Journal of Ethnopharmacology 67 (1999) 253 – 258

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Post-coital antifertility activity of Acalypha indica L.

Shivayogi P. Hiremath a,*, K. Rudresh a, Shrishailappa Badami a,
Saraswati B. Patil b, Somanath R. Patil b
a
Department of Chemistry, Gulbarga Uni6ersity, Gulbarga-585 106, India
b
Department of Zoology, Gulbarga Uni6ersity, Gulbarga-585 106, India

Received 1 August 1998; received in revised form 19 October 1998; accepted 2 November 1998

Abstract

Four successive solvent extracts of the whole plant Acalypha indica L. (Euphorbiaceae) were tested for post-coital
antifertility activity in female albino rats. Of these, the petroleum ether and ethanol extracts were found to be most
effective in causing significant anti-implantation activity. The antifertility activity was reversible on withdrawal of the
treatment of the extracts. Both the extracts at 600 mg/kg body weight showed estrogenic activity. Histological studies
of the uterus were carried out to confirm this estrogenic activity. © 1999 Elsevier Science Ireland Ltd. All rights
reserved.

1. Introduction 1935; Anonymous, 1948; Chopra et al., 1956;

One approach pursued to identify new antifer-
tility agents is the search for their presence in
natural sources. Many plant preparations are re-
ported to have fertility regulating properties and a
few have been tested for such effects. But so far
no single plant is available which can be devel-
oped further as a potent antifertility agent. Hence
the search continues. Acalypha indica L. (Euphor-
biaceae) is a weed widely distributed throughout
the plains of India. It has been reported to be
useful in treating pneumonia, asthma, rheumatism
and several other ailments (Kirtikar and Basu,
* Corresponding author.
Nadakarni and Nadakarni, 1982). This plant was
cited as an emmenagogue by Nadakarni and
Nadakarni (1982). The leaves of Acalypha grandis
have also been reported to possess contraceptive
activity (Bourdy and Walter, 1992). Several chem-
ical (Donw and Steyn, 1938; Talapatra et al.,
1981; Nohrstedt et al., 1982; Manzoor-i-Khuda et
al., 1985; Asolkar et al., 1992) and biological
(Caius and Mhaskar, 1923; Hiremath et al., 1993)
investigations have been carried out on this plant,
but, so far no antifertility testing has been done.
Hence, in continuation of our work on antifertil-
ity activity of plants (Hiremath and Hanumantha,
1990; Hiremath et al., 1990, 1994, 1996), we were
interested in subjecting Acalypha indica L. to an-
tifertility testing in female albino rats.

