Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

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:
other: GLP guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1999
Report Date:
1999
Reference Type:
publication
Title:
Developmental toxicity of 1,6-Hexamethylene diisocyanate (HDI) in the Sprague-Dawley rat
Author:
Astroff AB et al.
Year:
2000
Bibliographic source:
Teratology 62: 205-213

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Labs, Raleigh, NC
- Age at study initiation: 12-15 wks
- Weight at study initiation: Females: 237-325 g
- Housing: individual
- Diet: ad libitum (except during the exposure period)
- Water: ad libitum
- Acclimation period: at least 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 26
- Humidity (%): 30 - 70
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
other: air
Details on exposure:
TEST SUBSTANCE GENERATION:
HDI was generated as a vapor by passing filtered, dry air through liquid HDI in a grass bubbler. During vapor generation the bubbler containing HDl was immersed in a constant temperature water bath. The vaporized material was entrained with chamber intake air flow for mixing at the chamber head. Both bubbler temperature and air flow may have been adjusted to maintain desired chamber HDl concentrations and these parameters were monitored continuously with recordings at half-hour intervals (minimum) during eaeh six-hour exposure period.

EXPOSURE SYSTEM:
Chambers: The chambers used in this study were Hazleton H-2000 inhalation exposure chambers which are constructed of stainless steel with clear glass windows. Each chamber has an approximate volume of two cubic meters. The chambers are equipped with stainless steel, wire mesh cage-packs. Each cage-pack is fitted with removable feed troughs and an automatic watering system. The air supplied to the chamber passes through an activated charcoal trap and a HEPA filter before being conditioned to the desired temperature and relative humidity. These chambers have been used
previously for exposure of animals to HDI.

NOMINAL CHAMBER PARAMETERS (During Exposure):
Temperature: 22 ± 2°C; Relative Humidity: 50 ± 10%; Exhaust Flow: 650 ± 50 Lpm; Static Pressure: -0.25 to -1.0 inches of water relative to atmospheric.
To the extent possible these nominal values were maintained during each exposure period. During non-exposure periods (nights) nominal values for each chamber parameter were set to be maintained as Iisted above, with the exception that the range for RH shall be relaxed to be 40 - 70%. This was to accommodate expected inereases in RH due to chamber handling and animal care. The increase to 70% RH is in accordanee with AALAC guidelines governing care of rats.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Chamber samples were collected near the animal's breathing zone using two midget impingers connected in series. Samples were collected at a frequency that ensured that the average daily value was representative of the required concentration. At a minimum, three samples (one for the control
chamber) were collected per chamber per day. An acetonitrile solution (at least 10 mL per impinger) containing N-4-nitrobenzyl-N-n-propylamine (nitro reagent) was used to trap and derivatize HDl to a UV-absorbing compound. All midget impinger samples were assayed by an established high performance liquid chromatography method. The homogeneous distribution of HDI within the inhalation chambers has been confirmed in previous studies, under the conditions utilized in the present study.
Details on mating procedure:
Rats were co-housed with a maximum of two females per male. Following cohabitation, morning vaginal smears were taken and examined for the presence of sperm. Females found to be sperm-positive were randomized into groups and the sire number was documented. The day on which sperm was observed in the vaginal smear was designated day 0 of gestation for that female.
Duration of treatment / exposure:
days 0 - 19 of gestation
Frequency of treatment:
once daily for 6 hours
Duration of test:
cesarean section and sacrifice at day 20 of gestation
No. of animals per sex per dose:
30 females per dose
Control animals:
yes, sham-exposed
Details on study design:
DOSE SELECTION RATIONALE:
The concentrations of HDl used in this study were based on a 21-day inhalation toxicity study, a 90day inhalation toxicity study, a chronic inhalation toxicity/oncogenicity study, and a sensory irritation study. In the 21-day study, Sprague-Oawley rats were exposed to either 0, 0.005, 0.0175,
0.15, or 0.3 ppm HDl for 5 hours/day, 5 days/week for 3 weeks. Compound-related ocular and nasal irritation were observed in animals exposed to 0.0175, 0.15, and 0.3 ppm on days of exposure only. These findings were not observed during non-exposure days. There were no compound-related effects on body weight, feed consumption, clinical chemistry, hematology, urinalysis, or gross pathology. At 0.3 ppm, liver and kidney weights were decreased in females. The major findings for both sexes were histopathologie lesions of the nasal mucosa and minor changes in the larynx and trachea. This study demonstrated that the target site following HDl exposure was the nasal cavity. In the 90-day study, Fischer 344 rats were exposed to HDl concentrations of 0, 0.01, 0.04 and 0.14 ppm for 6 hours/day, 5days/week for approximately 13 weeks. The only compound-related findings were ocular irritation and histopathologic lesions of the anterior nasal cavity. Both findings were observed at all three concentrations, therefore, a clear NOEL was not established in this study. In the chronic/oncogenicity study, Fischer 344 rats were exposed to HDl concentrations of 0, 0.005, 0.025 and 0.175 ppm for 6 hours/day, 5 days/week for up to 2 years. Animals were evaluated following both one and two years of exposure. A maximum tolerated dose was achieved at the highest concentration based on decreased body weight and slight anemia in the females, and histopathologie lesions of the nasal cavity in both sexes. The lowest concentration (0.005 ppm) was shown to be a NOEL after one year of exposure. However, after two years of exposure 0.005 ppm was considered to be a NOAEL based on the observation of reversible lesions, indicative of responses to non-specific irritation. In the sensory irritation study, female Sprague-Dawley rats were exposed using the head-only technique, to 0, 0.10, 0.21, 0.79, and 4.42 ppm Mondur HX (100% HDl) for three hours. Following exposure the animals were held for a seven-day recovery period. A concentration dependent increase in the respiratory response (sensory irritation) was observed. The severity of the response culminated in the death of two rats at the 4.42 ppm dose level. The RD50 (concentration which was estimated to produee a 50% depression in respiratory frequency) for the last hour of a three-hour exposure was 1.69 ppm. The NOEL for this study was 0.1 ppm. Based on these results, and the projected exposure of the animals for approximately three weeks during the current study, the proposed concentrations were 0, 0.005, 0.05, and 0.3 ppm HDI.

