[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: 40CFR799.2155]
[Page 243-248]
TITLE 40--PROTECTION OF ENVIRONMENT
CHAPTER I--ENVIRONMENTAL PROTECTION AGENCY (CONTINUED)
PART 799_IDENTIFICATION OF SPECIFIC CHEMICAL SUBSTANCE AND MIXTURE
TESTING REQUIREMENTS--Table of Contents
Subpart B_Specific Chemical Test Rules
Sec. 799.2155 Commercial hexane.
(a) Identification of test substance. (1) ``Commercial hexane,'' for
purposes of this section, is a product obtained from crude oil, natural
gas liquids, or petroleum refinery processing in accordance with the
American Society for Testing and Materials Designation D 1836-83 (ASTM D
1836), consists primarily of six-carbon alkanes or cycloalkanes, and
contains at least 40 liquid volume percent n-hexane (CAS No. 110-54-3)
and at least 5 liquid volume percent methylcyclopentane (MCP; CAS No.
96-37-7). ASTM D 1836, formally entitled ``Standard Specification for
Commercial Hexanes,'' is published in 1986 Annual Book of ASTM
Standards: Petroleum Products and Lubricants, ASTM D 1836-83, pp. 966-
967, 1986, is incorporated by reference, and is available for public
inspection at the National Archives and Records Administration (NARA).
For information on the availability of this material at NARA, call 202-
741-6030, or go to: http://www.archives.gov/federal--register/code--of--
federal--regulations/ibr--locations.html. This incorporation by
reference was approved by the Director of the Office of the Federal
Register in accordance with 5 U.S.C. 522(a) and 1 CFR part 51. This
material is incorporated as it exists on the date of approval, and a
notice of any change in this material will be published in the Federal
Register. Copies of the incorporated material may be obtained from the
Non-Confidential Information Center (NCIC) (7407), Office of Pollution
Prevention and Toxics, U.S. Environmental Protection Agency, Room B-607
NEM, 401 M St., SW., Washington, DC 20460, between the hours of 12 p.m.
and 4 p.m. weekdays excluding legal holidays.
(2) The commercial hexane test substance, for purposes of this
section, is a product which conforms to the specifications of ASTM D1836
and contains at least 40 liquid volume percent but no more than 55
liquid volume percent n-hexane and no less than 10 liquid volume percent
MCP.
(b) Persons required to submit study plans, conduct tests, and
submit data. All persons who manufacture (including import) or process
or intend to manufacture or process commercial hexane, as defined in
paragraph (a)(1) of this section and other than as an impurity, from the
effective date of the final rule to the end of the reimbursement period
shall submit letters of intent to conduct testing, submit study plans,
conduct tests in accordance with part 792 of this chapter, and submit
data, or submit exemption applications, as specified in this section,
subpart A of this part, and part 790 of this chapter for single-phase
rulemaking. Persons who manufacture commercial hexane as a byproduct are
covered by the requirements of this section. Notwithstanding Sec.
790.50(a)(1) of this chapter, persons who notify EPA of their intent to
conduct neurotoxicity testing in compliance with paragraph (c)(7) of
this section may submit study plans for those tests less than 45 days
before beginning testing provided that EPA receives the study plans
before this testing begins.
(c) Health effects testing--(1) Subchronic inhalation toxicity--(i)
Required testing. (A) A subchronic inhalation toxicity test shall be
conducted with commercial hexane in accordance with Sec. 798.2450 of
this chapter except for the provisions in paragraphs (d)(4)(ii) and (5)
of Sec. 798.2450.
(B) For the purposes of this section, the following provisions also
apply:
(1) High dose level. The highest concentration should result in
toxic effects but neither produce an incidence of fatalities which would
prevent a meaningful evaluation nor exceed the lower explosive limit of
commercial hexane.
(2) Exposure conditions. Animals shall be dosed for 6 hours/day, 5
days/week for 90 days.
