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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / 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
Cross-reference
Reason / purpose for cross-reference:
reference to same study
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: no data
- Age at study initiation: no data
- Weight at study initiation: 272.2-325.1 g for males, 188.1-235-9 g for females
- Housing:no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum):no data
- Acclimation period:no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-27
- Humidity (%): 35-75
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12-12
Route of administration:
oral: gavage
Vehicle:
corn oil
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
no details available
Duration of treatment / exposure:
42-53 days
Frequency of treatment:
Once daily
Remarks:
Doses / Concentrations:
10, 50, 200 mg/kg bw/day
Basis:

No. of animals per sex per dose:
12 males and 17 females per group at 0 and 200 mg/kg bw/day and 12 males and 12 females per group at 10 and 50 mg/kg bw/day; 12 males and 12 females per group were used for mating. To investigate reversibility, a recovery period of 14 days was established for 5 males and 5 non-mated females at 0 and 200 mg/kg bw/day
Control animals:
yes, concurrent vehicle
Details on study design:
Post-exposure period: 14 days
Positive control:
not relevant
Observations and examinations performed and frequency:
Clinical observations and frequency:
General condition was observed 2 or 3 times a day throughout the administration period and once a day throughout the recovery period.

Body weight:
Body weight was measured on days 1 (before dosing), 4, 8, 11, 15, 18, 22, 25, 30, 32, 36, 39, and 42 of the administration period in males. In females, body weight was measured on days 1 (before dosing), 4, 8, 11, 15, 18, 22, 25, 30, 32, 36, 39, and 42 of the administration period. Body weight of pregnant females was measured on days 0, 7, 14, and 20 of gestation and days 0 and 4 of lactation. Body weight was measured at necropsy in both sexes.

Food consumption:
Food consumption was measured on days 1 (before dosing), 4, 8, 11, 15, 30, 32, 36, 39, and 42 of the administration period in males. In females, food consumption was measured on days 1 (before dosing), 4, 8, 11, 15, 30, 32, 36, 39, and 42 of the administration period, and on days 18, 22, and 25 in non-mated females. Food consumption of pregnant females was measured on days 1, 7, 14, and 20 of gestation and days 1 and 4 of lactation.

During the recovery period, body weight and food consumption were measured on days 1, 4, 8, 11, and 14 in both sexes.

Urinalysis:
Urinalysis was carried out for 5 males per group at 6 weeks of the administration period.

Functional observation battery:
Detailed clinical observation was carried out once a week in all animals throughout the administration period. In pregnant females, it was carried out on days 7, 14, and 20 of gestation and on day 4 of lactation. Sensory reaction test, grip strength, and motor activity were examined at 6 weeks of administration in males and on day 4 of lactation in pregnant females.

Hematology and blood biochemistry:
At necropsy, hematology and blood biochemistry were examined in 5 males and 5 females per group.
Sacrifice and pathology:
Necropsy was carried out at the day following the end of the administration and recovery periods.

Organ weights:
Organs were examined at necropsy. The brain, heart, liver, kidney, adrenal, thymus, thyroids, and spleen of 5 males and 5 females per group and the testis and epididymis of all males were weighed.

Microscopic examination:
The spleen, liver, and kidney in 5 males and 5 females of each group were microscopically examined at the end of the administration and recovery periods. The cerebrum, cerebellum, medulla oblongata, pituitary, thymus, thyroid, parathyroid, adrenal, heart, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, trachea, lung, urinary bladder, spinal cord, mesenteric lymph node, submaxillary lymph node, sciatic nerve, femur (included bone marrow), and sternum (included bone marrow) of 5 males and 5 females at 0 and 200 mg/kg bw/day were microscopically examined at the end of the administration period. The testis, epididymis, prostate, and seminal vesicles of 5 males at 0 and 200 mg/kg bw/day were microscopically examined at the end of the administration period. Further, the ovary, uterus, and vagina of 5 females at 0 and 200 mg/kg bw/day were microscopically examined at the end of the administration period. Additionally, kidney tissue was immunohistochemically examined by staining with anti-alpha2u-globulin.
Statistics:
Statistical methods: For numerical data, Dunnett's test (homogeneous) or Steel test (heterogeneous) was used. Steel Multiple comparison test was used for urinary of test paper method. Wilcoxon's rank sum test was used for detailed clinical observation data, sensory reaction test data. Fisher exact probability test for necropsy data and Mann-Whitney U-test method for histopathology data were used.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
There was no death in any group.
No compound-related changes were observed in any animal.

