v>EPA
United States
Environmental Protection
Agency
Health Effects Research
Laboratory
Research Triangle Park NC 27711
Research and Development
EPA-600/S1 -81 -035 May 1981
Project Summary
Teratology and Postnatal
Studies in Rats of the
Propylene Glycol Butyl
Ether and Isooctyl Esters of
2,4-Dichlorophenoxyacetic Acid
Timothy M. linger, Janet Kliethermes, Dan Van Goethem, and
Robert D. Short
The purpose of this study was to
evaluate the teratogenic potential of
the propylene glycol butyl ether
(PGBE) and isooctyl (IO) esters of 2,4-
dichlorophenoxyacetic acid (2,4-D).
Accordingly, groups of pregnant CD
rats received daily oral doses of PGBE
or IO equivalent to 0, 6.25, 12.5,
25.0, or 87.5 mg/kg/day of 2,4-D
from day 6 through day 15 of gesta-
tion, and fetuses were observed for
gross, soft tissue, and skeletal
defects. In addition, a postnatal study
was performed on rats receiving 0,
12.5, or 87.5 ME/kg/day PGBE or IO
to determine the effect of treatment
on growth and survival of pups. No
adverse effects were observed on
maternal welfare, nor was there any
evidence of embryo or fetal lethality in
any of the treatment groups. Of the
anomalies observed, the incidence of
lumbar (14th) rib buds was found to be
statistically increased in the groups
given the 87.5 mg/kg/day doses of
both PGBE and IO. No other anomaly
reached a level of statistical or
toxicological significance.
The number of pups per litter was
significantly reduced on postpartum
days 4 and 7 in dams receiving 87.5
ME/kg/day IO. However, mean body
weight remained normal. Postnatal
growth and survival of pups receiving
PGBE were not adversely affected. It
was concluded that PGBE and IO
caused minor embryotoxicity which
was not deleterious to growth and
survival, and therefore, was not tera-
togenic to offspring of treated rats.
This Project Summary was develop-
ed by EPA's Health Effects Research
Laboratory, Research Triangle Park,
NC, to announce key findings of the
research project that is fully docu-
mented in a separate report of the
same title (see Project Report ordering
information at back).
Introduction
The herbicide, 2,4-dichlorophenoxy-
acetic acid (2,4-D) and several of its
ester derivatives are commonly used for
brush and weed control. The terato-
genic potential of these compounds has
been investigated in laboratory animals.
Fetal pathology and an increased inci-
dence of skeletal anomalies were
observed in Wistar rats that received
100 to 150 mg/kg of 2,4-D on days 6 to
15 of gestation. In addition, an
increased frequency of skeletal defects
was also observed in rats that received
butyl, isooctyl, butoxyethanol, and
dimetnylamine derivatives of 2,4-D.
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In another teratology study, doses of
2,4-D up to a maximum tolerated dose
of 87 5 mg/kg/day or molar equivalents
of 2,4-D isooctyl (10) esters or 2,4-D
propylene glycol butyl ether (PGBE)
esters were administered to Sprague-
Dawley rats on days 6 to 1 5 of gestation.
The only anomalies which could be
related to treatment were decreased
fetal body weight, subcutaneous
edema, delayed ossification of bone,
lumbar ribs, and wavy ribs. Since treat-
ment did not affect fetal or neonatal
development and survival, these obser-
vations were classified as signs of
embryotoxicity and fetotoxicity. Terato-
genic effects, which were defined as
embryotoxicity which seriously inter-
fered with normal development and
survival of the offspring, were not
observed.
The present study was undertaken to
reevaluate the teratogenic potential of
10 and PGBE. The doses of 10 and PGBE
were selected as the molar equivalents
of 0, 6.25, 12.5, 25, and 87.5 mg/kg of
2,4-D Accordingly, pregnant rats were
treated with 10 or PGBE on days 6 to 1 5
of gestation, and their fetuses were
examined for defects. In addition, dams
from some groups were allowed to
deliver, and the growth and develop-
ment of their pups were monitored.