(ii) Reporting requirements. (A) The subchronic inhalation toxicity
test
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shall be completed and the final report submitted to EPA within 15
months of the effective date of the final rule.
(B) Interim progress reports shall be submitted to EPA for the
subchronic inhalation toxcity test at 6-month intervals beginning 6
months after the effective date of the final rule, until the final
report is submitted to EPA.
(2) Oncogenicity--(i) Required testing. (A) An oncogenicity test
shall be conducted with commercial hexane in accordance with Sec.
798.3300 of this chapter except for the provisions in paragraphs
(b)(3)(ii) and (6) of Sec. 798.3300.
(B) For the purposes of this section, the following provisions also
apply:
(1) High dose level. The high dose level should elicit signs of
minimal toxicity without substantially altering the normal life span and
should not exceed the lower explosive limit of commercial hexane.
(2) Administration of test substance. Animals shall be exposed to
commercial hexane by inhalation.
(ii) Reporting requirements. (A) The oncogenicity test shall be
completed and the final report submitted to EPA within 53 months of the
effective date of the final rule. The mouse portion of the oncogenicity
study shall be submitted by June 5, 1993.
(B) Interim progress reports shall be submitted to EPA for the
oncogenicity test at 6-month intervals beginning 6 months after the
effective date of the final rule, until the final report is submitted to
EPA.
(3) Reproduction and fertility effects--(i) Required testing. (A) A
reproduction and fertility effects test shall be conducted with
commercial hexane in accordance with Sec. 798.4700 of this chapter
except for the provisions in paragraphs (c)(3)(ii) and (5) of Sec.
798.4700.
(B) For the purposes of this section, the following provisions also
apply:
(1) High dose level. The highest dose level should induce toxicity
but not high levels of mortality in the parental (P) animals. In
addition, the highest dose level should not exceed the lower explosive
limit of commercial hexane.
(2) Administration of test substance. Animals shall be exposed to
commercial hexane by inhalation.
(ii) Reporting requirements. (A) The reproduction and fertility
effects test shall be completed and the final report submitted to EPA
within 29 months of the effective date of the final rule.
(B) Interim progress reports shall be submitted to EPA for the
reproduction and fertility effects test at 6-month intervals beginning 6
months after the effective date of the final rule, until the final
report is submitted to EPA.
(4) Inhalation developmental toxicity--(i) Required testing. (A) An
inhalation developmental toxicity test shall be conducted with
commercial hexane in accordance with Sec. 795.4350 of this chapter
except for the provisions in paragraph (e)(3)(iv) of Sec. 798.4350.
(B) For the purposes of this section, the following provisions also
apply:
(1) High dose level. Unless limited by the physical/chemical nature
or biological properties of the test substance, the highest
concentration level shall induce some overt maternal toxicity such as
reduced body weight or body weight gain, but not more than 10 percent
maternal deaths. In addition, the highest dose level should not exceed
the lower explosive limit of commercial hexane.
(2) [Reserved]
(ii) Reporting requirements. (A) The inhalation developmental
toxicity test shall be completed and the final report submitted to EPA
within 12 months of the effective date of the final rule.
(B) Interim progress reports shall be submitted to EPA for the
inhalation developmental toxicity test at 6-month intervals beginning 6
months after the effective date of the final rule, until the final
report is submitted to EPA.
(5) Mutagenic effects--gene mutations--(i) Required testing. (A)(1)
A Salmonella typhimurium reverse mutation assay shall be conducted with
commercial hexane in accordance with Sec. 798.5265 of this chapter
except for the provisions in paragraphs (d)(4) and (e) of Sec.
798.5265.
(2) For the purposes of this section, the following provisions also
apply:
(i) Metabolic activation. Bacteria shall be exposed to commercial
hexane both in the presence and absence of an appropriate metabolic
activation system.
