https://ntp.niehs.nih.gov/go/tr578abs

Abstract for TR-578

Toxicology and Carcinogenesis Studies of Ginkgo Biloba Extract in F344/N Rats and B6C3F1/N Mice (Gavage Studies)

CASRN: 90045-36-6
Synonyms/Common Names: Ginkgo; ginkgo biloba; fossil tree; maidenhair tree; Japanese silver apricot; baiguo; bai guo ye; kew tree; yinhsing (yin-hsing); ginkgo biloba
Report Date: March 2013

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Abstract

Ginkgo biloba extract has been used primarily as a medicinal agent in the treatment or prevention of cardiovascular and cerebrovascular dysfunction. Ginkgo biloba extract was nominated for study by the National Cancer Institute because of its widespread use as an herbal supplement to promote mental function and the limited availability of toxicity and carcinogenicity data. Furthermore, one of the major ingredients in Ginkgo biloba extract, quercetin, is a known mutagen. The Ginkgo biloba extract used in the current studies was procured from a supplier known to provide material to United States companies and contained 31.2% flavonol glycosides, 15.4% terpene lactones (6.94% bilo-balide, 3.74% ginkgolide A, 1.62% ginkgolide B, 3.06% ginkgolide C), and 10.45 ppm ginkgolic acid. Male and female F344/N rats and B6C3F1/N mice were administered Ginkgo biloba extract in corn oil by gavage for 3 months or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Escherichia coli, and mouse peripheral blood erythrocytes.

Three-month study in rats

Groups of 10 male and 10 female rats were administered 0, 62.5, 125, 250, 500, or 1,000 mg Ginkgo biloba extract/kg body weight in corn oil by gavage, 5 days per week for 14 weeks. Additional groups of 10 male and 10 female rats (clinical pathology study) were administered the same doses, 5 days per week for 23 days. All rats survived to the end of the study. Mean body weights of all dosed groups were similar to those of the vehicle control groups. Liver weights of all dosed groups of males and females were significantly greater than those of the vehicle control groups.

The incidences of hepatocyte hypertrophy in all dosed groups of males and in 500 and 1,000 mg/kg females were significantly greater than those in the vehicle control groups; there was a dose-related increase in severity of this lesion in males. Hepatocyte fatty change occurred in all dosed males. The incidences of thyroid gland follicular cell hypertrophy were significantly increased in 500 and 1,000 mg/kg males and in 1,000 mg/kg females. The incidences of pigmentation in the olfactory epithelium of the nose were significantly increased in 500 and 1,000 mg/kg males and in females administered 125 mg/kg or greater.

Three-month study in mice

Groups of 10 male and 10 female mice were administered 0, 125, 250, 500, 1,000, or 2,000 mg Ginkgo biloba extract/kg body weight in corn oil by gavage, 5 days per week for 14 weeks. One female mouse in the 1,000 mg/kg group died of a dosing accident during week 11. Mean body weights of 2,000 mg/kg females were significantly less than those of the vehicle control group. Ruffled fur was observed in two 1,000 mg/kg males between weeks 7 and 8 and all 2,000 mg/kg males between weeks 5 and 9. Liver weights of 250 mg/kg or greater males and all dosed groups of females were significantly greater than those of the vehicle control groups. Kidney weights of 2,000 mg/kg males were significantly less than those of the vehicle control group. The Markov transition matrix analyses indicate female mice in the 2,000 mg/kg group had a significantly higher probability of extended estrus than did the vehicle control females.

The incidences of hepatocytic hypertrophy were significantly increased in males and females in the 250 mg/kg or greater groups. Significantly increased incidences of focal hepatocytic necrosis occurred in 1,000 and 2,000 mg/kg males. The incidences of hyaline droplet accumulation in the respiratory epithelium of the nose were significantly increased in 500 mg/kg males and 1,000 and 2,000 mg/kg females. In the olfactory epithelium of the nose, the incidences of hyaline droplet accumulation were significantly increased in the 125 (female only), 500, and 1,000 mg/kg groups. Incidences of atrophy of the olfactory epithelium were significantly increased in the 1,000 mg/kg groups. The incidences of pigment accumulation in macrophages in the olfactory epithelium were significantly increased in males in the 500 mg/kg or greater groups and in 1,000 and 2,000 mg/kg females.