BODY WEIGHT AND WEIGHT GAIN (see attached figure)
A low value was observed in both sexes at 200 mg/kg bw/day throughout the administration period. During the recovery period, a lower body weight was observed in females, but their body weight gains throughout the recovery period were similar to those of the control group.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
A low value or a tendency toward a low value was observed in males at 200 mg/kg bw/day on days 4 and 15 of administration and in females at 200 mg/kg bw/day throughout the administration period. During the recovery period, females exhibited lower food consumption on day 1 of the recovery period, but food consumption after day 4 of the recovery period was similar to the control group. A decrease in food consumption was observed in females at 10mg/kg on day 15 of the administration period. However, it was not observed at 50mg/kg and not considered to be a dose-related effect.

HAEMATOLOGY
Males:
Administration period (Table 1): Decreases in erythrocyte count, hemoglobin, hematocrit, and MCHC, an increase in platelet count, and prolonged PT and APTT were observed at 200 mg/kg bw/day. A decrease in MCHC was observed at 50 mg/kg bw/day. A decrease in reticulocyte at 10 mg/kg was not considered to be a dose related effect because it was not observed at 50 mg/kg bw/day and higher.
Recovery period (Table 2): Decreases in hemoglobin and MCHC and an increase in reticulocyte ratio were observed at 200 mg/kg bw/day.

Females:
Administration period Table 3): A decrease in erythrocyte count and an increase in reticulocyte ratio were observed at 200 mg/kg bw/day. An increase in reticulocyte at 50 mg/kg bw/day was not considered to be a dose related effect. A shortening of the APTT was observed at 50 mg/kg bw/day and higher.
Recovery period (Table 4): Increases in MCV and MCH and a decrease in MCHC were observed at 200 mg/kg bw/day. An increase in basophile at 200 mg/kg was not considered to be a dose related effect because it was not observed at the end of administration period.

CLINICAL CHEMISTRY
Males:
Administration period (Table 5): Decreases in alpha1-globulin, glucose and chlorine and increases in alpha2-globulin, albumin, gamma-GTP, total cholesterol, and phospholipids were observed at 200 mg/kg bw/day. Increases in total cholesterol and phospholipids at 50 mg/kg bw/day were observed.
Females:
Administration period (Table 6): Decreases in alpha1-globulin and glucose and increases in total protein, A/G ratio, albumin, total cholesterol, and phospholipids were observed at 200 mg/kg bw/day. An increase in total cholesterol was observed at 50 mg/kg bw/day. A decrease in ALP at 50 mg/kg bw/day was not considered to be a dose related effect because it was not observed at 100 mg/kg bw/day.
Recovery period: A decrease in creatinine was observed at 200mg/kg bw/day. However it was not considered to be a dose related effect because this effect was not observed at the end of administration period.

URINALYSIS
No test substance-related changes were observed.

NEUROBEHAVIOUR
No compound-related changes were observed in any animal in the functional observation battery.

ORGAN WEIGHTS
Administration period (Tables 7 and 8): Increases in absolute and relative weights of the liver were observed in males at 50 mg/kg bw/day and above and in females at 200 mg/kg bw/day. In the kidney of males, absolute weight at 50 mg/kg bw/day and above, and relative weight at 10 mg/kg bw/day and above were significantly increased. A decrease in absolute thymus weight was observed in males at 200 mg/kg bw/day. There were increases in relative weights of the heart, thyroids, testes and epididymides in males at 200 mg/kg and of the heart in females at 200mg/kg. However, absolute weights of these organs were not changed, and no histopathological changes were observed in these organs. These changes were considered to be due to decreases in body weights. Increases in absolute and relative weights of the epididymides in males and of the thymus in females were observed at 50mg/kg, but these changes were not considered to be dose related effects because these effects were not observed at 200 mg/kg bw/day.
Recovery period (Tables 9 and 10): Increase in relative liver weight was observed in both sexes at 200 mg/kg bw/day. Furthermore, increases in absolute and relative weights of the kidneys were observed in males at 200 mg/kg bw/day. There were increases in absolute weight of the adrenal in males at 200mg/kg and relative weight of the kidney in females at 200 mg/kg bw/day. However, these effects were not considered to be dose related effects because these changes were not observed at the end of administration period.