(ii) Test performance. The assay shall be performed using the
desiccator method described as follows: The agar overlay plates shall be
placed uncovered in a 9-liter desiccator. A volume of
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the liquid test substance shall be added to the glass Petri dish
suspended beneath the porcelain shelf of the desiccator. The highest
exposure concentration should not result in a vapor concentration which
exceeds the lower explosive limit of commerical hexane. A magnetic
stirring bar to serve as a fan to assure rapid and even distribution of
the vapor shall be placed on the bottom of the inside of the desiccator.
The desiccator shall be placed on a magnetic stirrer within a 37[deg] C
room or chamber for 7 to 10 hours. The plates shall then be removed,
their lids replaced, followed by incubation for an additional 40 hours
at 37[deg] C before counting. An appropriate selective medium with an
adequate overlay agar shall be used. All plating should be done in at
least triplicate.
(B)(1) A gene mutation test in mammalian cells shall be conducted
with commercial hexane in accordance with Sec. 798.5300 of this chapter
except for the provisions in paragraphs (d)(3)(ii) and (4) of Sec.
798.5300 if the results from the Salmonella typhimurium test conducted
pursuant to paragraph (c)(5)(i)(A) of this section are negative.
(2) For the purposes of this section, the following provisions also
apply:
(i) Cell growth and maintenance. Appropriate culture media and
incubation conditions (culture vessels, CO2 concentrations,
temperature, and humidity) shall be used. The cell culture shall be
directly dosed by pipetting liquid commercial hexane mixed with liquid
DMSO into the culture medium. Cells shall be exposed to test substance
both in the presence and absence of an appropriate metabolic activation
system.
(ii) [Reserved]
(C)(1) A sex-linked recessive lethal test in Drosophila melanogaster
shall be conducted with commercial hexane in accordance with Sec.
798.5275 of this chapter except for the provisions in paragraphs (d)(5)
(ii) and (iii) of Sec. 798.5275, unless the results of both the
Salmonella typhimurium test conducted pursuant to paragraph (c)(5)(i)(A)
of this section and the mammalian cells in the culture gene mutation
test conducted pursuant to paragraph (c)(5)(i)(B) of this section, if
required, are negative.
(2) For the purposes of this section, the following provisions also
apply:
(i) Dose levels. For the initial assessment of mutagenicity, it is
sufficient to test a single dose of the test substance for screening
purposes. This dose should be the maximum tolerated dose, or that which
produces some indication of toxicity or shall be the highest dose
attainable and should not exceed the lower explosive limit of commercial
hexane. For dose-response purposes, at least three additional dose
levels should be used.
(ii) Route of administration. The route of administration shall be
by exposure to commercial hexane vapors.
(D)(1) Unless the results of the sex-linked recessive lethal test in
Drosophila melanogaster conducted with commercial hexane pursuant to
paragraph (c)(5)(i)(C) of this section are negative, EPA shall conduct a
public program review of all of the mutagenicity data available for this
substance. If, after this review, EPA decides that testing of commercial
hexane for causing heritable gene mutations in mammals is necessary, it
shall notify the test sponsor by certified letter or Federal Register
notice that testing shall be initiated in either the mouse visible
specific locus test or the mouse biochemical specific locus test. The
mouse visible specific locus test, if conducted, shall be performed for
commercial hexane in accordance with Sec. 798.5200 of this chapter
except for the provisions in paragraphs (d)(5)(ii) and (d)(5)(iii) of
Sec. 798.5200. The mouse biochemical specific locus test, if conducted,
shall be performed for commercial hexane in accordance with Sec.
798.5195 of this chapter except for the provisions in paragraphs
(d)(5)(ii) and (d)(5)(iii) of Sec. 798.5195.
(2) For the purposes of this section, the following provisions also
apply:
(i) Dose levels. A minimum of two dose levels shall be tested. The
highest dose tested shall be the highest dose tolerated without toxic
effects, provided that any temporary sterility induced due to
elimination of spermatogonia is of only moderate duration, as determined
by a return of males to fertility within 80 days of treatment, or shall
be the highest dose attainable below the lower explosive
[[Page 246]]
limit concentration of commercial hexane. Exposure shall be for 6 hours
a day. Duration of exposure shall be dependent upon the accumulated
total dose desired for each group.
