1,1-Bis(tert-butyldioxy)-3,3,5-trimethylcyclohexane was administered daily by oral gavage to male and female rats at doses of 0, 4, 20 and 100 mg/kg for 52 weeks and necropsied at the end of the weeks 13 and 52 for examination of toxicity.

At the end of the 13-week administration period, treatment-related changes were observed on blood biochemical examination, organ weight measurement and histopathological examination.

On blood biochemical examination, high values for the b-globulin ratio and total cholesterol and low values for the albumin ratio, A/G ratio and/or chloride were observed in both sexes receiving 100 mg/kg. Elevated total cholesterol was also noted in females of the 20 mg/kg group.

On organ weight measurement, high values for absolute and relative liver weights were observed in both sexes of the 100 mg/kg group, along with elevated relative kidney weights in females. High relative liver weights were also observed in females of the 20 mg/kg group.

Histopathological examination revealed treatment-related changes in the liver and mesenteric lymph nodes. In the liver, mild or moderate hypertrophy of centrilobular hepatocytes was evident in females of the 20 mg/kg group and both sexes of the 100 mg/kg group. In the mesenteric lymph nodes, mild accumulation of foam cells was observed in both sexes receiving 100 mg/kg.

At the end of the 52-week administration period, treatment-related changes were observed on hematological examination, blood biochemical examination, necropsy, organ weight measurement and histopathological examination.

On hematological examination, males of the 100 mg/kg group showed high values for leukocytes, neutrophils, lymphocytes, basophils, monocytes and large unstained cells in differential leukocyte counts, low values for mean corpuscular volume and mean corpuscular hemoglobin, and prolongation of prothrombin time and activated partial thromboplastin time. Low values for hemoglobin and/or hematocrit were observed in females of the 20 and 100 mg/kg groups.

On blood chemical examination, low values for the albumin ratio, A/G ratio and sodium, and high values for total protein, a1-globulin ratio, a2-globulin ratio, b-globulin ratio, alanine aminotransaminase, total cholesterol and/or phospholipids were observed in both sexes of the 100 mg/kg group.

On organ weight measurement, elevated relative liver weights were apparent in females of the 4 mg/kg group and high values for absolute and relative liver weights in females of the 20 mg/kg group and both sexes receiving 100 mg/kg group. In addition, high values for absolute and relative thyroid gland weights in males and for absolute kidney weights were observed in females of the 100 mg/kg group.

Histopathological examination revealed treatment-related changes in the liver, mesenteric lymph nodes, spleen, duodenum, jejunum and ileum. In the liver, mild or moderate hypertrophy was observed in males of the 20 mg/kg group and both sexes given 100 mg/kg. Mild or moderate fatty degeneration of periportal hepatocytes was observed in both sexes of all groups, but was only significantly different from the control level in 100 mg/kg females. Mild or moderate hyperplasia of the bile duct was observed in both sexes of the control, 20 and 100 mg/kg groups, again with significant differences from the controls limited to males given 100 mg/kg. In addition, mild or moderate accumulation of foam cells in the sinusoids (with lymphocytes) was observed in both sexes of the 20 and 100 mg/kg groups. In the kidneys, mild or moderate basophilia of tubules was observed in males of all groups, and the change was only significantly different from controls in the females of the 100 mg/kg group. In the mesenteric lymph nodes, mild, moderate or marked accumulation of foam cells was observed in both sexes given 20 or 100 mg/kg. In the spleen, mild or moderate accumulation of foam cells in the white pulp was observed in males of the 20 mg/kg and both sexes of the 100 mg/kg group. In the duodenum, mild accumulation of foam cells in the lamina propria was observed in both sexes receiving 100 mg/kg. In the jejunum, mild accumulation of foam cells in the lamina propria was similarly observed in both sexes given 100 mg/kg, along with mild accumulation of foam cells in Peyer's patches. In the ileum, mild accumulation of foam cells in the lamina propria was again observed in both sexes at 100 mg/kg.

From the above-described results, the NOAELs are considered to be 4 mg/kg/day for males and less than 4 mg/kg/day for females under the conditions of this study.

SUMMARIZED DATA FROM THE STUDY

1. Repeated Dose Oral Toxicity 1)

Purity	:	98.8 %
Test species/strain	:	Rat/Crj:CD(SD)IGS
Test method	:	Chronic Oral Toxicity Study
Route	:	Oral(gavage)
Dosage	:	0(vehicle), 4, 20, 100 mg/kg/day
Number of animals/group	:	Males, 10; females, 10
Vehicle	:	Corn oil
Administration period	:	Males and females, 52 weeks
Terminal killing	:	Males and females, 13 or 52 weeks
GLP	:	Yes

　Test Results:

Clinical observation revealed no test substance-related changes in any groups. Deaths occurred of one male each in the 4 and 100 mg/kg groups and of 2 males and 1 female of the 20 mg/kg group, and 1 male of the 4 mg/kg group became moribund, but these were not considered related to administration of the test substance.

