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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
repeated dose toxicity: inhalation
Remarks:
other: Carcinogenicity study
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
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Long-term toxicity and carcinogenicity studies with 2.4/2.6 toluene diisocyanate (80/20) in rats and mice
Author:
Loeser, E.
Year:
1983
Bibliographic source:
Toxicology Letters, 15 (1983) 71-81
Report Date:
1982

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: OECD 451 - Carcinogenicity studies
Deviations:
yes
Remarks:
6-9 weeks of age at study initiation, only 21 animals per sex/level, no detailed information on housing, diet and water, only 2 dose levels tested, no detailed information which tissues were examined
Principles of method if other than guideline:
Groups of male and female rats were exposed to 0,05 and 0.15 ppm of toluene-diisocyanate (TDI) by inhalation for 6 h/day, 5 days/week for approx. 2 years. Type and incidence of tumours and the number of tumour-bearing animals were recorded. Additionally clinical signs, mortality, body weight gain, haematology, biochemistry, urinalysis, and cytogenicity were recorded. At termination of the study necropsy and histopathological examination were performed and organ weights recorded.
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): production-grade toluene diisocyanate (TDI)
- 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
- Analytical purity: approx. 80% of the 2,4- and 20% of the 2,6-isomer
- Composition of test material, percentage of components: approx. 80% of the 2,4- and 20% of the 2,6-isomer
- Isomers composition: approx. 80% of the 2,4- and 20% of the 2,6-isomer

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, UK
- Age at study initiation: 6-9-week-old, randomly allocated to exposure and control groups

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: TDI-vapour was generated by the passage of dry nitrogen through liquid TDI maintained at 21 °C, The vapour produced was then diluted with air in all-glass systems to produce the exposure concentrations.
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless steel/glass cubical chambers with pyramidal ceilings of approx. 10 m³ were used.
- System of generating particulates/aerosols: TDI-vapour was generated by the passage of dry nitrogen through liquid TDI maintained at 21 °C, The vapour produced was then diluted with air in all-glass systems to produce the exposure concentrations.
- Temperature, humidity, pressure in air chamber: Temperature and humidity in the chambers were recorded daily.

TEST ATMOSPHERE
- Brief description of analytical method used: exposure levels have been monitored using both the U.E.I. Model 7000 TDI Tape monitor and a colorimetric method.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The exposure levels have been monitored using both the U.E.I. Model 7000 TDI Tape monitor and a colorimetric method, The actual levels were: 0.052 ± 0.007 ppm, 0.146 ± 0.014 ppm.
Duration of treatment / exposure:
108 (females) or 110 weeks (males) = approximately 2 years
Frequency of treatment:
6 h/day, 5 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0.0 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0.05 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0.15 ppm
Basis:
nominal conc.
No. of animals per sex per dose:
21 rats per sex per level = in total 126 (including animals for interim kill)
Control animals:
yes, concurrent no treatment
Details on study design:
- Dose selection rationale:
Range-finding study
A range-finding study was carried out at another laboratory, and has been reported [2] prior to starting the lifetime studies. Sprague-Dawley rats (Spartan), Fisher 344 rats (Charles River), CD-I mice (Charles River), and Syrian hamsters (Engle Labs.) in groups of 10 animals of each sex and strain were exposed to TDI-vapour for 6 h/day, 5 days/week for a total of 20-22 exposures at levels of 0 (control), 0.1 or 0.3 ppm. Exposure was conducted under dynamic airflow conditions in chambers with quadrangular, pyramidal ceilings. TDI-vapour was generated by metering a calculated amount of liquid TDI into a heated vaporization flask at 220°C. The actual chamber concentrations of TDI were monitored analytically using a modified colorimetric method based on Marcali [1],
Clinical observations and body weights were recorded; haematological (RBC, haemoglobin, haematocrit, total and differential WBC counts) and biochemical (BUN, SGPT, AP) parameters were measured at the end of the study for animals of each sex, species and strain.