(ii) Route of administration. Animals shall be exposed to commercial
hexane by inhalation.
(ii) Reporting requirements. (A) The gene mutation tests shall be
completed and final reports submitted to EPA as follows:
(1) The Salmonella typhimurium reverse mutation assay within 8
months of the effective date of the final rule.
(2) The gene mutation in mammalian cells assay within 17 months of
the effective date of the final rule.
(3) The sex-linked recessive-lethal test in Drosophila melanogaster
within 24 months of the effective date of the final rule.
(4) The mouse visible specific locus test or the mouse biochemical
specific locus test shall be completed and a final report shall be
submitted to EPA within 51 months of the date on which the test sponsor
is notified by EPA by certified letter or Federal Register notice that
testing shall be initiated.
(B) Interim progress reports for each test shall be submitted to EPA
for the gene mutation in mammalian cells assay and Drosophila sex-linked
recessive lethal test at 6-month intervals beginning 6 months after the
effective date of the final rule, until the applicable final report is
submitted to EPA.
(C) Interim progress reports for either the mouse visible specific
locus test or the mouse biochemical specific locus test shall be
submitted to EPA at 6-month intervals, beginning 6 months after EPA's
notification of the test sponsor that testing should be initiated, until
the applicable final report is submitted to EPA.
(6) Mutagenic effects--chromosomal aberrations--(i) Required
testing. (A)(1) An in vitro cytogenetics test shall be conducted with
commercial hexane in accordance with Sec. 798.5375 of this chapter
except for the provisions in paragraph (e)(3) of Sec. 798.5375.
(2) For the purposes of this section, the following provisions also
apply:
(i) Treatment with test substance. The test substance shall be added
in liquid form mixed with DMSO to the treatment vessels.
(ii) [Reserved]
(B)(1) An in vivo cytogenetics test shall be conducted with
commercial hexane in accordance with Sec. 798.5385 of this chapter
except for the provisions in paragraphs (d)(5) (ii), (iii) and (iv) of
Sec. 798.5385, if the in vitro test conducted pursuant to paragraph
(c)(6)(i)(A) of this section is negative.
(2) For the purposes of this section, the following provisions also
apply:
(i) Dose levels. For an initial assessment, one dose level of the
test substance may be used, the dose being the maximum tolerated dose
(to a maximum of 5,000 mg/kg), or that producing some indication of
cytotoxicity (e.g., partial inhibition of mitosis), or shall be the
highest dose attainable (to a maximum of 5,000 mg/kg) and should not
exceed the lower explosive limit of commercial hexane. Additional dose
levels may be used. For determination of dose-response, at least three
dose levels should be used.
(ii) Route of administration. Animals shall be exposed to commercial
hexane by inhalation.
(iii) Treatment schedule. The duration of exposure shall be for 6
hours per day for 5 consecutive days.
(C)(1) A dominant lethal assay shall be conducted with commercial
hexane in accordance with Sec. 798.5450 of this chapter except for the
provisions in paragraphs (d)(5) (ii) and (iii) of Sec. 798.5450, unless
both the in vitro and in vivo cytogenetics tests conducted pursuant to
paragraphs (c)(6)(i) (A) and (B) of this section are negative.
(2) For the purposes of this section, the following provisions also
apply:
(i) Dose levels. Normally, three dose levels shall be used. The
highest dose shall produce signs of toxicity (e.g., slightly reduced
fertility and slightly reduced body weight). The highest dose should not
exceed the lower explosive limit of commercial hexane. However, in an
initial assessment of dominant lethality, a single high dose may be
sufficient. Nontoxic substances shall be tested at 5 g/kg or, if this is
not practicable, then at the highest dose attainable.