No abnormalities were observed in the body weights or food consumption during the administration period.

<At the end of the 13-week administration period>

No changes related to administration of the test substance were observed on urinalysis, hematological examination and necropsy.

Blood biochemical examination revealed the following changes; elevation of the b-globulin ratio and a lowering of chloride in males of the 100 mg/kg group, high b-globulin ratios and total cholesterol and low values for the albumin ratio and the A/G ratio in females of the 100 mg/kg group, and high values for total cholesterol in females of the 20 mg/kg group.

Organ weight measurement revealed the following changes; high values for absolute and relative liver weights in both sexes of the 100 mg/kg group, a high value for relative kidney weights in females of the 100 mg/kg group, and a high value for relative liver weights in females of the 20 mg/kg group.

Histopathological examination revealed treatment-related changes in the liver and mesenteric lymph nodes. In the liver, mild or moderate hypertrophy of centrilobular hepatocytes was noted in 2 females of the 20 mg/kg group and 2 males and all females of the 100 mg/kg group. In the mesenteric lymph nodes, mild accumulation of foam cells was observed in 2 males and 1 female of the 100 mg/kg group.

<At the end of the 52-week administration period>

No changes related to administration of the test substance were observed on urinalysis.

Hematological examination revealed the following changes; high values for leukocytes and neutrophils, lymphocytes, basophils, monocytes and large unstained cells in differential leukocyte counts, low values for mean corpuscular volume and mean corpuscular hemoglobin and prolongation of prothrombin time and activated partial thromboplastin time in males of the 100 mg/kg group, low values for hemoglobin and hematocrit in females of the 100 mg/kg group, and low values for hemoglobin in females of the 20 mg/kg group.

Blood chemical examination revealed the following changes; low values for the albumin and A/G ratios and high values for the a2-globulin and b-globulin ratios and alanine aminotransaminase in males of the 100 mg/kg group, and low values for the albumin and A/G ratios and sodium and high values for total protein, the a1-globulin and b-globulin ratios, total cholesterol and phospholipids in females of the 100 mg/kg group.

Organ weight measurement revealed the following changes; high values for absolute and relative liver weights in both sexes of the 100 mg/kg group, high values for absolute and relative thyroid gland weights in males of the 100 mg/kg group, high values for absolute kidney weights in females of the 100 mg/kg group, high values for absolute and relative liver weights in females of the 20 mg/kg group, and high values for relative liver weights in females of the 4 mg/kg group.

Histopathological examination revealed treated-related changes in the liver, mesenteric lymph nodes, spleen, duodenum, jejunum and ileum. In the liver, mild or moderate hepatocyte hypertrophy was observed in 1 male of the 20 mg/kg group and 4 males and 3 females of the 100 mg/kg group. Mild or moderate fatty degeneration of periportal hepatocytes was observed in 6 males and 3 females of the control group, single rats of both sexes of the 4 mg/kg group, 4 males and 2 females of the 20 mg/kg group and 5 males and 7 females of the 100 mg/kg group. The only significant difference from controls was for females of the 100 mg/kg group. Mild or moderate hyperplasia of the bile ducts was observed in single rats of both sexes of the control group, 2 of each sex of the 20 mg/kg group, and 7 males and 1 female in the 100 mg/kg group. This change was only significantly different from the control group in males receiving 100 mg/kg. In addition, mild or moderate accumulation of foam cells in the sinusoids (with lymphocytes) was observed in 2 males and 1 female of the 20 mg/kg group and all males and 9 females of the 100 mg/kg group. In the kidneys, mild or moderate basophilia of tubules was observed in 5 males of the control group, 5 males and 3 females of the 4 and 20 mg/kg groups and 9 males and 5 females of the 100 mg/kg group, the change only being significantly different from the control level in the females receiving 100 mg/kg. In the mesenteric lymph nodes, mild, moderate or marked accumulation of foam cells was observed in 5 males and 3 females of the 20 mg/kg group and all rats of both sexes given 100 mg/kg. In the spleen, mild or moderate accumulation of foam cells in the white pulp was observed in 1 male of the 20 mg/kg and 4 rats of both sexes of the 100 mg/kg group. In the duodenum, mild accumulation of foam cells in the lamina propria was observed in 1 rat of each sex of the 100 mg/kg group. In the jejunum, mild accumulation of foam cells in the lamina propria was observed in 8 males and 6 females of the 100 mg/kg group, together with mild accumulation of foam cells in the Peyer's patches in 3 males and 1 female. In the ileum, mild accumulation of foam cells in the lamina propria was observed in 6 males and 1 female of the 100 mg/kg group.

From the above-described results, the NOAELs are considered to be 4 mg/kg/day for males and less than 4 mg/kg/day for females under the conditions of this study.