- Rationale for animal assignment (if not random): randomly allocated to exposure and control groups

- Other: An additional 5 rats per sex per level were exposed for 4 weeks to conduct the micronucleus test.
Positive control:
No data on positive controls available.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Clinical signs were recorded at regular intervals

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: clinical signs and ophthalmoscopic observations were recorded at regular intervals

BODY WEIGHT: Yes
- Time schedule for examinations: body weights were recorded at regular intervals

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: ophthalmoscopic observations were recorded at regular intervals
- Dose groups that were examined: 0 ppm, 0.05 ppm and 0.15 ppm

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Haematological (hemoglobin, RBC, PCV, total and differential, white cell count) were measured after 6, 12, 18 months of exposure and terminally.
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: blood biochemical parameters (urea nitrogen, alkaline phosphatase and alanine-aminotransferase) were measured after 6, 12, 18 months of exposure and terminally.
- Animals fasted: No data
- How many animals: all animals

URINALYSIS: Yes
- Time schedule for collection of urine: measured after 6, 12, 18 months of exposure and terminally
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Animals found dead, killed in extremis or scheduled for interim or terminal kills were subjected to gross and histopathology. Organ weights were recorded at all scheduled necropsies. The following tissues were examined histologically in both studies: adipose tissue, adrenal glands, aorta, blood film, brain (3 levels), caecum, colon, epididymides, eyes, femur (including bone marrow), gonads, heart, intestine, kidneys, liver, lungs (infused), lymph nodes, mammary gland, nasal turbinate (2 levels; mouse terminated, rat in progress), oesophagus, pancreas, pituitary, prostate, salivary glands, sciatic nerve, seminal vesicles, skeletal muscle (m. quadriceps), spinal cord (high cervical), spleen, stomach, thyroid, trachea, urinary bladder, uterus.
Other examinations:
Micronucleus test in rats
The results of the micronucleus test indicate that there was no dose- or treatment-related percentage increase of micronucleated erythrocytes in rats exposed to TDI at levels up to and including 0.15 ppm for 4 weeks.
Statistics:
No additional information on statistics available.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No exposure-related signs occurred during the study. At termination of the study total percentage deaths were 65% in controls, 67% low dose, 71% high dose males & 68% in controls, 75% low dose, 64% high dose females.
Mortality:
mortality observed, treatment-related
Description (incidence):
No exposure-related signs occurred during the study. At termination of the study total percentage deaths were 65% in controls, 67% low dose, 71% high dose males & 68% in controls, 75% low dose, 64% high dose females.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
in all groups during the main part of the study. Significantly less weight gain in the 0.15 ppm group in both sexes in the first 12 weeks of exposure. Weight increases between weeks 12 and 108 show comparable weight gains for all groups.
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
no effects observed
Description (incidence and severity):
No treatment-related changes in haematological parameters were recorded.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
No treatment-related changes in blood biochemical parameters were recorded.
Urinalysis findings:
no effects observed
Description (incidence and severity):
No treatment-related changes in urinary parameters were recorded.
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
The statistical analysis of the organ weights of the animals from interim and terminal kills did not reveal treatment-related differences.
Gross pathological findings:
no effects observed
Description (incidence and severity):
The examination of the tissues did not reveal any evidence of treatment-related effects. Macroscopically no changes were seen in the upper respiratory tract.
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
Microscopic evaluation of the tissues revealed some alterations in all groups which were considered incidental and not treatment-related. It should, however, be noted that histopathology of the nasal turbinates is still in progress.
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
Statistical evaluation demonstrated no treatment-related effect on the incidence of tumours, their multiplicity or malignancy.
Details on results:
CLINICAL SIGNS AND MORTALITY
No exposure-related signs occurred during the study. At termination of the study (at week 110 for males and at week 108 for females) total percentage deaths were 65% in controls, 67% low dose, 71% high dose males and 68% in controls, 75% low dose, 64% high dose females. Statistical analysis indicated that TDI did not significantly affect mortality.

BODY WEIGHT AND WEIGHT GAIN
Body weight gain was similar in all groups during the main part of the study. There was, however, significantly less weight gain in the 0.15 ppm group in both sexes in the first 12 weeks of exposure. The weight increases for the periods between weeks 12 and 108 show comparable weight gains for all groups.

OPHTHALMOSCOPIC EXAMINATION
No effects reported.

HAEMATOLOGY
No treatment-related changes in haematological parameters were recorded.

CLINICAL CHEMISTRY
No treatment-related changes in blood biochemical parameters were recorded.

URINALYSIS
No treatment-related changes in urinary parameters were recorded.

ORGAN WEIGHTS
The statistical analysis of the organ weights of the animals from interim and terminal kills did not reveal treatment-related differences.

GROSS PATHOLOGY
At necropsy the examination of the tissues from control and TDI-exposed animals did not reveal any evidence of treatment-related effects. Macroscopically no changes were seen in the upper respiratory tract.