GROSS PATHOLOGY
Administration period: Enlargement and discoloration of the kidneys were observed in 1, 3, and 4 males at 10, 50, and 200 mg/kg bw/day, respectively. Liver enlargement was observed in 2 males at 200 mg/kg bw/day.
Recovery period: Kidney enlargement was observed in 1 male at 200 mg/kg bw/day.

HISTOPATHOLOGY: NON-NEOPLASTIC
Administration period (Tables 11 and 12): Histopathological changes were observed in the liver and spleen of both sexes and in the kidneys of males. The histopathological findings were as follows: slight centrilobular hypertrophy of hepatocytes in the liver in 5 males and 3 females at 50 mg/kg bw/day and slight or moderate hypertrophy in all males and females at 200 mg/kg bw/day, hemosiderin deposits in the red pulp in the spleen of both sexes at 200 mg/kg bw/day, periportal fatty degeneration of hepatocytes in the liver in males at 50 mg/kg bw/day and above, basophilic renal tubules in the kidneys in males at 10 mg/kg bw/day and above and hyaline deposits in proximal tubular epithelial cells in the kidneys in males at 10 mg/kg bw/day and above, which indicates alpha2µ-globulin nephropathy.
Recovery period (Tables 13 and 14): Hemosiderin deposits were observed in the red pulp of the spleen of both sexes, and basophilic renal tubules were observed in the kidneys in males at 200 mg/kg bw/day. Periportal fatty degeneration of hepatocytes in the liver was observed in 1 male and 1 female at 200 mg/kg bw/day (4 males and 0 female at the end of the treatment period). Centrilobular hypertrophy of hepatocytes in the liver is not longuer observed after recovery.
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Sex:
male/female
Basis for effect level:
other: slight reversible hematological and liver effects
Dose descriptor:
LOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Sex:
male/female
Basis for effect level:
other: decreased body weight gain and slight reversible hematological and liver effects
Critical effects observed:
not specified
Table 1: Haematological parameters in males (end of treatment)
Dose (mg/kg bw/day)
 
0
 
10
 
50
 
200
 
Erythrocyte count (10^4/uL)
 
815(29)
 
820(21)
 
768(39)
 
743(28)**
 
Hemoglobin(g/dL)
 
14.9(0.5)
 
14.9(0.6)
 
14.2(0.3)
 
13.7(0.5)**
 
Hematocrit (%)
 
42.2(1.2)
 
42.9(1.6)
 
40.9(0.9)
 
39.8(1.8)*
 
MCHC (g/dL)
 
35.3(0.2)
 
34.9(0.2)
 
34.7(0.5)*
 
34.5(0.3)**
 
Reticulocyte (%)
 
2.3(0.2)
 
1.8(0.3)*
 
 2.2(0.2)
 
3.5(0.9)
 
Platelet count (10^4/uL)
 
95.1(5.9)
 
96.1(5.9)
 
106.6(9.1)
 
121.7(19.2)**
 
PT (sec)
 
14.1(1.1)
 
14.8(1.8)
 
16.5(4.2)
 
21.9(8.9)*
 
APTT (sec)
 
26.1(1.9)
 
27.9(4.3)
 
27.0(3.8)
 
38.1(7.5)**
 
*p<0.05, ** p<0.01; n=5
Mean (Standard Deviation)
Table 2: Haematological parameters in males (end of recovery)
Dose (mg/kg bw/day)
 
0
 
200
 
Hemoglobin (g/dL)
 
15.5(0.5)
 
14.5(0.6)*
 
MCHC (g/dL)
 
35.8(0.8)
 
34.7(0.6)*
 
Reticulocyte (%)
 
2.3(0.2)
 
2.8(0.3)*
 
Note: *, p<0.05; n=5
Mean (Standard Deviation)

Table 3: Haematological parameters in females (end of treatment)
Dose (mg/kg bw/day)
 
0
 
10
 
50
 
200
 
Erythrocyte count (10^4/uL)
 
656(36)
 
634(12)
 
640(21)
 
