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Repeated dose toxicity: inhalation

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Administrative data

sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1996-08-19 to 1996-11-19
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions

Data source

Reference Type:
study report

Materials and methods

Principles of method if other than guideline:
Method: other: NTP Test Protocol, see Test Conditions
GLP compliance:
Limit test:

Test material

Details on test material:
Tetralin was obtained from Sigma Aldrich Fluka Bulk Chemicals (St. Louis, MO) in two lots (00822JG and 07808LG) were used as a mixture.
The purity of each lot was determined to be greater than 97%.
Stability was monitored by the study laboratory during the 2‑week, 3‑month, and 2‑year studies using GC/FID. No degradation of the chemical occurred.

Test animals

Fischer 344
Details on test animals and environmental conditions:
- Source: Taconic, Germantown (New York, USA)
- Age: approximately 6 weeks at first exposure
- Number of animals: Total of 25 males and 20 females per dose = 5 male  renal toxicity rats + 10 male and 10 female core study rats
 + 10 male and  10 female clinical pathology rats
Feed was available ad libitum except during exposure and urine collectionperiods;
water was available ad libitum.
Rats and mice were housed individually.

Administration / exposure

Route of administration:
Type of inhalation exposure:
whole body
other: air
Details on inhalation exposure:
- Type of exposure: whole-body inhalation
Analytical verification of doses or concentrations:
Details on analytical verification of doses or concentrations:
The tetralin concentrations in the exposure chambers were monitored by an online gas chromatograph. Samples were drawn from each exposure chamber approximately every 24 minutes during each 6‑hour exposure period.
Chamber concentration uniformity was maintained throughout the studies.
Duration of treatment / exposure:
14 weeks
Frequency of treatment:
6 h/day, 5 days/week; additionally: last sunday before terminal sacrifice
Doses / concentrations
Doses / Concentrations:
7.5; 15; 30; 60; 120 ppm = 41.2; 82.4; 165; 330; 660 mg/m³
analytical conc.
No. of animals per sex per dose:
Control animals:
yes, concurrent no treatment
Details on study design:
Post-exposure period: sacrifice on day after last exposure


Observations and examinations performed and frequency:
- Clinical signs: twice daily including weekends
- Mortality: twice daily including weekends
- Body weight: weekly (core study rats)
- "Observations": weekly
- Hematology: Sampling from 10 animals per dose and sex on days 3 and 23  (after exposures, clinical pathology rats) and at terminal sacrifice  
(core study rats). Evaluations included: red blood cell count, volume of  packed cells and spun hematocrit, hemoglobin, mean corpuscular volume,   mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration,  white blood cell count, differential count (absolute), absolute  
reticulocyte count, platelet count, Morphological assessment.
- Biochemistry: Blood urea nitrogen, sorbitol dehydrogenase, alanine  aminotransferase, total protein, albumin, alkaline phosphatase, total  bile acids,  creatine kinase, creatinine.
- Urinalysis: 16-hour collection during week 12 on all surviving core  study animals, with access to water but not food. Measurements included:  
volume, specific gravity, appearance (visual inspection), microscopic  examination of sediment from centrifuged sample, glucose, protein,  
N-acetyl-beta-glucosaminidase, creatinine (to be used to normalize other  values), alkaline phosphatase, aspartate aminotransferase, lactate  
dehydrogenase, gamma glutamyl transaminase