HISTOPATHOLOGY: NON-NEOPLASTIC
Microscopic evaluation of the tissues revealed some alterations in all groups which were considered incidental and not treatment-related. It should, however, be noted that histopathology of the nasal turbinates is still in progress.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
Statistical evaluation demonstrated no treatment-related effect on the incidence of tumours, their multiplicity or malignancy.

Effect levels

Dose descriptor:
NOAEC
Effect level:
0.15 ppm
Sex:
male/female

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Range-finding study:

Male rats of both the Fisher 344 and Sprague-Dawley strain exposed to 0.1 or 0.3 ppm TDI showed a slight but significant reduction in body weight gain. A greater incidence of sneezing occurred in the Sprague-Dawley males suggesting slight respiratory irritation. No adverse effects were observed in the mouse and hamster study.

Rats did not exhibit changes in the lower respiratory tract (histopathology of upper respiratory tract in progress) nor was there any trend of increased mortality in any group. Rats exposed at 0.15 ppm had significantly less increase in weight, only in the first 12 weeks of the study. No specific target tissue has been identified. The tumour pattern in the rat study was similar in the control and in the treated groups and corresponded to historical data. The study demonstrated no evidence of any neoplastic response. These results are supported by tests on mutagenicity. There was no evidence of a mutagenic response in vivo as demonstrated by the results of a micronucleus test on rats and mice exposed for four weeks to TDI at exposure concentrations up to and including 0.15 ppm. As far as the incomplete rat study can be interpreted, levels which cause no primary irritative effect also do not result in any other effect of toxicological significance. As a result of these studies the current occupational exposure levels are not considered to represent a serious health hazard. The possible sensitization to TDI, however, needs special attention.

TABLE I
TDI: LONG-TERM INHALATION STUDY IN RATS
Synopsis of tumour incidence (malignant tumours in brackets).
  Atmospheric concentrations of TDI (ppm)
  0 0.05 0.15
Sex ¿ ¿ ¿ ¿ ¿ ¿
Number of animals examined 104 104 104 105 104 105
Skin/Adnexa/Glands  
carcinoma (basal/squamous etc.) (5) (3) (3) (2) (6) 0
adenoma 0 0 0 0 3 0
papilloma 9 1 5 3 3 0
Mammary gland  
benign tumour (fibroadenoma etc.) 4 24 4 27 2 22
multiple benign tumours 1 55 0 42 1 52
carcinoma + benign tumour (1) (9) 0 (9) 0 (14)
multiple carcinoma + benign tumour 0 (3) 0 (1) 0 0
mammary tumour (unconf.) 0 1 0 0 0 0
Subcutis/muscle/bone  
fibroma 29 1 22 1 35 4
fibrosarcoma (1) (2) (2) 0 (2) 0
lipoma 10 3 6 0 6 3
osteoma 0 0 0 0 1 0
osteosarcoma (1) 0 (1) 0 (1) (1)
lymphangioma 0 0 0 0 1 0
histiocytoma (1) 0 (4) 0 (1) 0
rhabdomyosarcoma 0 0 (1) 0 0 0
sarcoma (unclass.) 0 0 (1) 0 0 0
Haemopoietic/lymphoreticular  
malignant lymphoma (6) (1) 0 (1) (3) (3)
haemangioma 1 1 1 2 4 0
thymus: thymoma 0 (1) 0 1+ (1) 0 1
squamous carcinoma 0 (1) 0 0 0 0
Uterus  
polyp - 5 - 2 - 3
leiomyoma   0 - 1 - 0
sarcoma - 0 - 0 - (1)
Cervix  
stromal tumour - 2 - 2 - 0
Pancreas  
islet cell adenoma 1 1 2 0 3 2
Liver  
angiosarcoma 0 (3) 0 (1) 0 (3)
carcinoma 0 0 (1) 0 0 (1)
cholangiocarcinoma 0 0 (1) 0 0 0
Jejunum  
sarcoma 0 0 (1) 0 0 0
Stomach  
adenocarcinoma 0 0 0 (1) 0 0
Caecum  
schwannoma 0 0 0 0 1 0
leiomyosarcoma 0 0 0 0 0 (1)
Rectum  
polyp 0 0 0 1 0 0
Lungs  
adenoma 2 3 3 0 1 1
Body cavities, membranes, surfaces etc.  
lipoma 0 1 0 0 0 0
mesothelioma 0 0 (1) 0 1 0
liposarcoma 0 0 0 0 (1) 0
haemangioma 0 0 0 0 1 0
lymphangioma 0 0 1 0 0 0
Adrenals  
phaeochromocytoma 1 0 1 0 1 0
cortical adenoma 2 1 1 0 0 1
cortical carcinoma 0 0 0 0 (1) 0
Pituitary  
adenoma 53 64 32 62 38 67
Thyroid  
c-cell carcinoma 0 0 0 (1) 0 0
c-cell adenoma 11 7 3 1 7 4
follicular adenoma 1 0 0 0 0 1
Parathyroids  
adenoma 0 1 0 0 0 0
Brain  
meningioma 0 0 0 0 2 0
astrocytoma 1 0 1 0 0 0
oligodendroglioma 1 0 0 0 0 0
Kidneys  
carcinoma 0 0 0 0 0 (1)
liposarcoma (1) 0 (1) 0 0 0
mesenchymal tumour 1 0 0 0 0 0
lipomatous tumour 1 0 3 0 0 0
nephroblastoma 0 0 1 0 0 0
Testes  
Leydig cell adenoma 2 - 2 - 1 -
Epididymides  
mesothelioma 0 - 1 - 0 -
Prostate  
adenocarcinoma 0 - 0 - (1) -
Ovaries  
granulosa-theca cell - 2 - 0 - 1
Heart  
angiosarcoma 0 (1) 0 0 0 0
Eyes  
leiomyoma 0 0 0 1 1 1
Miscellanous  
squamous cell carcinoma of unknown origin (1) 0 0 0 0 0
Nasal turbinates No macroscopically diagnosed tissue masses. Histopathology in progress.