607(37)*
 
Reticulocyte (%)
 
4.4(0.7)
 
7.0(1.8)*
 
5.7(2.1)
 
6.4(0.5)*
 
APTT (sec)
 
19.6 (1.6)
 
17.8(1.0)
 
17.1(0.8)*
 
16.8(1.4)**
 


    
Table 4: Haematological parameters in females (end of recovery)


    

      

        
Dose (mg/kg bw/day)

        
0

        
200

      

      

        
MCV (fL)

        
52.3(0.9)

        
54.8(1.2)**

      

      

        
MCH (pg)

        
18.9(0.4)

        
19.5(0.3 )*

      

      

        
MCHC (g/dL)

        
36.2(0.4)

        
35.6(0.3)*

      

      

        
Basophile (%)

        
0.1(0.0)

        
0.2(0.1)*

      

    

    
Note: *, p<0.05, **; p<0.01; n=5


    


    
Table 5: Blood biochemistry parameters in males (end of treatment)


    

      

        
Dose (mg/kg bw/day)

        
0

        
10

        
50

        
200

      

      

        
alpha1-globulin (%)

        
17.9(2.6)

        
17.8(1.6)

        
16.8(2.3)

        
13.7(1.4)*

      

      

        
alpha2-globulin (%)

        
7.6(0.7)

        
8.0(0.4)

        
8.2(0.6)

        
8.8(0.8)*

      

      

        
Albumin (g/dL)

        
2.92(0.22)

        
3.04(0.18)*

        
2.93(0.16)

        
3.43(0.30)*

      

      

        
gamma-GTP (IU/L)

        
0.3(0.2)

        
0.5(0.4)

        
0.2(0.3)

        
0.8(0.4)*

      

      

        
Total cholesterol (mg/dL)

        
60(10)

        
73(17)

        
85(8)*

        
96(15)**

      

      

        
Phospholipid (mg/dL)

        
106(10)

        
126(20)

        
141(8)*

        
171(26)**

      

      

        
Glucose (mg/dL)

        
124(9)

        
117(16)

        
111(23)

        
79(9)**

      

      

        
Chlorine (mEq/L)

        
108.4(2.3)

        
107.6(1.6)

        
107.0(1.4)

        
105.1(2.1)*

      

    

    

      Note: 
      *, p<0.05 **; p<0.01; n=5      

Mean (Standard Deviation)      


    


    


    
Table 6: Blood biochemistry parameters in females (end of treatment)


    

      

        
Dose (mg/kg bw/day)

        
0

        
10

        
50

        
200

      

      

        
Total protein (g/dL)

        
5.2(0.3)

        
5.4(0.2)

        
5.4(0.5)

        
6.2(0.4)**

      

      

        
A/G Ratio

        
1.19(0.07)

        
1.12(0.12)

        
1.20(0.09)

        
1.36(0.11)*

      

      

        
alpha1-globulin (%)

        
18.4(1.2)

        
17.6(1.8)

        
17.9(0.7)

        
15.5(2.1)*

      

      

        
Albumin (g/dL)

        
2.85(0.18)

        
2.83(0.21)

        
2.92(0.25)

        
3.57(0.29)**

      

      

        
ALP (IU/L)

        
241(37)

        
223(75)

        
132(30)*

        
177(90)

      

      

        
Total cholesterol (mg/dL)

        
74(3)

        
80(7)

        
97(8)*

        
120(22)*

      

      

        
Phospholipid (mg/dL)

        
144(8)

        
145(10)

        
173(19)

        
223(32)**

      

      

        
Glucose (mg/dL)

        
121(7)

        
112(12)

        
107(14)

        
93(8)**

      

    

    
Note: *, p<0.05 **; p<0.01; n = 5
Mean (Standard Deviation)


    

      

    
Conclusions:
t-butyl mercaptan induced an increase of kidney weights in males exposed to 50 and 200mg/kg bw/d and a chronic hyaline droplet nephropathy in all males, a rat-specific effect which is with no relevance for human risk assessment (Hard et al., 2009). Based on the decreased body weight gain in both males and females at 200 mg/kg bw/day, the slight reversible hematological changes and the slight or moderate reversible hepatocellular hypertrophy observed in males at 50 and 200 mg/kg bw/d and in females at 200 mg/kg bw/d, the LOAEL is considered to be 200 mg/kg bw/day and the NOAEL is considered to be 50 mg/kg bw/day.