- Assessment of kidneys after 2 weeks (5 renal toxicity rats), at 6 weeks  (5 male clinical pathology rats), and at terminal  sacrifice (5 male core  study  rats): Histopathology and evaluation of cell proliferation  (positive control: cross section of small intestine) in left kidney,  measurement of 
a2u-globulin in right kidney
-  Sperm morphology & vaginal cytology (SMVCE): Vaginal cytology was  evaluated for 12 days during the last 2 weeks of the study in all  remaining 
females in the 0-, 30-, 60-, and 120-ppm groups. Epididymal  sperm concentration, spermatid heads/testis, and left caudal, epididymal  & 
testicular weights were evaluated in surviving males from the same  groups.
Sacrifice and pathology:
- Macroscopic: Complete necropsy; weights of liver, thymus, right kidney,  right testis, heart, lungs.
- Microscopic: Complete histopathology on all 0- and 120-ppm-rats  included the following tissues: adrenal glands, brain, clitoral glands,  
esophagus, femur (including bone marrow & joint surfaces), gross lesions,  tissue masses, regional lymph nodes, heart, aorta, large intestine  
(cecum, colon, rectum), small intestine (duodenum, jejunum, ileum),  kidneys (left only for males), larynx, liver, lungs, mainstem bronchi,  lymph 
nodes (mandibular, mesenteric, bronchial, mediastinal), mammary  glands & adjacent skin, nasal cavity & nasal turbinates (three sections),  ovaries,  pancreas, parathyroid glands, pituitary glands, preputial  glands, prostate, salivary glands, spleen, stomach (forestomach &  glandular), testes / 
epididymis / seminal vesicles, thymus, thyroid  glands, trachea, urinary bladder, uterus.   
Target tissues identified at 120 ppm were examined at lower  concentrations to no-effect level or lowest exposure concentrations.  
Gross lesions were examined in all groups.
Other examinations:
At the end of the studies, sperm samples were collected from core study male animals in the 0, 30, 60, and 120 ppm groups for sperm motility evaluations. The following parameters were evaluated: spermatid heads per testis and per gram testis, spermatid counts, and epididymal spermatozoal motility and concentration. The left cauda, left epididymis, and left testis were weighed. Vaginal samples were collected for up to 12 consecutive days prior to the end of the studies from core study females exposed to 0, 30, 60, or 120 ppm for vaginal cytology evaluations. The percentage of time spent in the various estrous cycle stages and estrous cycle length were evaluated.
STATISTICAL METHODS: A modified Dunnett's t-test (Xybion Path / Tox  System; Cedar Knolls, New Jersey) was used to compare the treated groups  
to the control group with respect to body and organ weights, and  organ:body weight ratios. Corresponding statistics for hematology, and  clinical 
chemistry were calculated using the Statistical Analysis System  (SAS Institute; Berkeley, California).

Results and discussion

Results of examinations

Clinical signs:
no effects observed
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
reduced body weight at 120 ppm males
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:
effects observed, treatment-related
Description (incidence and severity):
modest regenerative anemia was observed in both sexes,  primarily in groups exposed to 60 and 120 ppm
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
erum alanine aminotransferase decreased at 60 and 120 ppm (both sexes)
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
dark stained urine
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Olfactory necrosis and regeneration,
Histopathological findings: neoplastic:
no effects observed
Details on results:
- Mortality and time to death: no mortalities in any group
- Clinical signs: no clinical abnormalities in any group
- Body weight gain: lower by 6.1 % (males) and 5.7 % (females),  respectively, in highest dose groups
- Clinical chemistry: Minimal nephropathy was observed in males in the  higher exposure groups. Clinical chemistry data were consistent with  
nephropathy. serum alanine aminotransferase decreased at 60 and 120 ppm (both sexes)
- Haematology: A modest regenerative anemia was observed in both sexes,  primarily in groups exposed to 60 and 120 ppm.
- Urinalysis:    Dark-stained urine at 30, 60, and 120 ppm.    Urine aspartate aminotransferase values significantly higher in males  (ca. 2.5) and 
females (ca. 17 times control values) at the 120 ppm level.   Urine lactic dehydrogenase (LDH):creatinine ratio significantly, but  modestly 
increased  in the two highest dose levels, LDH activity increased  in 120 ppm females group.
- Organ weights:    
Kidney: Increased right kidney:body weight ratio in males (15, 60, and  120 ppm) and females (15 ppm and higher); mean absolute  right kidney  
weight slightly increased in all treated groups;   
Liver:body weight ratios increased in males ( 120 ppm) and  females (60 and 120 ppm); mean absolute liver weight slightly increased  
in all groups exposed;
- Gross pathology: no gross observations in any dose group
- Histopathology:   Olfactory necrosis and regeneration, confirming the irritation  potential of 1,2,3,4-tetrahydronaphthalene. 
The NOAEC for nasal lesions  was 15 ppm in males and in females.    
Hyaline droplet accumulation in kidneys of males increased slightly  with increased exposure; a NOAEC was not clear.   
Minimal nephropathy in males in the higher exposure groups
- Other: Concentrations of a2u-globulin generally increased with exposure  concentration and time on study in all groups of male F344/N rats

Effect levels

open allclose all
Dose descriptor:
Effect level:
82.4 mg/m³ air
Based on:
act. ingr.
Basis for effect level:
other: nasal lesions
Dose descriptor:
Effect level:
41.2 mg/m³ air
Based on:
act. ingr.
Basis for effect level:
other: hyaline droplet formation; not relevant for human safety
Dose descriptor:
Effect level:
41.2 mg/m³ air
Based on:
act. ingr.
Basis for effect level:
other: nasal lesions

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Incidences of selected nonneoplastic lesions in F344/N rats in the 3-month inhalation study of tetrahydronaphthalene



7.5 ppm

15 ppm

30 ppm

60 ppm

120 ppm

Kidney Male Accumulation Hyaline droplet

week 2


2/5 (1.0)

3/5 (1.0)

4/5 (1.8)

5/5 (1.8)

5/5 (1.6))

week 6

5/5 (1.4)