Two-year study in rats

Groups of 50 male and 50 female rats were administered 0, 100, 300, or 1,000 mg Ginkgo biloba extract/kg body weight in corn oil by gavage, 5 days per week for 104 or 105 (females) weeks. Additional groups of 10 male and 10 female rats (special study) were administered the same doses, 5 days per week for 14 weeks. Survival of 1,000 mg/kg males was significantly less than that of the vehicle controls. At week 14, all dosed groups of males and 1,000 mg/kg females had increased levels of thyroid stimulating hormone compared to those of the vehicle control groups. There were no significant decreases in the levels of triiodothyronine or total thyroxine. Mean body weights of 300 mg/kg males and females were less (10% or more) than those of the vehicle controls after week 93, and those of 1,000 mg/kg males and females were less after week 89. Clinical findings included ruffled fur in seven, eight, and 10 males in the 100, 300, and 1,000 mg/kg groups, respectively, beginning at week 89; four vehicle control males also had ruffled fur.

Liver weights were significantly increased in all dosed groups of special study rats at 14 weeks. In the liver at 2 years, incidences of hepatocellular adenoma were slightly increased in 100 and 300 mg/kg males. Significantly increased incidences of nonneoplastic lesions at 2 years included hepatocyte hypertrophy and bile duct hyperplasia in all dosed groups of males and females, focal fatty change in all dosed groups of females, cystic degeneration in 100 and 1,000 mg/kg males, and oval cell hyperplasia and necrosis in 1,000 mg/kg males.

In the thyroid gland, incidences of follicular cell adenoma were slightly increased in 300 and 1,000 mg/kg males and 300 mg/kg females. Single incidences of follicular cell carcinoma occurred in the 300 and 1,000 mg/kg female groups. There were significantly increased incidences of follicular cell hypertrophy in all dosed groups of males and females and follicle hyperplasia in all dosed groups of males.

In the nose, adenoma of the respiratory epithelium occurred in two females receiving 300 mg/kg. Except for respiratory epithelium hyperplasia in 100 mg/kg females, the incidences of transitional epithelium and respiratory epithelium hyperplasia were significantly increased in all dosed groups of males and females. Except for olfactory epithelium respiratory metaplasia in 100 mg/kg females, the incidences of atrophy, respiratory metaplasia, nerve atrophy, and pigmentation were significantly increased in the olfactory epithelium of all dosed groups of males and females. Incidences of goblet cell hyperplasia in the respiratory epithelium were significantly increased in 300 and 1,000 mg/kg males and females, and incidences of chronic active inflammation were significantly increased in 1,000 mg/kg males and females. The incidence of submucosa fibrosis was significantly increased in 1,000 mg/kg males.

The incidences of mononuclear cell leukemia in 300 and 1,000 mg/kg males were significantly greater than that in the vehicle controls.

Dose-related increased severity of kidney nephropathy was noted in all dosed groups of males.

Two-year study in mice

Groups of 50 male and 50 female mice were administered 0, 200, 600, or 2,000 mg Ginkgo biloba extract/kg body weight in corn oil by gavage, 5 days per week for 104 weeks. Survival of 600 and 2,000 mg/kg males was significantly less than that of the vehicle controls; survival of 600 mg/kg females was significantly greater than that of the vehicle controls. Mean body weights of 600 and 2,000 mg/kg males were less (10% or more) than those of the vehicle controls after weeks 85 and 77, respectively; mean body weights of 2,000 mg/kg females were generally less than those of the vehicle controls between weeks 17 and 69 and after week 93.

In the liver, there were significantly increased incidences of hepatocellular adenoma in all dosed groups of females, hepatocellular carcinoma in all dosed groups of males and 2,000 mg/kg females, and hepatoblastoma in all dosed groups of males and 600 and 2,000 mg/kg females. The increased incidences of these neoplasms were primarily due to increased incidences of multiple adenoma, carcinoma, and hepatoblastoma. Except for the incidences of hepatocellular carcinoma or hepatoblastoma (combined) in 200 and 600 mg/kg females, the incidences of hepatocellular adenoma or carcinoma (combined), hepatocellular carcinoma or hepatoblastoma (combined), and hepatocellular adenoma, hepatocellular carcinoma, or hepatoblastoma (combined) were significantly increased in all dosed groups of males and females. Significantly increased incidences of nonneoplastic liver lesions included hypertrophy in all dosed groups of males and females, erythrophagocytosis in all dosed groups of males and in 600 and 2,000 mg/kg females, hematopoietic cell proliferation, inflammation, and necrosis in 600 and 2,000 mg/kg males, and cytoplasmic vacuolization, eosinophilic focus, and mixed cell focus in all dosed groups of females.

In the thyroid gland, two incidences each of follicular cell adenoma occurred in the 600 and 2,000 mg/kg male groups. The incidence of follicle hyperplasia was significantly increased in 2,000 mg/kg males, and the incidences of follicular cell hypertrophy were significantly increased in 2,000 mg/kg males and 600 and 2,000 mg/kg females.