References:
Hard GC, Johnson KJ and Cohen SM (2009) A comparison of rat chronic progressive nephropathy with human renal disease-implications for human risk assessment. CRIT-REV-TOXICOL, 39, 332-346.
Executive summary:

      In 
      a combined repeated-dose/reproductive/developmental toxicity screening 
      study (OECD TG 422), Crl:CD(SD) rats were administered 0, 10, 50 or 200 
      mg/kg bw/day of t-butyl mercaptan by gavage in corn oil daily for 42-53 
      days and a satellite group in the control and high dose were monitored 
      without dosing for 2-weeks for recovery. Group sizes were12 males and 17 
      females at 0 and 200 mg/kg bw/day and 12 males and 12 females at 10 and 
      50 mg/kg bw/day; 12 males and 12 females per group were used for mating 
      and a recovery period of 14 days was established for 5 males and 5 
      non-mated females at 0 and 200 mg/kg bw/day.  
    


    

      There 
      was no mortality, clinical signs of toxicity or changes in functional 
      observational battery measurements. Decreased body weight was observed 
      in both sexes at 200 mg/kg bw/day throughout the administration 
      period. During the recovery period, a lower body weight was observed in 
      females, but body weight gains throughout the recovery period were 
      similar to those of the control group. Decreased food consumption was 
      observed in males at 200 mg/kg bw/day on days 4 and 15 of administration 
      and in females at 200 mg/kg bw/day throughout the administration 
      period. During the recovery period, females exhibited lower food 
      consumption on day 1 of the recovery period, but food consumption after 
      day 4 of the recovery period was similar to the control group. There was 
      no effect on urinalysis measurements. For males, decreases in 
      erythrocyte count, hemoglobin, hematocrit, and MCHC, an increase in 
      platelet count, prolonged PT and APTT, decreased a-1-globulin, glucose 
      and chlorine, and increased a-2-globulin, albumin, g-GTP, total 
      cholesterol, and phospholipids were observed at 200 mg/kg bw/day.  
      Slight increases in total cholesterol and phospholipids and a decrease 
      in MCHC were observed at 50 mg/kg bw/day. During the recovery period, 
      decreases in hemoglobin and MCHC and an increase in reticulocyte ratio 
      were observed at 200 mg/kg bw/day. For females, decreased erythrocyte 
      count, increased reticulocyte ratio, decreases in a-1-globulin and 
      glucose and increases in total protein, A/G ratio, albumin, total 
      cholesterol, and phospholipids were observed at 200 mg/kg bw/day. A 
      shortening of the APTT was observed at 50 and 200 mg/kg bw/day, and an 
      increase in total cholesterol was observed at 50 mg/kg bw/day. During 
      the recovery period, increases in MCV and MCH, a decrease in MCHC and a 
      decrease in creatinine were observed at 200 mg/kg bw/day. At necropsy, 
      enlargement and discoloration of the kidneys were observed in 1, 3, and 
      4 males at 10, 50, and 200 mg/kg bw/day, respectively and liver 
      enlargement was observed in 2 males at 200 mg/kg bw/day; after the 
      recovery period, kidney enlargement was observed in 1 male at 200 mg/kg 
      bw/day. No gross findings were recorded for females. Increases in 
      absolute and relative liver weights were observed in males (50 and 200 
      mg/kg bw/day) and females (200 mg/kg bw/day). Kidney weight in males at 
      50 and 200 mg/kg bw/day and relative weight at all doses were 
      significantly increased. A decrease in absolute thymus weight was 
      observed in males at 200 mg/kg bw/day. Following the recovery period, 
      increased relative liver weight was observed in both sexes and increases 
      in absolute and relative weights of the kidneys were observed in males 
      at 200 mg/kg bw/day. Histopathological changes were observed in the 
      liver and spleen of both sexes and in the kidneys of males 
      including: slight and/or moderate hepatocellular centrilobular 
      hypertrophy in males at 50 and 200 mg/kg bw/day and in females at 200 
      mg/kg bw/day; hemosiderin deposits in the red pulp in the spleen of both 
      sexes at 200 mg/kg bw/day; periportal fatty degeneration of hepatocytes 
      in males at 50 and 200 mg/kg bw/day; basophilic renal tubules and 
      hyaline deposits in proximal tubular epithelial cells in the kidneys in 
      males at all doses which were considered to be indicative of 
      a-2µ-globulin nephropathy (a rat-specific effect which is with no 
      relevance for human risk assessment). Following the recovery period, 
      hemosiderin deposits were observed in the red pulp of the spleen of both 
      sexes, basophilic renal tubules were observed in the kidneys in males, 
      and periportal fatty degeneration of hepatocytes was observed in 1 male 
      and 1 female at 200 mg/kg bw/day.  Based on the decreased 
      body weight gain in both males and females at 200 mg/kg bw/day, the 
      slight reversible hematological changes and the slight or moderate 
      reversible hepatocellular hypertrophy observed in males at 50 and 200 
      mg/kg bw/d and in females at 200 mg/kg bw/d, the LOAEL is considered to 
      be 200 mg/kg bw/day and the NOAEL is considered to be 50 mg/kg bw/day.
    
