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Respiratory sensitisation

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Administrative data

Endpoint:
respiratory sensitisation: in vivo
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable well-documented publication, which meets basic scientific principles

Data source

Reference
Reference Type:
publication
Title:
Provocation of Respiratory Allergy in Guinea Pigs Following Inhalation of Free Toluene Diisocyanate
Author:
Aoyama, K.,Huang, J., Ueda, A.,Matsushita, T.
Year:
1994
Bibliographic source:
Arch. Environ. Contam. Toxicol. 26, 403-407 (1994)
Report Date:
1993

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
An animal exposure experiment, simulating a workplace exposure situation was made to compare toluene diisocyanate (TDI) concentrations which resulted in antibody production with those which elicited pulmonary responses. Groups of guinea pigs were exposed to inhaled TDI from 0.02 to 1.0 ppm (µg/g) for 3 h/day on 5 consecutive days. Three weeks later the animals were challenged with 0.02 ppm of free TDI for 15 min. TDI specific antibodies and pulmonary responses were evaluated. Using an experimental model in which animals were sensitized and challenged by inhalation of free TDI, the present study was performed in order to compare the TDI concentration which resulted in antibody production with that which elicited a pulmonary response, and secondly, to provide an experimental basis for setting up OELs of chemical sensitizers.
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Toluene diisocyanate
- Molecular formula (if other than submission substance): C9H6N202
- Molecular weight (if other than submission substance): 174.15 g/mol
- Smiles notation (if other than submission substance): CC(C)c1c(N=C(=O))c(C(C)C)c(N=C(=O))c(C(C)C)c1
- Structural formula attached as image file (if other than submission substance): see Fig.
- Substance type: aromatic diisocyanate
- Physical state: liquid
- Isomers composition: 80/20 mixture of the 2,4- and 2,6-isomers

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS:
- Source:
- Age at study initiation:
- Weight at study initiation: 300-350 g
- Housing: group housed two per cage
- Diet (e.g. ad libitum): food ad libitum
- Water (e.g. ad libitum): water ad libitum
- Acclimation period: 1 week

Test system

Route of induction exposure:
inhalation
Route of challenge exposure:
inhalation
Remarks:
induction: whole body, challenge head-nose-only
Vehicle:
other: clean dry air
Concentration:
Induction: 0 ppm, 0.02 ppm, 0.2 ppm, 0.6 ppm, 1 ppm
Challenge: 0.02 ppm
No. of animals per dose:
6 females
Details on study design:
RANGE FINDING TESTS:

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 5
- Exposure period: 5 days
- Test groups: 5
- Control group: 1
- Site: not applicable inhalation
- Frequency of applications: once daily
- Duration: 3 h
- Concentrations: 0 ppm, 0.02 ppm, 0.2 ppm, 0.6 ppm and 1 ppm

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 1
- Exposure period: 15 min
- Test groups: Groups 2 - 6 induced with 0.02 ppm, 0.2 ppm, 0.6 ppm and 1 ppm
- Control group: Group 1 - no induction = 0 ppm TDI
- Site: not applicable inhalation
- Concentrations: 0.02 ppm were used for challenge
- Evaluation (hr after challenge): during challenge and up to 60 min after challenge
- Other: Twenty-six days after the first induction exposure, the animals were placed in individual body plethysmographs with the head of each animal extending through a latex dam into an inhalation chamber, and were challenged by exposure to 0.02 ppm TDI for 15 min. The concentration of TDI at challenge was chosen because a short exposure to that level was previously shown not to be an irritant to guinea pigs.

OTHER:
The actual TDI concentrations (2-+- SD) were 1.128 +/- 0.125, 0.64 +/- 0.078, 0.218 +/- 0.046 and 0.022 +/- 0.003 ppm at induction, and 0.019 +/- 0.002 ppm at challenge.

TDI aerosol was generated by passing air through a midget impinger containing TDI solution and was led into the inhalation chamber after dilution with clean, dry air to achieve the appropriate concentration. TDI concentrations were determined by the method of Marcali (1957).
Challenge controls:
Six animals of an additional group were sham-exposed.
Positive control substance(s):
toluene diisocyanate (TDI)
Negative control substance(s):
other: clean dry air

Results and discussion

Any other information on results incl. tables

Table2. Pulmonary responses following ; inhalation challenge to free toluene diisocyanate (TDI)
Group Induction conc,(ppm) Challengeaconc, (ppm) Pulmonary responsea Responding animals/total
I 1.0 0.02 145±12 5/6
II 0.6 0.02 161±40 4/6
III 0.2 0.02 168±32 6/6
IV 0.02 0.02 120±3 0/6
V 0 0.02 114±7 0/6
ax ±SD of % pre-challenge respiratory rate demonstrated in individual animals. Percent pre-challenge rate used here was derived from the most severe response exhibited in individual animals during an observation period.
Production of Specific Antibodies

The time course of IgG antibody production was investigated by ELISA. Serum which gave an absorbance of 0.054 or greater was defined as having a positive IgG response to TDI.