In the forestomach, the incidences of inflammation, epithelium hyperplasia, and epithelium hyperkeratosis were significantly increased in all dosed groups of males and in 2,000 mg/kg females; the incidences of epithelium ulcer were significantly increased in 2,000 mg/kg males and females.

In the nose, the incidences of hyaline droplet accumulation in the olfactory epithelium were significantly increased in 2,000 mg/kg males and females; the incidences of pigmentation in the olfactory epithelium were significantly increased in 2,000 mg/kg males and 600 and 2,000 mg/kg females.

Genetic toxicology

Ginkgo biloba extract was mutagenic in S. typhimurium strains TA98 and TA100, and in E. coli strain WP2 uvrA/pKM101, with and without exogenous metabolic activation. Results of a peripheral blood micronucleus test in male and female B6C3F1/N mice administered Ginkgo biloba extract for 3 months by gavage were negative in males but judged to be equivocal in females based on a significant trend test.

Conclusions

Under the conditions of these 2-year gavage studies, there was some evidence of carcinogenic activity of Ginkgo biloba extract in male F344/N rats based on increased incidences of thyroid gland follicular cell adenoma. The increased incidences of mononuclear cell leukemia and hepatocellular adenoma may have been related to Ginkgo biloba extract administration. There was some evidence of carcinogenic activity of Ginkgo biloba extract in female F344/N rats based on increased incidences of thyroid gland follicular cell neoplasms. Increased occurrence of respiratory epithelium adenomas in the nose may have been related to Ginkgo biloba extract administration. There was clear evidence of carcinogenic activity of Ginkgo biloba extract in male B6C3F1/N mice based on increased incidences of hepatocellular carcinoma and hepatoblastoma. The increased incidences of thyroid gland follicular cell adenoma were also related to Ginkgo biloba extract administration. There was clear evidence of carcinogenic activity of Ginkgo biloba extract in female B6C3F1/N mice based on increased incidences of hepatocellular adenoma, hepatocellular carcinoma, and hepatoblastoma.

Administration of Ginkgo biloba extract resulted in increased incidences of nonneoplastic lesions in the liver, thyroid gland, and nose of male and female rats and mice and the forestomach of male and female mice. Increased severity of nephropathy in male rats was also due to administration of Ginkgo biloba extract. 

Studies

Summary of the Two-year Carcinogenesis and Genetic Toxicology Studies of Ginkgo Biloba Extract
  Male
F344/N Rats
Female
F344/N Rats
Male
B6C3F1/N Mice
Female
B6C3F1/N Mice
Doses in corn oil by gavage 0, 100, 300, or 1,000 mg/kg 0, 100, 300, or 1,000 mg/kg 0, 200, 600, or 2,000 mg/kg 0, 200, 600, or 2,000 mg/kg
Body weights 300 mg/kg group ≥10% less than the vehicle control group after week 93; 1,000 mg/kg group ≥10% less than the vehicle control group after week 89 300 mg/kg group ≥10% less than the vehicle control group after week 93; 1,000 mg/kg group ≥10% less than the vehicle control group after week 89 600 and 2,000 mg/kg groups ≥10% less than the vehicle control group after week 85 and 77, respectively 2,000 mg/kg group ≥10% less than the vehicle control group between weeks 17 and 69 and after week 93
Survival rates 38/50, 37/50, 31/50, 16/50 37/50, 27/50, 37/50, 32/50 34/50, 27/50, 21/50, 23/50 31/50, 36/50, 43/50, 36/50
Nonneoplastic effects Liver: hepatocyte, hypertrophy (1/50, 17/50, 26/50, 27/50); bile duct, hyperplasia (32/50, 43/50, 46/50, 46/50); oval cell, hyperplasia (0/50, 1/50, 1/50, 10/50); degeneration, cystic (4/50, 14/50, 10/50, 14/50); necrosis (1/50, 4/50, 6/50, 7/50)

Thyroid gland
: follicular cell hypertrophy (13/50, 37/50, 41/49, 41/45); follicle, hyperplasia (0/50, 7/50, 9/49, 5/45)

Kidney
: severity of nephropathy (1.7, 2.0, 2.4, 2.9)

Nose
: transitional epithelium, hyperplasia (2/50, 18/49, 43/49, 31/50); respiratory epithelium, hyperplasia (14/50, 28/49, 45/49, 35/50); olfactory epithelium, atrophy (1/50, 26/49, 37/49, 31/50); olfactory epithelium, respiratory metaplasia (9/50, 30/49, 40/49, 32/50); nerve, olfactory epithelium, atrophy (0/50, 17/49, 14/49, 23/50); olfactory epithelium, pigmentation (0/50, 39/49, 42/49, 30/50); inflammation, chronic active (33/50, 32/49, 38/49, 46/50); goblet cell, respiratory epithelium, hyperplasia (20/50, 18/49, 41/49, 34/50); submucosa, fibrosis (0/50, 0/49, 0/49, 8/50)
Liver: hepatocyte, hypertrophy (7/50, 15/50, 27/50, 33/50); bile duct, hyperplasia (11/50, 31/50, 31/50, 33/50); fatty change, focal (11/50, 25/50, 30/50, 25/50)