Results
Pregnant rats were exposed to 0,
6.25, 12.5, 25.0, or 87.5 ME/kg/day of
10 or PGBE for 10 days starting on day 6
of gestation. Mortality was not observed
in rats receiving up to 87.5 ME/kg/day
IO or PGBE. While rats which received
25 0 ME/kg/day had significantly
reduced body weights during gestation,
this was determined not to be treatment-
related since this effect occurred before
treatment began. Further, all other
dams gained weight normally during
gestation.
Embryo or fetal lethality, as measured
by percent viable fetuses, percent early
resorptions, number of dams with com-
plete resorptions, and fetal weight, was
not observed with IO or PGBE.
None of the gross anomalies observed
in the rats were increased to a statisti-
cally significant level by treatment with
either PGBE or IO.
Soft tissue anomalies observed in rats
treated with either PGBE or IO were not
increased to a significant level. How-
ever, hydronephrosis, both marked and
slight, was observed only in the treated
animals. This anomaly, though, occur-
red at very low incidences in all dose
levels in rats treated with both PGBE
and IO, and showed no dose-related
response.
Skeletal anomalies observed in rats
treated with PGBE or IO consisted
mainly of minor variations in the degree
of ossification There was a significantly
increased number of fetuses with
lumbar (14th) rib buds in the 875
ME/kg/day dose levels of both IO and
PGBE. Additionally, in one pup treated
with 87.5 ME/kg/day of IO some major
bone malformations of the axial
skeleton, including fused ribs, fused
centri, unossified vertebrae, and fusion
of vertebrae, were observed.
These scattered effects are consid-
ered to be within normal biological
variation. When the significance level is
set up at p <0.05, it is expected that one
anomaly in twenty in each dose group
will be significantly different by random
variation The differences observed
here were not dose-related and did not
include clusters of related anomalies
Therefore, these differences have been
considered lexicologically unimportant
In addition, a postnatal growth and
survival study was performed on rats
receiving 0, 12.5, or 87.5 ME/kg/day
PGBE or IO Pup body weights for both
compounds were normal. In rats
receiving 87.5 ME/kg/day IO, there
was a reduced number of pups per litter
on days 4 and 7 postpartum. This effect,
however, represents a whole litter loss
by one of the dams at this dose level
Since the survival of pups from other
dams in this treatment group was
similar to the control, it was concluded
that this effect was not treatment-
related.
In a previous study, PGBE and IO were
given orally to rats in similar doses on
days 6 and 15 of gestation, and an in-
creased incidence of fetuses with
subcutaneous edema was observed.
This anomaly was not observed in the
present study in any of the dose levels
studied. However, the same study
reported a significant increase in the
incidence of lumbar ribs in the 87.5
ME/kg/day dose levels of both IO and
PGBE. An increase in the appearance of
lumbar ribs was also reported in
another study using Wistar rats at only
slightly higher maximum doses of PGBE
andIO.
Although the occurrence of lumbar
ribs has no morphological or functional
consequence to the fetuses and appears
spontaneously in control animals, it
falls within the scope of embryotoxicit
Since this effect is not detrimental to tr
normal development and survival <
fetuses, it is not classified as a "terati
genie" effect.
In summary, the results of this stuc
indicate that (a) PGBE or IO produce r
adverse effects on maternal welfare <
pup viability, (b) PGBE or IO exhib
embryotoxicity of insignificant detr
mental effect; and (c) PGBE or IO do n<
adversely affect postnatal growth an
survival of pups.
Accordingly, these results indicat
that PGBE and IO are not teratogenic i
CDŽ rats
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Timothy M. Linger, Janet Kliethermes, Dan Van Goethem, and Robert D Short
are with Midwest Research Institute, Kansas City, MO 64110.
William F. Durham is the EPA Project Officer (see below).
The complete report, entitled "Teratology and Postnatal Studies in fiats of the
Propy/ene Glycol Butyl Ether andIsooctylEsters of 2,4-Dichlorophenoxy acetic
Acid," (Order No. PB 81-191 140; Cost $6.50, subject to change) will be
available only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA 22161
Telephone: 703-487-4650
The EPA Project Officer can be contacted at:
Health Effects Research Laboratory
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
i US GOVERNMENT PRINTING OFFICE 1961 -757-012/7123
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United States
Environmental Protection
Agency
Center for Environmental Research
Information
Cincinnati OH 45268
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