Chloral hydrate is used medically as a sedative or hypnotic and as a rubefacient in topical preparations, and it is often given to children as a sedative during dental and other medical procedures. Chloral hydrate is used as a central nervous system depressant and sedative in veterinary medicine and as a general anesthetic in cattle and horses. It is a byproduct of the chlorination of water and has been detected in plant effluent after the bleaching of softwood pulp. Chloral, the anhydrous form of chloral hydrate, is used as a synthetic intermediate in the production of insecticides and herbicides. Chloral hydrate was nominated for study by the Food and Drug Administration based upon widespread human exposure and its potential hepatotoxicity and the toxicity of related chemicals. One goal of the study was to assess the effect of the animal's age and the duration of dosing on the tumorigenicity of chloral hydrate. Beginning on post-natal day 28, female B6C3F1 mice received chloral hydrate (99.5% pure) in water by gavage for 3, 6, or 12 months, 2 years, or as a single dose;on postnatal day 15, male and female B6C3F1 mice received a single dose by gavage. Tumorigenicity was assessed for 2 years after the initial dose. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, Drosophila melanogaster, and mouse bone marrow cells.
Groups of female mice (regimens A, B, C, and D) and groups of male mice (regimen E) received chloral hydrate in distilled water by gavage; control groups received distilled water only. In regimen A, groups of 48 female mice received 0, 25, 50, or 100 mg chloral hydrate/kg body weight 5 days per week for 105 weeks beginning when they were 28 days old. In regimen B, 24 female mice received 0 mg/kg and three groups of 48 female mice received 100 mg/kg 5 days per week beginning when they were 28 days old. Eight mice from the 0 and 100 mg/kg groups were killed at 3, 6, and 12 months. The remaining mice were held without dosing for the duration of the 2-year study. In regimen C, groups of 48 female mice received a single dose of 0, 10, 25, or 50 mg/kg when they were 28 days old and were held for 105 weeks. In regimens D and E, groups of 48 female and 48 male mice, respectively, received a single dose of 0, 10, 25 or 50 mg/kg when they were 15 days old and were held for 105 weeks. Additional groups of four mice from regimens C, D, and E and mice killed at 3 or 6 months from regimen B (eight mice per group) were designated for hepatic cell proliferation analyses; mice killed at 3 or 6 months in regimen B were also designated for apoptosis analyses.
Survival and Body Weights
Survival of all dosed mice in all regimens was similar to that of the vehicle control groups. Mean body weights of 100 mg/kg female mice in regimen B dosed for 3 or 6 months were generally greater than those of the vehicle controls during the second year of the study. Mean body weights of 25 mg/kg male mice in regimen E were generally less than those of the controls beginning at week 19; mean body weights of 10 and 50 mg/kg mice were generally less than those of the vehicle controls beginning at week 80.
A dose-related and significant increase in the incidence of pars distalis adenoma occurred in regimen A 100 mg/kg females. There was also a time-related increase in the incidence of adenoma in female mice administered 100 mg/kg for up to 24 months in regimen B, and the increase in the incidence of this neoplasm at 24 months was significant. There was a significant increase in the severity of pars distalis hyperplasia in regimen A 100 mg/kg female mice.
Chloral hydrate was mutagenic in vitro and in vivo. It induced mutations in Salmonella typhimurium strain TA100, with and without liver S9 activation; an equivocal response was obtained in S. typhimurium strain TA98 in the absence of S9, and no mutagenicity was detected with strain TA1535 or TA1537, with or without S9. Chloral hydrate was shown to produce chromosomal damage in mammalian cells. It induced significant increases in sister chromatid exchanges and chromosomal aberrations in cultured Chinese hamster ovary cells, with and without S9. Results of sex-linked recessive lethal (SLRL) tests in Drosophila melanogaster were inconclusive. Chloral hydrate, administered by feeding, produced an inconclusive increase in SLRL mutations in the germ cells of male flies. Results of an in vivo mouse bone marrow micronucleus test with chloral hydrate were positive.
Under the conditions of this two-year gavage study, there was equivocal evidence of carcinogenic activity of chloral hydrate in female B6C3F1 mice treated continuously for two years based on increased incidences of pituitary gland pars distalis adenoma. No increased incidences of neoplasms were seen in female B6C3F1 mice that received a single dose of chloral hydrate at 15 or 28 days of age or in male B6C3F1 mice that received a single dose of chloral hydrate at 15 days of age. No hepatic carcinogenicity was seen under all dosing conditions.