Data source
Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report date:
2006
Materials and methods
Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
The histopathological examination of the reproductive organs was only performed on 5 animals/sex of the control and top dose.
GLP compliance:
yes
Limit test:
no
Test material
Constituent 1
Chemical structure
Reference substance name:
2-methylpropane-2-thiol
EC Number:
200-890-2
EC Name:
2-methylpropane-2-thiol
Cas Number:
75-66-1
Molecular formula:
C4H10S 
IUPAC Name:
2-methylpropane-2-thiol
Test animals
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: no data
- Age at study initiation: no data
- Weight at study initiation: 272.2-325.1 g for males, 188.1-235-9 g for females
- Housing:no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum):no data
- Acclimation period:no data
					
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-27
- Humidity (%): 35-75
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12-12
Administration / exposure
Route of administration:
oral: gavage
Vehicle:
corn oil
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
no details available
Duration of treatment / exposure:
Exposure period: Males: 42 days; Females: 42-53 days from 14 days before mating to day 4  of lactation

Premating exposure period (males): 2 weeks

Premating exposure period (females): 2 weeks

Duration of test: 10, 50, 200 mg/kg bw/day
Frequency of treatment:
Once daily
Doses / concentrations
Remarks:
Doses / Concentrations:
10, 50, 200 mg/kg bw/day
Basis:

No. of animals per sex per dose:
Males: 12
Females: 17 for control and top dose, 12 for low and mid doses
Control animals:
yes, concurrent vehicle
Examinations
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS:  Yes 
General condition was observed 2 or 3  times a day throughout the administration period
					
DETAILED CLINICAL OBSERVATIONS:  Yes
Detailed clinical observation was carried out once a week in all animals throughout the administration period. In pregnant females, it was carried out on days 7, 14, and 20 of gestation and on day 4 of lactation. Sensory reaction test, grip strength, and motor activity were examined at 6 weeks  of administration in males and on day 4 of lactation in pregnant females.
					
BODY WEIGHT: Yes 
Body weights were measured on days 1 (before dosing), 4, 8, 11, 15, 18, 22, 25, 30, 32, 36,  39, and 42 of administration for males, For females, body weight was  measured on days 1 (before dosing), 4, 8, 11, 15, 18, 22, 25, 30, 32, 36,  39, and 42 of administration, except for pregnant females for whom it was  measured on days 0, 7, 14, and 20 of gestation and days 0 and 4 of  lactation. Further, it was measured at necropsy in both sexes. 
					
FOOD CONSUMPTION : 
Food consumption was measured on days 1 (before dosing), 4, 8, 11, 15,  30, 32, 36, 39, and 42 of administration for males. For females, food  consumption was measured on days 1 (before dosing), 4, 8, 11, 15, 30, 32,  36, 39, and 42 of administration, except for pregnant females for whom it  was measured on days 1, 7, 14, and 20 of gestation and days 1 and 4 of  lactation
					
WATER CONSUMPTION :  No
Oestrous cyclicity (parental animals):
yes
Sperm parameters (parental animals):
No
Litter observations:
STANDARDISATION OF LITTERS
Performed on day 4 postpartum: no
					
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain
					
GROSS EXAMINATION OF DEAD PUPS:
 yes, for external and internal abnormalities; possible cause of death was not determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
Necropsy was carried out at the day following the end of the administration and recovery periods
					
GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
- The testis and epididymis of all males were weighed. 
				