0378-8741/99/$ - see front matter © 1999 Elsevier Science Ireland Ltd. All rights reserved.
PII: S 0 3 7 8 - 8 7 4 1 ( 9 8 ) 0 0 2 1 3 - X
254 S.P. Hiremath et al. / Journal of Ethnopharmacology 67 (1999) 253–258
2. Materials and methods
The whole plant of Acalypha indica L. was
collected from the fields in and around Gulbarga
during August and September, 1996 and was au-
thenticated at the Herbarium, Department of
Botany, Gulbarga University, Gulbarga (HGUG
No.279). The plant was shade dried, powdered
and subjected to Soxhlet extraction (500 g) succes-
sively and separately with petroleum ether (60 –
80°C, 2 l), chloroform (2 l), ethanol (95%, 2 l) and
distilled water (2 l). The extracts were concen-
trated to dryness in a flash-evaporator under re-
duced pressure and controlled temperature
(50 – 60°C). The petroleum ether and chloroform
extracts yielded dark brown solids weighing 20.0
and 32.0 g, respectively. All the extracts were
stored in a refrigerator. The extracts were pre-
pared in Tween − 80 (1%), suspended in distilled
water and they were administered to the rats at
doses of 300 and 600 mg/kg body weight orally by
means of an intragastric catheter.
Colony-bred female albino rats (Wistar strain),
weighing (150–200 g), were maintained under
controlled standard animal house conditions with
access to food and water ad libitum. Vaginal
smears from each rat were monitored daily. Only
rats with normal estrous cycles were selected.
Anti-implantation activity was determined as de-
scribed by Khanna and Chaudhury (1968). Rats
found in proestrous phase of the cycle were caged
with males of proven fertility, in the ratio of 2:1
and examined the following morning for evidence
of copulation. Rats exhibiting thick clumps of
spermatozoa in their vaginal smears were sepa-
rated and that day was designated as day 1 of
pregnancy and those rats were divided into nine
groups containing eight rats in each group. The
extracts were administered at 300 and 600 mg/kg
body weight orally from days 1 to 7 of pregnancy.
Control rats received the vehicle (Tween-80, 1%)
only. On day 10, laparotomy was performed un-
der light ether anesthesia and semisterile condi-
tions. The uteri were examined to determine the
number of implantation sites. The rats were al-
lowed to recover and deliver after full term. Each
pup was weighed and examined for gross defects.
The litters were allowed to grow to check post-na-
tal growth and monitor any congenital
abnormalities.
The petroleum ether and ethanol extracts were
found to be the most active of the extracts of
Acalypha indica L., hence they were subjected to a
detailed investigation for potential estrogenic and
antiestrogenic activity. Colony-bred (Wistar
strain) immature female albino rats, 21–23 days
old, weighing between 35 and 45 g, were bilater-
ally ovariectimised by dorsolateral approach un-
der light ether anaesthesia and sterile conditions.
They were divided into six groups consisting of
eight rats each. The first group served as a control
end received vehicle only (Tween-80, 1%). The
second group received ethinyl estradiol in olive
oil, 1 mg/rat per day, subcutaneously. The third
and fourth groups received the petroleum ether
and ethanol extracts at a dose of 600 mg/kg body
weight, respectively. The fifth and sixth groups
received, in addition to ethinyl estradiol, a test
dose of the petroleum ether and ethanol extracts
at 600 mg/kg body weight, respectively. All the
above treatments were given for 7 days. On the
8th day, the rats were sacrificed by decapitation,
the uteri dissected out and surrounding tissues
removed. The uteri were blotted on filter papers
and weighed quickly on a sensitive balance and
fixed in Bouin’s fluid for 24 h. The tissues were
dehydrated and embedded in paraffin. The
paraffin sections were cut at 6 mm and stained
with haematoxylin–eosin for histological observa-
tions. The diameter of the uterus, thickness of the
endometrium and the height of the endometrial
epithelium were measured in 20 randomly selected
sections using an ocular micrometer. Statistical
analysis was carried out using Student’s t-test.
The results were judged significant if PB 0.05.
3. Results
Of the four extracts of Acalypha indica L. eval-
uated for post-coital antifertility activity, the
petroleum ether extract at 300 and 600 mg/kg
significantly inhibited pregnancy in three of eight
rats with a mean number of implants of 5.8759
1.76 (PB 0.05) and six of eight rats with a mean
number of implants of 2.1259 1.39 (PB 0.001),
 S.P. Hiremath et al. / Journal of Ethnopharmacology 67 (1999) 253–258 255

respectively (Table 1). The ethanol extract at
doses of 300 and 600 mg/kg body weight also
showed anti-implantation activity in three of eight
and five of eight rats, with a mean number of
implants of 6.00 91.76 (P B 0.05) and 3.25 9 1.60
(P B0.001), respectively. However, both the doses
of the chloroform and distilled water extracts
were found to be ineffective and the number of
implantation sites in these cases were comparable
with the control rats.
No toxic effects were observed either by gross
visual examination or in the weight of animals.
After discontinuation of treatment, all the animals
were mated. This resulted in pregnancy and deliv-
ery of normal litters, indicating that the action of
the extracts was reversible.
The estrogenic and the antiestrogenic activity of
the petroleum ether and ethanol extracts is shown
in Tables 2 and 3. Oral administration of the
petroleum ether and ethanol extracts at 600 mg/
kg body weight caused a significant increase in
uterine weight in immature rats (versus control,
P B 0.001). The uterotrophic potency, as shown
by the weight of the uterus, is about 37% of that
of the ethinyl estradiol in the case of the
petroleum ether and 32% of that of ethinyl estra-
diol in the case of the ethanol extract, respec-
tively. The uterotrophic changes, such as the
diameter of the uterus, (P B 0.001), thickness of
the endometrium (PB 0.001) and height of the
endometrial epithelium (P B 0.05), were signifi-
cantly increased when compared with control rats.
The uteri of these rats were inflated and full of
fluid resembling the proestrous/estrous uterus.
The epithelium of the endometrium consisted of
spindle-shaped cells with basal nuclei. The stroma
consisted of loose and edematous fibroblast-type
cells. The treated rats showed open vaginas, while
all the control rats had closed vaginas. Examina-
tion of the vaginal smears of treated rats revealed
predominantly cornified and nucleated epithelial
cells. However, their number was less than in
ethinyl estradiol-treated rats.
It appears that the petroleum ether and ethanol
extracts have weak estrogenic activity, but no
antiestrogenic activity at 600 mg/kg dose.
4. Discussion
In the present study, Acalypha indica L. was
tested for its anti-implantation and estrogenic
properties. Among the four extracts tested at two
different doses, the petroleum ether and ethanol
extracts at 600 mg/kg body weight dose were
more potent in their anti-implantation activity, as

