes, and liver weights (absolute but not relative) were profoundly and significantly reduced. Audible respiration and nasal discharge were observed; stomach and intestines were gas filled and adrenal glands were enlarged.
These were no effects on gestational parameters. The maternal blood gas results include the statistically significant changes at 1.0 ppm.
Based on these results 1.0 ppm was deemed too toxic for use in the definitive study. Therefore, the target exposure concentrations for the definitive study were 0.00, 0.02, 0.10, and 0.50 ppm.

Examinations

Maternal examinations:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Maternal clinical signs were taken daily.

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were measured on gestation day 0, 6, 9, 12, 16, and 21.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Food consumption was measured every 3 days.

WATER CONSUMPTION: Yes
- Time schedule for examinations: - Water consumption was measured every 3 days.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: yes

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:external, soft tissue, skeletal
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- External examinations: Yes:
All live fetuses were anesthetised by hypothermia, and were weighed, sexed and examined for external malformations including cleft palate, and variations.

- Soft tissue examinations: Yes:
Approximately ½ of the live fetuses in each litter were examined for thoracic and abdominal visceral abnormalities.

- Skeletal examinations: Yes:
Intact fetuses in each litter were eviscerated, fixed in ethanol, and then processed for skeletal staining and examined for skeletal malformations and variations.

- Head examinations: Yes:
The fetuses examined viscerally were decapitated and their heads fixed in Bouin's solution for examination of craniofacial structures.

Statistics:

Quantitative continuous variables were intercompared via Levene's test, ANOVA, and T-tests with Bonferroni probabilities. Nonparametric data were analyzed using the Kruskal-Wallis test followed by the Mann-Whitney U test. Incidence data were compared using Fisher's Exact Test. The fiducial limit of 0.05 (two-tailed) was used as the criterion for significance.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Description (incidence and severity):

Treatment-related clinical signs of toxicity were limited to audible respiration in dams at 0.5 ppm and red nasal discharge at 0.02, 0.1, and 0.5 ppm.

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Maternal weight gain was significantly depressed over the exposure period for 0.5 ppm group. Maternal weight gain was significantly reduced early in the exposure period, gd 6 - 9, at 0.02 and 0.1 ppm.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Maternal food consumption was reduced at the high dose from exposure day 9 onward. In addition, food consumption was reduced at 0.02 ppm for gd 18 - 21 due to the dam with one fully resorbed litter eating at a nonpregnant rate in this group.

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Maternal developmental toxicity

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Details on maternal toxic effects:

Maternal toxic effects: yes

Details on maternal toxic effects:
Maternal weight gain was significantly depressed over the exposure period for 0.5 ppm group. Significant body weight reduction, significant body weight gain reduction and significantly reduced food consumption were observed.

Effect levels (maternal animals)

Key result

Dose descriptor:

NOAEC

Effect level:

0.1 ppm

Based on:

test mat.

Basis for effect level:

body weight and weight gain

Results (fetuses)

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

An increased incidence of poorly ossified cervical centrum 5 at 0.5 ppm was reported (exhibited a statistically significant increase relative to controls), indicating minimal fetotoxicity or this variation could be described as a common rat skeletal variation.

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects: no effects

Details on embryotoxic / teratogenic effects:
No embryotoxicity or teratogenicity was observed at any exposure concentration employed.

Effect levels (fetuses)

Key result

Dose descriptor:

NOAEC

Effect level:

0.1 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

skeletal malformations

Fetal abnormalities

Key result

Abnormalities:

no effects observed

Overall developmental toxicity

Key result

Developmental effects observed:

no