Thyroid gland
: follicular cell, hypertrophy (15/49, 41/50, 45/49, 48/49)

Nose
: transitional epithelium, hyperplasia (0/49, 6/49, 32/50, 36/46); respiratory epithelium, hyperplasia (9/49, 6/49, 19/50, 34/46); olfactory epithelium, atrophy (0/49, 18/49, 25/50, 37/46); olfactory epithelium, respiratory metaplasia (8/49, 4/49, 32/50, 37/46); nerve, olfactory epithelium, atrophy (0/49, 15/49, 22/50, 33/46); olfactory epithelium, pigmentation (0/49, 37/49, 43/50, 40/46); inflammation, chronic active (22/49, 16/49, 26/50, 38/46); goblet cell, respiratory epithelium, hyperplasia (6/49, 2/49, 18/50, 35/46)
Liver: hypertrophy (3/50, 19/50, 35/50, 23/50); erythrophagocytosis (0/50, 4/50, 11/50, 7/50); hematopoietic cell proliferation (4/50, (9/50, 12/50, 14/50); inflammation (28/50, 35/50, 42/50, 39/50); necrosis (9/50, 15/50, 17/50, 19/50)

Thyroid gland
: follicle hyperplasia (2/49, 1/49, 7/50, 25/50); follicular cell hypertrophy (2/49, 0/49, 2/50, 38/50)

Forestomach
: inflammation (11/50, 24/50, 21/50, 45/50); epithelium hyperplasia (14/50, 27/50, 27/50, 45/50); epithelium hyperkeratosis (11/50, 24/50, 24/50, 46/50); epithelium ulcer (7/50, 10/50, 12/50, 24/50)

Nose
: olfactory epithelium, hyaline droplet accumulation (18/50, 16/50, 15/50, 28/50); olfactory epithelium, pigmentation (0/50, 1/50, 3/50, 13/50)
Liver: hypertrophy (0/50, 18/50, 37/50, 37/50); erythrophagocytosis (0/50, 3/50, 7/50, 16/50); vacuolization cytoplasmic (18/50, 38/50, 44/50, 35/50); eosinophilic focus (26/50, 39/50, 43/50, 45/50); mixed cell focus (7/50, 27/50, 31/50, 31/50)

Thyroid gland
: follicular cell hypertrophy (1/49, 5/48, 9/49, 39/48)

Forestomach
: inflammation (4/50, 6/50, 5/50, 19/50); epithelium hyperplasia (8/50, 18/50, 11/50, 20/50); epithelium hyperkeratosis (3/50, 11/50, 5/50, 20/50); epithelium ulcer (1/50, 1/50, 1/50, 11/50)

Nose
: olfactory epithelium, hyaline droplet accumulation (5/50, 3/50, 12/50, 17/50); olfactory epithelium, pigmentation (0/50, 1/50, 6/50, 13/50)
Neoplastic effects Thyroid gland: follicular cell, adenoma (2/50, 1/50, 3/49, 5/45) Thyroid gland: follicular cell, adenoma (1/49, 0/50, 3/49, 1/49); follicular cell, carcinoma (0/49, 0/50, 1/49, 1/49) Liver: hepatocellular carcinoma (22/50, 31/50, 41/50, 47/50); hepatoblastoma (3/50, 28/50, 36/50, 38/50)

Thyroid gland
: follicular cell, adenoma (0/50, 0/50, 2/50, 2/50)
Liver: hepatocellular adenoma (17/50, 37/50, 41/50, 48/50); hepatocellular carcinoma (9/50, 10/50, 15/50, 44/50); hepatoblastoma (1/50, 1/50, 8/50, 11/50)
Equivocal findings Mononuclear cell leukemia: (9/50, 12/50, 22/50, 21/50)

Liver
: hepatocellular adenoma (0/50, 3/50, 3/50, 0/50)
Nose: respiratory epithelium, adenoma (0/49, 0/49, 2/50, 0/46) None None
Level of evidence of carcinogenic activity Some evidence Some evidence Clear evidence Clear evidence
Genetic Toxicology
Assay Results
Bacterial gene mutations:
 
Positive in S. typhimurium strains TA98 and TA100 with and without S9; positive in E. coli strain WP2 uvrA/pKM101 with and without S9
Micronucleated erythrocytes
Mouse peripheral blood in vivo:
Negative in males and equivocal in females