[Code of Federal Regulations]
[Title 40, Volume 31]
[Revised as of July 1, 2006]
From the U.S. Government Printing Office via GPO Access
[CITE: 40CFR795.225]
[Page 63-65]
TITLE 40--PROTECTION OF ENVIRONMENT
CHAPTER I--ENVIRONMENTAL PROTECTION AGENCY (CONTINUED)
PART 795_PROVISIONAL TEST GUIDELINES--Table of Contents
Subpart D_Provisional Health Effects Guidelines
Sec. 795.225 Dermal pharmacokinetics of DGBE and DGBA.
(a) Purpose. The purpose of these studies is to determine:
(1) The absorption of diethylene glycol butyl ether (DGBE) after
administration by the dermal route.
(2) The biotransformation of DGBE administered dermally.
(3) The dermal absorption of DGBE and diethylene glycol butyl ether
acetate (DGBA).
(b) Test procedures--(1) Animal selection--(i) Species. The species
utilized for investigating DGBE and DGBA shall be the rat, a species for
which historical
[[Page 64]]
data on the toxicity and carcinogenicity of many compounds are available
and which is used extensively in percutaneous absorption studies.
(ii) Animals. Adult female Sprague Dawley rats shall be used. The
rats shall be 7 to 8 weeks old and weigh 180 to 220 grams. Prior to
testing, the animals shall be selected at random for each group. Animals
showing signs of ill health shall not be used.
(iii) Animal care. (A) The animals should be housed in
environmentally controlled rooms with 10 to 15 air changes per hour. The
rooms should be maintained at a temperature of 25 2 [deg]C and humidity of 50 10
percent with a 12-hour light/dark cycle per day. The rats should be
isolated for at least 7 days prior to use.
(B) During the acclimatization period, the rats should be housed in
cages on hardwood chip bedding. All animals shall be provided with
conventional laboratory diets and water ad libitum.
(2) Administration of DGBE and DGBA--(i) Test substances. These
studies require the use of \14\C-labeled DGBE and DGBA. The use of
\14\C-DGBE and \14\C-DGBA is required for the determinations in
paragraphs (a) (1), (2), and (3) of this section because they will
facilitate the work and improve the reliability of quantitative
determinations.
(ii) Dosage and treatment. (A) Two doses of DGBA shall be used in
the study, a ``low'' dose and a ``high'' dose. Three doses of DGBE shall
be used in the study, a neat ``low'' dose, an aqueous ``low'' dose, and
neat ``high'' dose. When administered dermally, the ``high'' dose level
should ideally induce some overt toxicity such as weight loss. The
``low'' dose level should correspond to a no observed effect level.
(B) For dermal treatment, the doses shall be applied in a volume
adequate to deliver the prescribed doses. The backs of the rats should
be lightly shaved with an electric clipper shortly before treatment. The
dose shall be applied with a micropipette on a specific area (for
example, 2 cm\2\) on the freshly shaven skin.
(iii) Washing efficiency study. Before initiation of the dermal
absorption studies described in paragraph (b)(2)(iv)(A) of this section,
an initial washing efficiency experiment shall be performed to assess
the extent of removal of the applied DGBE and DGBA by washing with soap
and water. Groups of four rats should be lightly anesthetized with
sodium pentobarbital. These animals shall then be treated with dermal
doses of test substance at the low dose level. Soon after application (5
to 10 minutes) the treated animals shall be washed with soap and water
then housed in individual metabolism cages for excreta collection. Urine
and feces shall be collected at 8, 24, and 48 hours following dosing.
Collection of excreta shall continue every 24 hours if a significant
amounts of DGBE, DGBA, or metabolites continue to be eliminated.
(iv) Determination of absorption, biotransformation, and excretion.
(A) Eight animals shall be dosed once dermally with the low dose of
\14\C-DGBE.
(B) Eight animals shall be dosed once dermally with the high dose of
\14\C-DGBE.
(C) Eight animals shall be dosed once dermally with the low dose of
\14\C-DGBA.
(D) Eight animals shall be dosed once dermally with the high dose of
\14\C-DGBA.
(E) The high and low doses of \14\C-DGBE and \14\C-DGBA shall be
kept on the skin for 24 hours. After application, the animals shall be
placed in metabolism cages for excreta collection. After 24 hours, any
test material remaining on the skin will be washed off and the
containment cell removed. Radiolabeled material in the wash will be
accounted for in the total recovery. Urine and feces shall be collected
at 8, 24, 48, 72, and 96 hours after dosing, and if necessary, daily
thereafter until at least 90 percent of the dose has been excreted or
until 7 days after dosing, whichever occurs first.
(3) Observation of animals--(i) Urinary and fecal excretion. The
quantities of total \14\C excreted in urine and feces by rats dosed as
specified in paragraph (b)(2)(iv) of this section shall be determined at
8, 24, 48, 72 and 96 hours after dosing, and if necessary, daily
thereafter until at least 90 percent of the dose has been excreted or
until 7 days after dosing (whichever occurs first). Four animals from
each group shall be used for this purpose.
[[Page 65]]
(ii) Biotransformation after dermal dosing. Appropriate qualitative
and quantitative methods shall be used to assay urine specimens
collected from rats dosed with DGBE as specified in paragraph (b)(2)(iv)
of this section. Any metabolite which comprises greater than 10 percent
of the dose shall be identified.
(c) Data and reporting--(1) Treatment of results. Data shall be
summarized in tabular form.
(2) Evaluation of results. All observed results, quantitative or
incidental, shall be evaluated by an appropriate statistical method.
(3) Test report. In addition to the reporting requirements as
specified in the TSCA Good Laboratory Practice Standards, in part 792,
subpart J of this chapter, the following specific information shall be
reported:
(i) Species, strain, and supplier of laboratory animals.
(ii) Information on the degree (i.e., specific activity for a
radiolabel) and sites of labeling of the test substances.
(iii) A full description of the sensitivity and precision of all
procedures used to produce the data.
(iv) Relative percent absorption by the dermal route for rats
administered low and high doses of \14\C-DGBE and \14\C-DGBA.
(v) Quantity of isotope, together with percent recovery of the
administered dose, in feces and urine.
(vi) Biotransformation pathways and quantities of DGBE and
metabolites in urine collected after administering single high and low
dermal doses to rats.
[53 FR 5946, Feb. 26, 1988, as amended at 54 FR 41834, Oct. 12, 1989]