Hexamethylene diisocyanate

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

traditional method

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Hexamethylene diisocyanate

EC Number:

212-485-8

EC Name:

Hexamethylene diisocyanate

Cas Number:

822-06-0

Molecular formula:

C8H12N2O2

IUPAC Name:

1,6-diisocyanatohexane

Constituent 2

Reference substance name:

Desmodur H

IUPAC Name:

Desmodur H

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Iffa Credo (Brussels, Belgium)
- Strain: Wistar, ICO:WU (IOPS CPB)
- Age at study initiation: 2-3 months
- Weight at study initiation: males: 173-209 g; females: 164-189 g
- Housing: individual
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): approx. 50
- Air changes (per hr): approx. 10
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure:

nose only

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: aluminium inhalation chamber (two-chamber system)
- Exposure chamber volume: about 3.8 l
- Method of holding animals in test chamber: Plexiglas exposure tubes applying a directed-flow nose-only exposure principle
- Source and rate of air: compressed air was supplied by Boge compressors; > 236 air exchanges per hour
- Method of conditioning air: air was conditioned (i.e. freed from water, dust, and oil) automatically by a VIA compressed air dryer
- Method of particle size determination: samples for the analysis of the aerodynamic particle-size distribution were taken in the vicinity of the breathing zone. The primary objective of these measurements, however, was to determine whether a condensation aerosol would occur. These samples were taken using a TSl-Laser Velocimeter APS 3300, including diluter TSI Model 3302 (TSI Inc., St. Paul, MN, USA). Particle size measurements were conducted once per exposure.
- Treatment of exhaust air: the exhaust air was purified via glass bubbler containing 10% aqueous NaOH solution and passed through cotton-wool/activated charcoal and HEPA filters. These filters were disposed of by Bayer AG.
- Temperature, humidity in air chamber: temperature and humidity measurements are also performed by the computerized HP 3852A
Data Acquisition and Control System using FTFI I sensors (ELKA ELEKTRONIK, Lüdenscheid, Germany) The position of the measuring probe was at the exhaust location. Temperature and humidity data are integrated for 30-seconds and displayed accordingly. The hurnidity sensors are calibrated using saturated salt solutions according to Greenspan (1977) and Pauluhn (1994) in a two-point calibration at 33% (MgCh) and at 75% (NaCl) relative humiduy The calibration of the temperature sensors is also checked at two temperatures using reference thermometers.

TEST ATMOSPHERE
- Brief description of analytical method used: the nominal concentration was calculated taking into account the actually evaporated mass of test substance (difference of weight of the glass bubbler before and after exposure) devided by total airflow through the chamber. The test atmosphere was determined by HPLC after derivatization of the isocyanate functionality. Samples were taken by using glass powder filled tubes containing nitroreagent as scavenging agent.
- Samples taken from breathing zone: yes

RESULTS OF PARTICLE-SIZE ANALYSES
- In the 55, 107, 120 and 151 mg/m3 exposure groups, the total mass of aerosol detected was 0.4, 6.3, 13.8 and 21.3 mg/m3, resp. based on CMAD calculation.
- Particle size distribution: In the 55, 107, 120 and 151 mg/m3 exposure groups 51, 64, 67 and 71 % ,resp. of particles were < 3 µm.
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): In the 55, 107, 120 and 151 mg/m3 exposure groups MMAD was 2.98, 2.54, 2.46 and 2.34 µm, resp. (GSD: 1.75, 1.60, 1.60 and 1.60, resp.).

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

analytical concentrations: 0, 55, 107, 120, 151 mg/m3;
nominal concentrations: 0, 64, 114, 153, 169 mg/m3

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 28 days
- Frequency of observations and weighing: clinical signs were examined several times on the day of exposure and twice daily therafter; body weights were measured before exposure, on days 3 and 7, and weekly thereafter.
- Necropsy of survivors performed: yes
- Other examinations performed: rectal temperature

Statistics:

-Necropsy flndings: If specific findings occur from the respiratory tract of surviving rats they are evaluated statistically using the pairwise Fisher test after the R x C chi-squared test.
-Body weights: Means and single standard deviations of body weights are calculated. Since in acute studies individual group means may differ prior to commencement of the first exposure, the body weight gain was statistically evaluated for each group. For these evaluations a one-way ANOVA (vide infra) is used.
-Calculation of the LC50: If calculation of a median lethal concentration (LC50) is possible, it is performed by computer (HP 3000) according to the method of AP. Rosiello, I.M. Essigmann, and G.N. Wogan (1977) as modified by Pauluhn (1983). This method is based on the maximurn-likelihood method of C.I. Bliss (1938). If only 2 pairs of values with greater than 0% lethality and less than 100% are available then the first linear approximation is based on these values and a homogeneity test is not performed. The interpolated concentration at 50% lethality in this case was designated at approximate LC50.
-Analysis of variance (ANOVA): This parametric method checks for normal distribution of data by comparing the median and mean The groups are compared at a confidence level of (1-alpha)= 95% (p=0.05) The test for the between-group homogeneity of the variance employed Box's test if more than 2 study groups were compared with each other. If the above F-test shows that the intra-group variability is greater than the inter-group variability, this is shown as "no statistical difference between the groups". If a difference is found then a pairwise post-hoc comparison is conducted (1- and 2-sided) using the Games and Howell modification of the Tukey-Kramer significance test.