[[Page 247]]
(ii) Route of administration. Animals shall be exposed to commercial
hexane by inhalation.
(iii) Treatment schedule. The duration of exposure shall be for 6
hours per day for 5 consecutive days.
(D)(1) A heritable translocation test shall be conducted with
commercial hexane in accordance with Sec. 798.5460 of this chapter
except for the provisions in paragraphs (d)(5) (ii) and (iii) of Sec.
798.5460, if the results of the dominant lethal assay conducted pursuant
to paragraph (c)(6)(i)(C) of this section are positive and if, after a
public program review, EPA issues a Federal Register notice or sends a
certified letter to the test sponsor specifying that the testing shall
be initiated.
(2) For the purposes of this section, the following provisions also
apply:
(i) Dose levels. At least two dose levels shall be used. The highest
dose level shall result in toxic effects (which shall not produce an
incidence of fatalities which would prevent a meaningful evaluation) or
shall be the highest dose attainable or 5 g/kg body weight and should
not exceed the lower explosive limit of commercial hexane.
(ii) Route of administration. Animals shall be exposed to commercial
hexane by inhalation.
(iii) Reporting requirements. (A) The chromosomal aberration tests
shall be completed and the final reports submitted to EPA as follows:
(1) The in vitro cytogenetics test within 15 months of the effective
date of the final rule.
(2) The in vivo cytogenetics test within 19 months of the effective
date of the final rule.
(3) The dominant lethal assay within 28 months of the effective date
of the final rule.
(4) The heritable translocation test within 25 months of the date of
EPA's notification of the test sponsor by certified letter or Federal
Register notice that testing shall be initiated.
(B) Interim progress reports for each test shall be submitted to EPA
for the in vivo cytogenetics and the dominant lethal assays at 6-month
intervals beginning 6 months after the effective date of the final rule,
until the applicable final report is submitted to EPA.
(C) Interim progress reports shall be submitted to EPA for the
heritable translocation assay at 6-month intervals beginning 6 months
after the date of EPA's notification of the test sponsor that testing
shall be initiated, until the final report is submitted to EPA.
(7) Neutrotoxicity--(i) Required testing. (A)(1) A schedule-
controlled operant behavior test shall be conducted with commercial
hexane in accordance with Sec. 798.6500 of this chapter except for the
provisions in paragraphs (d)(5)(i), (6) and (7) of Sec. 798.6500.
(2) For the purposes of this section, the following provisions also
apply:
(i) High dose level. The highest dose shall produce clear behavioral
effects or life-threatening toxicity. In addition, the highest dose
should not exceed the lower explosive limit of commercial hexane.
(ii) Duration and frequency of exposure. Animals shall be dosed once
for 4 to 6 hours.
(iii) Route of administration. Animals shall be exposed to
commercial hexane by inhalation.
(B)(1) A functional observation battery shall be conducted with
commercial hexane in accordance with Sec. 798.6050 of this chapter
except for the provisions in paragraphs (d)(4)(i), (5), and (6) of Sec.
798.6050.
(2) For the purposes of this section, the following provisions also
apply:
(i) High dose level. The highest dose shall produce clear behavioral
effects or life-threatening toxicity. In addition, the highest dose
should not exceed the lower explosive limit of commercial hexane.
(ii) Duration and frequency of exposure. Animals shall be dosed for
6 hours/day, 5 days/week for 90 days.
(iii) Route of exposure. Animals shall be exposed to commercial
hexane by inhalation.
(C)(1) A motor activity test shall be conducted with commercial
hexane in accordance with Sec. 798.6200 of this chapter except for the
provisions in paragraphs (d)(4)(i), (5), and (6) of Sec. 798.6200.
(2) For the purposes of this section, the following provisions also
apply:
(i) High dose level. The highest dose shall produce clear effects on
motor activity of life-threatening toxicity. In addition, the highest
dose should not
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exceed the lower explosive limit of commercial hexane.