TABLE II
TDI: LONG-TERM INHALATION STUDY IN RATS
Summary of total tumour incidence
  Males Females
  0 ppm 0.05 ppm 0.15 ppm 0 ppm 0.05 ppm 0.15 ppm
Total animals 104 104 104 104 105 105
Tumour bearers 86 67 81 105 98 101
Animals with malignant tumours 9 3 10 3 4 5
Animals with benign tumours 69 53 64 84 80 76
Animals with both malignant and benign tumours 8 11 7 18 14 20

TABLE III
MICRONUCLEUS TEST ON RATS EXPOSED TO TDI-VAPOUR FOR 4 WEEKS, PERCENTAGE OF MICRONUCLEATED ERYTHROCYTES, MEANS OF 5 ANIMALS PER SEX PER LEVEL ± STANDARD DEVIATION, 1000 RBC PER ANIMAL EXAMINED
  Atmospheric concentration of TDI (ppm)
  0 0.05 0.15
Sex ¿ ¿ ¿ ¿ ¿ ¿
Rats 0.6 ± .9 0.5 ± .2 0.9 ± .4 0.8 ± .l 0.8 ± .2 0.8 ± .4

Applicant's summary and conclusion

Conclusions:
The study is considered to be reliable (reliability Klimisch 2). The validity criteria of the test system were fulfilled. The test material did induce slight signs of toxicity but no increased incidence of tumours. The test material was considered to be not carcinogenic after exposure via the inhalation route under the conditions of the test. The results of the study did not reveal a substantial carcinogenic potential of TDI, when administered via inhalation to rats.
Executive summary:

The carcinogenicity of the test material was investigated in rats by Loeser et al (1983). The test was conducted similar to OECD TG451. Groups of male and female rats (21 animals per sex per group) were exposed to 0,05 and 0.15 ppm of toluene-diisocyanate (TDI) by inhalation for 6 h/day, 5 days/week for approx. 2 years. Type and incidence of tumours and the number of tumour-bearing animals were recorded. Additionally clinical signs, mortality, body weight gain, haematology, biochemistry, urinalysis, and cytogenicity were recorded. At termination of the study necropsy and histopathological examination were performed and organ weights recorded. No exposure-related signs occurred during the study. At termination of the study (at week 110 for males and at week 108 for females) total percentage deaths were 65% in controls, 67% low dose, 71% high dose males and 68% in controls, 75% low dose, 64% high dose females. Male and female rats of the 0.15 ppm group gained less weight during the first 12 weeks of the study. Type and incidence of tumours and the number of tumour-bearing animals did not indicate any carcinogenic effect. Haematology, biochemistry, urinalysis, and cytogenicity did not reveal any untoward effect. Histopathological examination in the rat study has not been completed, but no effect in the respiratory tract or in any other tissue has yet been seen.