Results and discussion

Effect levels

Sex:

male/female

Dose descriptor:

LC50

Effect level:

124 mg/m³ air

95% CL:

>= 111 - <= 140

Exp. duration:

4 h

Remarks on result:

other: NOAEL: < 55 mg/m3 air

Mortality:

Exposure to vapour concentrations of 55 mg/m3 did not induce test substance related mortality. Exposure to 107 mg/m3 air and above was followed by a characteristic type of mortality pattern occurring within post exposure days 1 through 6. Marked gender specific differences in susceptibility was not observed (details see table 1).

Clinical signs:

other: see "Other findings"

Body weight:

Decreased body weights were observed in all groups exposed to the test compound (at 55 mg/m3 and above).

Gross pathology:

Gross necropsy (among others) revealed less collapsed, discolorated (dark-red) lungs with serous mucus in the trachea. The lung associated lymph nodes were enlarged.

Other findings:

All animals showed normal reflexes. One rat of group 5 (151 mg/m3) showed mildly changed reflexes due to its moribund state.
Concentration-dependent effect on body temperature were observed at 55 mg/m3 and above (details see table 1).

Clinical Signs:
Exposures to concentrations of 55 mg/m3 and above were followed by concentration-dependent signs indicative of respiratory tract irritation, suchas bradypnea, dyspnea, laboured breathing pattern, rales, nostril/snout area with red encrustations, cyanosis, prostration (lying on belly), reduced motility, ungroomed haircoat, hypothermia, decrease in body weights, and piloerection. The duration of signs (maximum duration up to day 28) and, in most instances, was dependent on respiratory signs (see also table 1 for onset and duration of signs).

Any other information on results incl. tables

Table 1: Acute inhalation toxicity (vapour) of HDI

Sex	Analytical concentration (mg/m3)	Toxicological results	Onset and duration of signs	Rectal temperature(°C)	Onset of mortality
male	0	0 / 0 / 5	---	38.0	---
	55	0 / 5 / 5	0d - 8d	30.7 *	---
	107	2 / 5 / 5	0d - 28d	30.2 *	1d
	120	1 / 5 / 5	0d - 28d	28.2 *	2d
	151	5 / 5 / 5	0d - 2d	28.1 *	1d - 3d
female	0	0 / 0 / 5	---	38.4	---
	55	0 / 5 / 5	0d - 7d	31.2 *	---
	107	1 / 5 / 5	0d - 14d	29.6 *	2d
	120	3 / 5 / 5	0d - 28d	28.2 *	2d - 4d
	151	3 / 5 / 5	0d - 28d	27.6 *	3d - 6d

Toxicological results:

number of dead animals / number of animals with signs after cessation of exposure / number of animals exposed

* p<0.01 (Tukey-Kramer post hoc test)

Applicant's summary and conclusion

Executive summary:

Exposure to vapour concentrations of 55 mg/m3 did not induce test substance related mortality. Exposure to 107 mg/m3 air and above was followed by a characteristic type of mortality pattern occurring within post exposure days 1 through 6. Marked gender specific differences in susceptibility could not be observed. Exposures to concentrations of 55 mg/m3 and above were followed by a concentration-dependent signs indicative of respiratory tract irritation, such as bradypnea, dyspnea, laboured breathing pattern, rales, nostrils/muzzle with red encrustations, cyanosis, prostration (lying on belly), reduced motility, ungroomed haircoat, hypothermia, decreased body weights, and piloerection. The duration of signs (maximum duration up to day 28), in most instances, was dependent on respiratory signs. Gross necropsy (among others) revealed less collapsed, discolorated (dark-red) lungs with serous mucus in trachea. The lung associated lymph nodes were enlarged. Clinical observations and necropsy findings support the conclusion that a causal relationship between lethality and lung damage existed.

The evaporated test substance proved to have a high acute inhalation toxicity to rats (LC50 - vapour, 4 h: 124 mg/m3 air). Cumulative evidence suggest that there is a causal relationship of local effects to the respiratory tract and the observed findings.