(ii) Duration and frequency of exposure. Animals shall be dosed for
6 hours/day, 5 days/week for 90 days.
(iii) Route of exposure. Animals shall be exposed to commercial
hexane by inhalation.
(D)(1) A neuropathology test shall be conducted with commercial
hexane in accordance with Sec. 798.6400 of this chapter except for the
provisions in paragraphs (d)(4)(i), (5), and (6) of Sec. 798.6400.
(2) For the purposes of this section, the following provisions also
apply:
(i) High dose level. The highest dose shall produce clear behavior
effects or life-threatening toxicity. In addition, the highest dose
should not exceed the lower explosive limit of commercial hexane.
(ii) Duration and frequency of exposure. Animals shall be dosed for
6 hours/day, 5 days/week for 90 days.
(iii) Route of exposure. Animals shall be exposed to commercial
hexane by inhalation.
(ii) Reporting requirements. (A) The schedule-controlled operant
behavior, functional observation battery, motor activity, and
neuropathology tests shall be completed and the final reports submitted
to EPA within 15 months of the effective date of the final rule.
(B) Interim progress reports for each test shall be submitted to EPA
for the schedule-controlled operant behavior, functional observation
battery, motor activity, and neuropathology tests at 6-month intervals
beginning 6 months after the effective date of the applicable final
rule, until the applicable final report is submitted to EPA.
(8) Pharmacokinetics--(i) Required testing. (A) Pharmacokinetics
testing shall be conducted in rats in accordance with Sec. 795.232 of
this chapter, except for paragraph (c)(1)(ii) of Sec. 795.232.
(B) For the purposes of this section, the following provisions also
apply:
(1) Test animals. Adult male and female rats shall be used for
testing. The rats shall be 9 to 11 weeks old and their weight range
should be comparable from group to group. The animals shall be purchased
from a reputable dealer and shall be permanently identified upon
arrival. The animals shall be selected at random for the testing groups,
and any animal showing signs of ill health shall not be used.
(2) Species and strain. The rat strain used shall be the same as the
strain used in the subchronic and chronic tests required under Sec.
798.2450(d)(1)(i) and Sec. 798.3300(b)(1)(i).
(ii) Reporting requirements. (A) The inhalation and dermal
pharmacokinetics tests shall be completed and the final report submitted
to EPA by August 21, 1992.
(B) Interim progress reports shall be submitted to EPA for the
inhalation and dermal pharmacokinetics tests at 6-month intervals,
beginning 6 months after the effective date specified in paragraph
(d)(1) of this section, until the final report is submitted to EPA.
(d) Effective date. (1) The effective date of this final rule is
November 17, 1988, except for the provisions of paragraphs
(c)(2)(ii)(A), (c)(5)(i)(D), (c)(5)(ii)(A)(4), (c)(5)(ii)(C), (c)(8)(i)
and (c)(8)(ii)(A) of this section. The effective date for paragraphs
(c)(5)(i)(D), (c)(5)(ii)(A)(4) and (c)(5)(ii)(C) of this section is May
21, 1990. The effective date for paragraphs (c)(8)(i) and (c)(8)(ii)(A)
of this section is June 12, 1992. The effective date of paragraph
(c)(2)(ii)(A) is September 8, 1994.
(2) The guidelines and other test methods cited in this rule are
referenced as they exist on the effective date of the final rule.
[53 FR 3392, Feb. 5, 1988, as amended at 53 FR 38953, Oct. 4, 1988; 55
FR 634, Jan. 8, 1990; 55 FR 7325, Mar. 1, 1990; 55 FR 12643, Apr. 5,
1990; 57 FR 24961, June 12, 1992; 58 FR 34205, June 23, 1993; 59 FR
46357, Sept. 8, 1994; 60 FR 34467, July 3, 1995; 69 FR 18803, Apr. 9,
2004]