5/5 (1.2)

5/5 (1.6)

5/5 (2.0)

5/5 (2.4)

5/5 (2.4)

week 14

10/10 (1.0)

10/10 (1.1)

10/10 (2.0)

10/10 (2.0)

10/10 (2.0)

10/10 (2.0)

Nose Olfactory Epithelium necrosis





4*/10 (1.8)










*Significantly different (P<0.05) from the chamber control group by the Fisher exact test

**Significantly different (P<0.01) from the chamber control group by the Fisher exact test

aNumber of animals with lesions

bnumber of animals examined

cAverage severity of grade of lesions in affected animals -1= minimal, 2=mild, 3=moderate, 4 =marked

Organ Weights and Organ-Weight-to-Body-Weight Ratios for F344/N Rats in the 3-Month Inhalationstudy with tetrahydronaphthalenea



7.5 ppm

15 ppm

30 ppm

60 ppm

120 ppm


Necropsy body wt


294 ± 8


301 ± 7


299 ± 6


301 ± 8


289 ± 8


276 ± 5

Heart           Absolute

0.82 ± 0.03

0.84 ± 0.03

0.83 ± 0.01

0.84 ± 0.02

0.84 ± 0.03

0.77 ± 0.02


2.791 ± 0.034

2.776 ± 0.037

2.775 ± 0.025

2.795 ± 0.052

2.891 ± 0.041

2.800 ± 0.027

R. Kidney


0.86 ± 0.03

0.93 ± 0.03

0.94 ± 0.02

0.92 ± 0.03

0.92 ± 0.03

0.90 ± 0.02


2.930 ± 0.037

3.082 ± 0.070*

3.143 ± 0.043*

3.051 ± 0.032*

3.181 ± 0.044**

3.270 ± 0.032**



8.48 ± 0.31

8.88 ± 0.25

9.59 ± 0.31*

9.00 ± 0.32

8.61 ± 0.34

8.65 ± 0.25


28.80 ± 0.56

29.53 ± 0.36

31.99 ± 0.62**

29.86 ± 0.44

29.76 ± 0.49

31.26 ± 0.52**


Necropsy body wt

183± 4

190 ± 4

184 ± 3

180 ± 4

178 ± 4

173 ± 3

R. Kidney


0.60 ± 0.01

0.63 ± 0.02

0.64 ± 0.01

0.64 ± 0.01

0.64 ± 0.02*

0.66 ± 0.01**



3.275 ± 0.044

3.314 ± 0.060

3.492 ± 0.041**

3.543 ± 0.059**

3.622 ± 0.056**

3.827 ± 0.048**

*Significantly different (P<0.05) from the chamber control group by William’s or Dunnett’s test

**Significantly different (P<0.01) from the chamber control

a Organ weights (absolute weights) and body weights are given in grams; organ-weight-to-body-weight ratios (relative weights) are given as

mg organ weight/g body weight (mean ± standard error).


Applicant's summary and conclusion

A LOAEC of 7.5 ppm (41.2 mg/m³) was observed based on hyaline droplet formation. This was not considered relevant for humans because of the species specific mechanism of toxicity. A NOAEC for local effects of 7.5 ppm (41.2 mg/m³) was observed based on increased incidences of olfactory epithelium necrosis and of olfactory epithelium regeneration in 60 and 120 ppm rats.
Executive summary:

Groups of 10 male and 10 female rats were exposed to tetrahydronaphthalene at concentrations of 0, 7.5, 15, 30, 60, or 120 ppm, 6 hours plus T90 (12 minutes) per day, 5 days per week for 14 weeks. The same exposure concentrations were given to additional groups of 10 male and 10 female clinical pathology study rats for up to 6 weeks and five male renal toxicity rats for 2 weeks.

All rats survived to the end of the study. Mean body weights of 120 ppm male rats were significantly less than those of the chamber controls. During the first 4 weeks of exposure, dark-stained urine was observed in the catch pans of rats exposed to 30, 60, or 120 ppm. Tetrahydronaphthalene induced a minimal decrease in the erythron in both sexes that resulted in a hematopoietic response.

Increased urine aspartate aminotransferase and urine lactate dehydrogenase activities (males and females) and glucose/creatinine ratio (males), suggestive of renal injury were observed. The absolute kidney weights of 60 and 120 ppm females and the relative kidney weights of males and females exposed to 15 ppm or greater were significantly greater than those of the chamber controls. Concentrations of [alpha]2u‑globulin in the kidney of exposed male rats were generally greater than those of the chamber controls at all time points and greater at 6 and 14 weeks than at 2 weeks. There were significantly increased incidences of olfactory epithelium necrosis and of olfactory epithelium regeneration in rats exposed to 15 ppm (82.4 mg/m³) and higher.