IgG antibodies were detected 6 days after the first exposure in several animals of the 1.0, 0.6 and 0.2 ppm groups (4/6, 2/6, and 2/6, respectively). At 13 days, the antibodies were detected in all animals of these groups. The peak mean absorbance in these groups was reached 20 days following the first exposure. However, IgG antibodies were not detected from animals exposed to 0.02 ppm and control animals. PCA titres showed a linear correlation to TDI concentration at induction, as did IgG antibody production (p < 0.01). The findings were consistent with the results of our previous study using TDI-GSA conjugates at challenge. IgE antibodies were not detected from any animal.

Pulmonary Response

Animals were challenged by inhalation for 15 min of 0.02 ppm free TDI 26 days after the first induction exposure. Pulmonary responses are shown in Table 2. None of the six animals exposed to 0.02 ppm TDI at induction showed significant pulmonary responses (Group IV). By contrast, most of the animals exposed to TDI above 0.2 ppm (1.0, 0.6 and 0.2 ppm groups) displayed significant pulmonary responses. There was no linear correlation between TDI concentration at induction and the intensity of pulmonary response upon challenge to free TDI (p > 0.05). Further, no significant difference in the proportion of animals showing a pulmonary response was found among these three groups (p > 0.05, Fisher's test).

Comparison of Antibody Production and Pulmonary Response

A comparison was made between antibody production and the experience of pulmonary response. As presented in Table 3, the proportion of animals showing a pulmonary response was not closely related to the antibody production. Although animals which had no antibody production did not display pulmonary responses, animals with high antibody production did not inevitably exhibit pulmonary responses. In contrast, some of the animals showing low antibody production were found to give pulmonary responses.

Table3. Comparison of antibody production and pulmonary response in guinea pig exposed to toluene diisocyanate (TDI)
Group Induction conc, (ppm)  ELISA absorbance a PCA titerb Pulmonary responsec
I 1.0 0.285 29 -
0.262 29 +
0.237 28 +
0.228 27 +
0.187 27 +
0.175 26 +
II 0.6 0.274 2io +
0.208 29 +
0.222 26
0.195 2s ++
0.093 25 +
0.171 24 -
III 0.2 0.190 26 +
0.156 25 +
0.153 24 +
0.130 24 + +
0.103 23 +
0.068 22 +
IV 0.02 0.030 nd -
0.028 nd -
0.027 nd -
0.015 nd -
0.003 nd -
0.009 nd -
V 0 0.038 nd -
0.041 nd -
0.001 nd -
0.001 nd -
0.015 nd -
0.003 nd -
nd=Not detected
a An absorbance/>0.054 (2 + 2SD of control responses) was defined as IgG antibody positive
 bTitre of IgG 1 antibodies
cResponse (+): an increase in respiratory rate to 127 % (Mean +/- 3SD of control response) or more of the pre-challenge rate within the challenge and 60 min after challenge. (++): 193 % (Mean +/- 10 SD of control response) or more. (-): 126 % or less.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Conclusions:
The publication is based on experiments with guinea pigs exposed via inhalation to toluene diisocyanate and is evaluating the potential of causing respiratory sensitisation. It is considered to be of high quality (reliability Klimisch 2). The validity criteria of the test system are fulfilled. The test material did induce sensitisation and should be classified accordingly.
Executive summary:

Aoyama and co-workers conducted an animal exposure experiment, in which animals were sensitized and challenged by inhalation of free TDI, so

simulating a workplace exposure situation to compare toluene diisocyanate (TDI) concentrations which resulted in antibody production with those which elicited pulmonary responses and secondly, to provide an experimental basis for setting up OELs of chemical sensitizers. Therefore groups of guinea pigs were exposed to TDI from 0.02 to 1.0 ppm for 3 h/day on 5 consecutive days. Three weeks later the animals were challenged with 0.02 ppm of free TDI for 15 min. TDI specific antibodies and pulmonary responses were evaluated. Specific antibody production showed a linear correlation to TDI concentration at induction. An induction level of 0.02 ppm TDI failed to stimulate antibody production in animals. Most of the animals exposed to TDI levels above 0.2 ppm displayed significant pulmonary responses, but no correlation was found between TDI concentration at induction and the intensity of pulmonary response upon challenge to free TDI. These results indicated that there was a threshold concentration of 0.02 ppm TDI for antibody production and for the development of pulmonary response. Although it failed to demonstrate sensitizing potency, 0.02 ppm free TDI did elicit a pulmonary response in some of the animals sensitized to TDI at levels ranging from 0.2 to 1.0 ppm. It was also found that exposure to TDI at a level lower than its threshold concentration for sensitisation may elicit a response in previously sensitized individuals. The intensity of pulmonary response was not notably influenced by TDI concentration at induction levels above 0.2 ppm. This result may be related to several factors such as solubility, hapten concentration, or size of hapten. This finding is also of great importance in extrapolating from the results of an animal model to an industrial situation. However, free TDI was able to elicit immediate-onset reactions, within 60 min following the challenge. No close association was seen between antibody production and the experience of pulmonary response. High antibody production did not stick to the development of pulmonary response. On the other hand, several animals with low antibody titres displayed positive pulmonary responses. This discrepancy might be derived from differences in the ability to release histamine due to the different distribution of specific antibodies in the lungs of animals.