Table 1
Effect of extracts of Acalypha indica L. on implantation in rats when fed orally from days 1 to 7 of pregnancya

Treatment Dose (mg/kg No. of rats having no Mean number of % of rats having no
body weight) implantation sites on day 10 implants 9 S.E implantation sites on day 10

Control – Nil 11.000 9 0.46 Nil

Petroleum ether 300 3 05.875 9 1.76* 37.5
extract 600 6 02.125 9 1.39** 75.0
Chloroform 300 Nil 10.500 90.29 Nil
extract 600 Nil 11.625 90.37 Nil
Ethanol extract 300 3 06.000 91.76* 37.5
600 5 03.2509 1.60** 62.5
Distilled water 300 Nil 10.0009 0.46 Nil
extract 600 Nil 08.375 9 0.70 Nil

a
Each group consisted of eight rats.
* PB0.05; when compared with control.
** PB0.001; when compared with control.
256 S.P. Hiremath et al. / Journal of Ethnopharmacology 67 (1999) 253–258
Table 2
Estrogenic and anti-estrogenic activity of the petroleum ether and ethanol extracts of Acalypha indica L.
Group Treatment (dose)
I Control
II Ethinyl estradiol (1 mg/rat per day)
III Petroleum ether extract (600 mg/kg)
IV Ethanol extract (600 mg/kg)
V Ethinyl estradiol (1 mg/rat per day)+petroleum ether
extract (600 mg/kg)
VI Ethinyl estradiol (1 mg/rat per day)+ethanol extract
(600 mg/kg)
a
+, nucleated epithelial cells; ++, nucleated and cornified cells; +++, cornified cells.
* PB0.001; when compared with control.
** PB0.01; when compared with ethinyl estradiol.
75 and 62.5% of the rats failed to show any
implantation sites, respectively. However, the
chloroform and distilled water extracts were inac-
tive, as the number of implantation sites in these
cases were comparable with the control rats.
The loss of implantation caused by the
petroleum ether and ethanol extracts may be due
to antizygotic, blastocytotoxic or anti-implanta-
tion activity as described by Hafez (1970).
The petroleum ether and the ethanol extracts
also exhibited estrogenic activity as shown by the
significant increase in uterine weight, diameter of
the uterus, thickness of endometrium, height of
the endometrial epithelium and vaginal epithelial
cornification in immature rats.
It is well known that for implantation exact
equilibrium of estrogen and progesterone is essen-
tial, and any disturbance in the level of these
hormones may cause infertility (Psychoyos, 1966).
The compound of hormonal values usually dis-
turbs the hormonal milieu in the uterus and pro-
vokes an infertility effect. In this study, the
histological evidence of the uterus treated with
petroleum ether and ethanol extracts clearly sup-
ports an unfavourable uterine milieu. Therefore,
the anti-implantation activity may be due to es-
trogenic activity, causing the expulsion of ova
from the tube, disrupting the luteotrophic activity
of the blastocyst (Pincus, 1965; Anderson, 1972).
Uterine weight (mg/100 g body weight; Vaginal
mean9S.E.) cornification
071.98 902.64 Nil
330.34 9 17.59* +++a
123.799 07.28* + to ++a
108.52 909.94* + to ++a
258.13 9 05.44 *,**
+++a
402.62 9 12.81*,** +++a
It is interesting to note that they possess around
35% of the estrogenic efficacy of ethinyl estradiol
and thus may reduce some of the unwanted side
effects of estrogens.
Simultaneous administration of ethinyl estra-
diol and petroleum ether extract caused a highly
significant increase in the uterine weight when
compared with control (P B0.001). But, the de-
gree of uterotrophic potency was less than that
produced by ethinyl estradiol (P B 0.01), when
compared with standard ethinyl estradiol. It also
caused a highly significant increase in uterine di-
ameter, thickness of the endometrium end height
of the endometrial epithelium (versus control,
PB 0.001). The simultaneous administration of
ethinyl estradiol and the ethanol extract also
caused a highly significant increase in the uterine
weight (versus control, PB 0.001). However, the
extent of uterotrophic response was greater than
that produced by ethinyl estradiol alone (P B
0.01). These observations have also been confi-
rmed when the uterotrophic changes, such as the
diameter of the uterus, thickness of the en-
dometrium and height of the endometrial epithe-
lium, were compared with the control and the
standard ethinyl estradiol treatments.
It appears that the petroleum ether and the
ethanol extracts have weak estrogenic activity
when given alone, but the petroleum ether extract
 S.P. Hiremath et al. / Journal of Ethnopharmacology 67 (1999) 253–258 257