Examinations

Statistics:
The data were analyzed with the litter as the primary experimental unit using applications provided by TASC. Parametric data (including dam body weights) were analyzed using an Analysis of Variance (ANOVA), and if significant differences were observed, a Dunnett's Test was performed. Fetal and
placental weights were specifically analyzed via the Healy's Test if significance was observed in the ANOVA. Nonparametric data (e.g. Iitter size and number of corpora lutea) were first analyzed by the Kruskal-Wallis test and then subjected to Dunn's Test if significant differences were identified. Nonparametric dichotomous data (e.g. number normal/abnormal) were initially analyzed by the Chi-Square Test and if significance was observed between groups then by the Fisher's Exact Test with the Bonferroni adjustment. A p-value less than or equal to 0.05 was considered statistically significant.
Historical control data:
yes

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
0.300 ppm:
No mortality; no clinical signs; no test compound related effects on maternal body weight, uterine weight, and net body weight; microscopic changes within the nasal cavity.
0.050 ppm:
No mortality; no clinical signs; no test compound related effects on maternal body weight, uterine weight, and net body weight; microscopic changes within the nasal cavity (to a lesser extent compared to the 0.300 ppm exposure group).
0.005 ppm:
No mortality; no clinical signs; no test compound related effects on maternal body weight, uterine weight, and net body weight; no microscopic changes.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEC
Effect level:
0.005 ppm (nominal)
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEC
Effect level:
0.3 ppm (nominal)
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
All dose groups:
No effects on reproductive parameters; no embryotoxicity; no litter effects; no fetal external, visceral, and skeletal malformations.

Effect levels (fetuses)

Dose descriptor:
NOAEC
Effect level:
0.3 ppm (nominal)
Basis for effect level:
other: teratogenicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Maternal toxicity was demonstrated in the 0.300 and to a lesser extent in the 0.050 ppm exposure groups. No maternal effects were noted in the 0.005 ppm dose group. Test compound-related maternal effects were restricted to histopathological findings, and included acanthosis, hyperkeratosis, inflammation of the nasal turbinates, and more seriously, degeneration of the olfactory epithelium. No pathological alterations were noted in the larynx, trachea, or lungs in any dose group. No test compound-related effects were observed on any reproductive parameters, or any embryonic endpoints, including pre/post-implantation loss and resorptions. There were no effects on litter size or the number of fetuses per implantation site and no effects on fetal or placental weights were observed. No test compound-related fetal external, visceral, or skeletal findings were observed. No effect on the fetal or litter incidence of total malformations or variations was observed and there was no difference in the incidence of malformations between males and females.

1,6-Hexamethylene diisocyanate (HDI), administered as described in this study, produced maternal effects (nasal turbinate histopathology) at concentrations of 0.050 and 0.300 ppm. No developmental toxicity was observed at any concentration level. Therefore, the maternal no-observed-effect-level (NOEL) was 0.005 ppm HDI and the developmental NOEL was 0.300 ppm HDI.

Analytically confirmed overall (for the entire study) mean HDI vapour concentrations were 0.005, 0.052 and 0.308 ppm.

Applicant's summary and conclusion

Executive summary:

In a developmental toxicity study (OECD TG 414) with 1,6-hexamethylene diisocyanate (HDI) rats were exposed, via whole-body exposure, to HDI vapour concentrations of 0, 0.005, 0.050, or 0.300 ppm for 6 hours/day on days 0 through 19 of gestation. Analytically confirmed overall (for the entire study) mean HDI vapour concentrations were 0.005, 0.052 and 0.308 ppm. Maternal toxicity was demonstrated in the 0.300 and to a lesser extent in the 0.050 ppm exposure groups. No maternal effects were noted in the 0.005 ppm dose group. Test compound-related maternal effects were restricted to histopathological findings, and included acanthosis, hyperkeratosis, inflammation of the nasal turbinates, and more seriously, degeneration of the olfactory epithelium. No pathological alterations were noted in the larynx, trachea, or lungs in any dose group. No test compound-related effects were observed on any reproductive parameters, or any embryonic endpoints, including pre/post-implantation loss and resorptions. There were no effects on litter size or the number of fetuses per implantation site and no effects on fetal or placental weights were observed. No test compound-

related fetal external, visceral, or skeletal findings were observed. No effect on the fetal or litter incidence of total malformations or variations was observed and there was no difference in the incidence of malformations between males and females.

In summary, HDI produced maternal effects (nasal turbinate histopathology) at concentrations of 0.050 and 0.300 ppm. No developmental toxicity was observed at any concentration level. Therefore, the maternal no-observed-effect-level (NOEL) was 0.005 ppm and the developmental NOEL was 0.300 ppm.