§ 799.9305 TSCA Repeated dose 28-day oral toxicity study in rodents.

(a) Scope—(1) Applicability. This section is intended to meet testing requirements of the Toxic Substances Control Act (TSCA) (15 U.S.C. 2601).

(2) Source. The source material used in developing this TSCA test guideline is the Office of Prevention, Pesticides and Toxic Substances (OPPTS) harmonized test guideline 870.3050 (July 2000, final guidelines). This source is available at the address in paragraph (h) of this section.

(b) Purpose. (1) In the assessment and evaluation of the toxic characteristics of a chemical, the determination of oral toxicity using repeated doses may be carried out after initial information on toxicity has been obtained by acute testing. This study provides information on the possible health hazards likely to arise from repeated exposure over a relatively limited period of time. The method comprises the basic repeated dose toxicity study that may be used for chemicals on which a 90-day study is not warranted (e.g., when the production volume does not exceed certain limits) or as a preliminary to a long term study. The duration of exposure should normally be 28 days although a 14-day study may be appropriate in certain circumstances; justification for use of a 14-day exposure period should be provided.

(2) This section places emphasis on neurological effects as a specific endpoint, and the need for careful clinical observations of the animals, so as to obtain as much information as possible, is stressed. The method should identify chemicals with neurotoxic potential, which may warrant further in-depth investigation of this aspect. In addition, the method may give an indication of immunological effects and reproductive organ toxicity.

(c) Definitions. The definitions in section 3 of TSCA and in 40 CFR Part 792—Good Laboratory Practice Standards apply to this section. The following definitions also apply to this section.

Dosage is a general term comprising of dose, its frequency and the duration of dosing.

Dose is the amount of test substance administered. Dose is expressed as weight (g, mg) or as weight of test substance per unit weight of test animal (e.g., mg/kg), or as constant dietary concentrations (parts per million (ppm)).

No-observed-effects level (NOEL) is the maximum dose used in a study which produces no adverse effects. The NOEL is usually expressed in terms of the weight of a test substance given daily per unit weight of test animals (milligrams per kilograms per day).

(d) Principle of the test. The test substance is orally administered daily in graduated doses to several groups of experimental animals, one dose level per group for a period of 28 days. During the period of administration the animals are observed closely, each day for signs of toxicity. Animals which die or are sacrificed during the test are necropsied and at the conclusion of the test surviving animals are sacrificed and necropsied.

(e) Description of the method—(1) Selection of animal species. The preferred rodent species is the rat, although other rodent species may be used. Commonly used laboratory strains of young healthy adult animals should be employed. The females should be nulliparous and non-pregnant. Dosing should begin as soon as possible after weaning and, in any case, before the animals are 9 weeks old. At the commencement of the study the weight variation of animals used should be minimal and not exceed ±20% of the mean weight of each sex. Where a repeated dose oral study is conducted as a preliminary to a long term study, preferably animals from the same strain and source should be used in both studies.

(2) Housing and feeding conditions. The temperature in the experimental animal room should be 22 °C (±3 °C). Although the relative humidity should be at least 30% and preferably not to exceed 70% other than during room cleaning, the aim should be 50-60%. Lighting should be artificial, the sequence being 12 hours light, 12 hours dark. For feeding, conventional laboratory diets may be used with an unlimited supply of drinking water. The choice of diet may be influenced by the need to ensure a suitable admixture of a test substance when administered by this method. Animals may be housed individually, or be caged in small groups of the same sex; for group caging, no more than five animals should be housed per cage.

(3) Preparation of animals. Healthy young adult animals must be randomly assigned to the control and treatment groups. Cages should be arranged in such a way that possible effects due to cage placement are minimized. The animals are identified uniquely and kept in their cages for at least 5 days prior to the start of the study to allow for acclimatization to the laboratory conditions.