HISTOPATHOLOGY / ORGAN WEIGHTS
The  testis, epididymis, prostate, and seminal vesicles of 5 males at 0 and  200 mg/kg bw/day were microscopically examined at the end of the  administration period.  Further, the ovary, uterus, and vagina of 5  females at 0 and 200 mg/kg bw/day were microscopically examined at the  end of the administration period.  
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring  were sacrificed at 5 days of age.  
- These animals were subjected to postmortem macroscopic examination for gross abnormalities.
Statistics:
Statistical methods: X2 test was used for mating index, males and females  fertility indices, gestation index, and delivery index were used.  Wilcoxon Rank Sum Test Method for implantation index, death birth index,  live birth index, and viability index on day 4 were used.
Reproductive indices:
Estrous cycle, number of copulated, number of pregnant females, mating length, mating index (# of  pairs with successful copulation/# of pairs mating × 100), males or  females fertility indices (# of pregnant animals/#of animals with  successful mating×100), number of females with live pups, gestational  length, number of corpora lutea, number of implantations, number of pups  delivered, number of live pups delivered, gestation index (#  of females with live pups/# of pregnant females × 100), implantation  index (# of implants/# of corpora lutea × 100), delivery index (# of pups  born/# of implants × 100), death birth index (number of stillborns/number  of litter × 100), were determined.
Offspring viability indices:
Sex ratio, live birth index (# of live pups born/# of pups born ×  100), and viability index on day 4 (# of live pups on postnatal day (PND)  4 /# of live pups born × 100) were determined.
Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
Clinical signs:
no effects observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not examined
Reproductive function / performance (P0)
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Details on results (P0)
BODY WEIGHT AND FOOD CONSUMPTION  (PARENTAL ANIMALS)
A low  body weight value was observed in both sexes at 200 mg/kg bw/day throughout the administration period. During the recovery period, a lower body weight was observed in females, but their body weight gains throughout the recovery period were similar to those of the control group.

A low food consumption value or a tendency toward a low  value was observed in males at 200 mg/kg bw/day on days 4 and 15 of administration and in females at 200 mg/kg bw/day throughout the administration period. During the recovery period, females exhibited lower food consumption on day 1 of the recovery period, but food consumption after day 4 of the recovery period was similar to the control group. A decrease in food consumption was observed in females at 10mg/kg on day 15 of the administration period. However, it was not observed at 50mg/kg and not considered to be a dose-related effect.
					
ORGAN WEIGHTS (PARENTAL ANIMALS)
There were increases in relative weights of the testes and epididymides in males at 200 mg/kg. However, absolute weights of these organs were not changed, and no histopathological changes were observed in these organs. These changes were considered to be due to decreases in body weights.
Increases in absolute and relative weights of the epididymides were observed at 50mg/kg, but these changes were not considered to be dose related effects because these effects were not observed at 200 mg/kg bw/day.
Effect levels (P0)
open allclose all
Dose descriptor:
NOAEL
Remarks:
parental toxicity
Effect level:
50  mg/kg bw/day (actual dose received)
Sex:
male/female
Basis for effect level:
other: decreased body weight gain at 200 mg/kg/day
Dose descriptor:
NOAEL
Remarks:
reproductive performance and developmental toxicity
Effect level:
>= 200  mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: no effect
Results: F1 generation
General toxicity (F1)
Clinical signs:
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Histopathological findings:
not examined
Details on results (F1)
BODY WEIGHT (OFFSPRING)
Decreases in body weight of live pups on PND 4 were observed in both sexes at 200 mg/kg bw/day.
Effect levels (F1)
Dose descriptor:
NOAEL
Remarks:
neonatal toxicity
Generation:
F1
Effect level:
50  mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: Decreased pup body weights
Overall reproductive toxicity
Reproductive effects observed:
not specified
Any other information on results incl. tables
Reproductive performance of rats


    

      

        
Dose (mg/kg bw/day)

        
0

        
10

        
50

        
200

      

      

        
Number of females eximaned

        
12

        
12

        
12

        
12

      