Table 3
Histological changes in the uterus and endometrium after treatment with petroleum ether and ethanol extract of Acalypha indica L.

Treatment (dose) Diameter of uterus Thickness of Height of endometrial

(mm9 S.E.) endometrium (mm9S.E.) epithelium (mm9 S.E.)

Control 338.999 3.30 52.35 9 2.10 19.59 92.25

Ethinyl estradiol (1 mg/rat per day) 812.519 4.27** 240.71 92.19** 46.03 9 2.60*
Petroleum ether extract (600 mg/kg) 625.069 1.44** 85.06 9 2.23** 32.5291.19*
Ethanol extract (600 mg/kg) 512.069 2.15** 78.39 9 1.59** 29.58 91.01*
Ethinyl estradiol (1 mg/rat per day)+ 919.899 3.00 **,****
288.44 95.41 **,****
54.38 91.86**,***
petroleum ether extract (600 mg/kg)
Ethinyl estradiol (1 mg/rat per day)+ethanol 859.369 1.12**,**** 249.10 91.08**,**** 50.35 90.69**,***
extract (600 mg/kg)

* PB0.05; when compared with control.

** PB0.001; when compared with control.
*** PB0.05; when compared with ethinyl estradiol.
**** PB0.001; when compared with ethinyl estradiol.

has shown anti-estrogenic activity when given
along with standard ethinyl estradiol. However, the
ethanol extract did not show any anti-estrogenic
activity when given along with ethinyl estradiol at
the tested dose.
The phytochemical studies reported on Acalypha
indica L. revealed the presence of several sterols in
the petroleum ether extract (Talapatra et al., 1981).
A flavone kaempferol (Asolkar et al., 1992) has also
been reported from the ethanolic extract. In our
phytochemical studies of this plant two flavonoids,
chrysin and galangin (Hiremath et al., 1998), have
been isolated from the ethanol extract. The prelim-
inary investigations of the antifertility studies re-
vealed that the two newly isolated flavonoids have
promising antifertility activity. Several sterols (Hall
and Fraser, 1983) and flavonoids (Pincus, 1965;
Psychoyos, 1966; Khanna and Chaudhury, 1968;
Hafez, 1970; Anderson, 1972) have been reported
to possess antifertility activity. Therefore, the anti-
implantation activity of the extracts of Acalypha
indica L. might be due to the presence of such
compounds.
Acknowledgements
The authors are thankful to Professor Malcolm
Hooper, University of Sunderland, Sunderland,
UK for useful discussions, and to M/s. Wyeth
Laboratories Ltd., Bombay, for supplying a free
sample of ethinyl estradiol. One of the authors
(KR) is thankful to AICTE, New Delhi, for finan-
cial assistance.
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