      

        
Count of estrus

        
3.67(0.78)

        
3.92(0.29)

        
3.83(0.58)

        
3.83(0.39)

      

      

        
Estrus cycle

        
3.97(0.10)

        
4.00(0.00)

        
4.03(0.10)

        
4.00(0.00)

      

      

        
No. of pairs mating

        
12

        
12

        
12

        
12

      

      

        
No. of pairs with successful mating

        
12

        
12

        
12

        
12

      

      

        
No. of pregnant females

        
12

        
12

        
12

        
11

      

      

        
Duration of mating

        
2.75(1.42)

        
3.33(2.84)

        
3.58(3.18)

        
3.36(1.80)

      

      

        
Fertility index (%)

        
100

        
100

        
100

        
91.67

      

    

    

      
    


    

      Terminal delivery of F0 dams
    


    

      

        
Dose (mg/kg bw/day)

        
0

        
10

        
50

        
200

      

      

        
No. of females with live pups

        
12

        
12

        
12

        
11

      

      

        
Gestational length (day)

        
22.08(0.29)

        
22.50(0.52)

        
22.33(0.65)

        
22.18(0.40)

      

      

        
# of corpora lutea

        
15.08(2.11)

        
14.58(2.61)

        
15.67(2.15)

        
15.36(1.96)

      

      

        
# of implantation sites

        
14.25(1.76)

        
13.42(3.00)

        
14.92(2.75)

        
14.45(3.00)

      

      

        
# of pups delivered

        
13.75(1.82)

        
13.08(3.34)

        
14.08(3.20)

        
13.64(2.77)

      

      

        
Sex ratio (male/female)

        
0.83

        
1.20

        
0.92

        
0.88

      

      

        
# of live pups on day 4

        
13.75(1.82)

        
13.00(3.28)

        
13.75(3.11)

        
13.64(2.77)

      

      

        
Viability index on day 4

        
98.79

        
98.08

        
 96.97

        
99.33

      

      

        
Body weight of live newborns (g)

        

        

        

        

      

      

        
    Male Day 0

        
6.4(0.5)

        
6.9(0.6)

        
6.7(0.7)

        
6.2(0.2)

      

      

        
    Male Day 4

        
10.3(0.8)

        
10.3(2.2)

        
10.0(2.4)

        
8.6(0.9)**

      

      

        
    Female Day 0

        
6.1(0.6)

        
6.4(0.6)

        
6.3(0.6)

        
5.9(0.2)

      

      

        
    Female Day 4

        
9.8(0.9)

        
9.8(2.0)

        
 9.5(2.4)

        
8.2(0.9)**

      

      

        
Number with external anomalies

        
0

        
0

        
0

        
1
(exencephaly
open eyelid
protruding tongue)

      

    

    

      **: 
      P<0.01      

      


    
Applicant's summary and conclusion
Conclusions:
The test substance had no effects on any reproductive or developmental  parameter. The NOAEL for reproductive and developmental toxicity was considered to be >= 200 mg/kg bw/day and the NOAEL for parental and neonatal toxicity is considered to be 50 mg/kg/day.
Executive summary:

      The OECD Test Guideline 422 study 
      conducted for t-butyl mercaptan is described in Section 3.1.5. In this 
      study, groups of male and female Sprague-Dawley rats (12-17/sex/dose) 
      were administered 0, 10, 50 or 200 mg/kg bw/day of t-butyl mercaptan by 
      gavage in corn oil daily for 42-53 days. The animals were dosed daily 
      for 2 weeks prior to mating, during mating and gestation, and the 
      females were dosed for 4 days post-partum after which the adult females 
      and their pups were terminated. There were no treatment-related effects 
      at any dose on reproductive and developmental parameters including 
      mating index, fertility index, duration of gestation, gestation index, 
      total number of pups born, live birth index, number of pups alive and 
      viability index on day 4 of lactation or sex ratio. Decreases in body 
      weight of live pups on PND 4 were observed in both sexes at 200 mg/kg 
      bw/day. All reproductive organs from adult animals were normal grossly 
      and microscopically. The NOAEL for reproductive performance and 
      developmental toxicity was 200 mg/kg bw/day and the NOAEL for parental 
      and neonatal toxicity was considered to be 50 mg/kg bw/day.