International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 3, Issue 1, 2011

ResearchArticle

ANTIDEPRESSANTACTIVITYOFMETHANOLICEXTRACTOFPASSIFLORAFOETIDALEAVES
INMICE

SANTOSHP,VENUGOPLR,NILAKASHAS,KUNJBIHARIS,DR.MANGALAL
NationalInstituteofPharmaceuticalEducationandResearch(NIPER),Guwahati,Assam,IndiaEmail:dr_mlahkar@rediffmail.com
Received:03Oct2010,RevisedandAccepted:05Nov2010

ABSTRACT
Passiflora foetida (Passifloraceae), popularly known as stinking passion flower, is an herbaceous climber that has been widely used in Mexican
traditionalmedicineforthetreatmentofdifferentcentralnervoussystem(CNS)disorders.Nevertheless,theavailablescientificinformationabout
thisspeciesisscarceandtherearenoreportsrelatedtoitspossibleeffectontheCNS.Inthiswork,theeffectsofmethanolicextractofleavesofP.
foetida (PF) were evaluated in mice using behavioral tests sensitive to clinically effective antidepressant compounds. The extract (100,200 and
300mg/kg), administered intraperitoneally, was able to decrease the immobility time of mice dosedependently when subjected to both tail
suspensionandforcedswimtestsandtheeffectsarecomparabletothatofstandarddrugsi.e.,fluoxetine(20mg/kg)andimipramine(15mg/kg).
Neither the extracts of PF nor fluoxetine, at the doses tested, produced significant effects on locomotor activity when subjected to open field
behavioraltest.TheseresultsdemonstratedthatPFhadspecificallyantidepressanteffectsinvivo.Inconclusion,thepresentstudysuggestedthat
PFextractspossessedpotentialantidepressanteffectswhichcouldbeoftherapeuticinterestforusinginthetreatmentofpatientswithdepressive
disorders.
Keywords:Passiflorafoetida,antidepressantactivity,tailsuspensiontest,forcedswimtest,openfieldtest.

INTRODUCTION Hyderabad. The botanical nomenclature of the plants was duly

According to the World Health report1, approximately 450 million
peoplesufferfromamentalorbehavioraldisorder.Thisamountsto
12.3%oftheglobalburdenofdisease,andwillriseto15%by20202.
Psychiatricillnessisalsooftenassociatedwithsuicideandthereare
between10and20millionsuicideattemptseveryyear.Depression
isthemostprevalentmentaldisorderanddepressionisrecognized
to be symptomatically, psychologically and biologically
heterogeneous3.The disorder was characterized by apathy, loss of
energy, retardation of thinking and activity, as well as profound
feelingsofgloominess,despairandsuicidalideation.Inspiteofthe
availability of antidepressant drugs like tricyclic antidepressants,
selective reversible inhibitors of monoamine oxidaseA (MAOA),
selective serotonin reuptake inhibitors (SSRIs) and selective
noradrenaline reuptake inhibitors (SNRIs), depression continue to
beamajormedicalproblem4.Basicneuroscienceoffersthepromise
of improving our understanding of disease pathophysiology,
identifyingnovelmechanismsthatcanbetargetedbymoreeffective
pharmacotherapiesandscreeningofherbalsourcesofdrugs.These
considerations implicate the search for new antidepressant agents
that have a fast onset of action, with less side effects and a wider
safetymargin.Variousplantsarebeingusedincomplementaryand
alternativemedicinesformanagementofmooddisorders.
identified by using standard floras and also crosschecked with
Herbariumrecords.Theplantmaterialwasshadedriedfor10days
andpulverized.
Preparationofextract
The dried plant material was crushed into fine particles (powder)
usingamixer.Thepowderedplantmaterial(500g)waspackedina
soxhlet apparatus and subjected to continuous hot percolation for
8h using methanol (1:4) as solvent. Extract obtained was passed
through the Whatman filter paper No.1 and the methanol was
evaporated(at400C)withthehelpofheatingmantleanddriedina
desiccator.
Animals
MaleSwissalbino miceweighing between 20 25 gwere used for
the present study. The animals were maintained under standard
environmentalconditions(25+2Candrelativehumidityof45to
55%) andwere fed with standardpellet diet and waterad libitum.
The study was approved by Institutional Animal Ethics Committee
(01/NIPER/CPCSEA/351). CPCSEA guidelines were adhered during
themaintenanceandexperiment.Allexperiments werecarried out
between10:0017:00hours.

Onthebasisoftheaboveinformation,theleavesofPFwasselected
for evaluating its anxiolytic and antidepressant activity due to its
traditional use in the management of anxiety, stress, insomnia,
hysteria,skininflammation,coughandfever.Chemicalconstituents
in PF include hydrocyanic acid, groups of flavonoids and harman
alkaloids5. Some reports have pointed out the harman alkaloids as
the bioactive constituents of Passiflora incarnata Linn, one of the
species of Passiflora that have been extensively studied chemically
and biologically6,7. Harman alkaloids were also found to be present
inPF.UnlikelyP.incarnata;sofartherehasbeennoscientificreport
in literature about the antidepressant activity (in experimental
animal models) of this species. Therefore, the present study has
been undertaken to investigate the effect of methanolic extract of
passiflorafoetidaLinn.ondepressioninmice.
Drugs&chemicals
Fluoxetine hydrochloride (FLUDAC, Cadila Pharmaceuticals,
Ahmedabad, India) and imipramine hydrochloride (SigmaAldrich,
St. Louis, USA) were used reference standards for antidepressant
activity.
Experimentalprotocols
Overnight fasted animals were selected randomly on the day of
experiment for administration of vehicle, standard drug and study
drug.Theanimalswereacclimatizedonehourbeforeforbehavioral
tests. Thirty minutes and 1 hour time interval between drug
administration and behavioral tests were maintained in case of
intraperitonealandoraladministrationsrespectively.

MATERIALS&METHODS
Plantmaterial
The leaves of P.foetida L. were collected from botanical garden in
madhapur, Hyderabad, Andhrapradesh. The plant material was
identified and authenticated by the botanist, Botanical garden,
The animals were divided into five groups of six animals each as
follows:
GroupI(n=6)Control,receiveddistilledwater,i.p
GroupII(n=6)(Standard)Imipramine(forcedswimtest)15mg/kg
& fluoxetine (tail suspension test) 20 mg/kg, p.o; Diazepam
(1mg/kg,i.p)inopenfieldtest.
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GroupIII(n=6)MEPF100mg/kg,i.p which was significant (135.333.19, 113.172.81 and 96.172.45

GroupIV(n=6)MEPF200mg/kg,i.p
GroupV(n=6)MEPF300mg/kg,i.p
The antidepressant activity was carried out using two different
models.Furthertheeffectofdrugswasevaluatedinopenfieldtest.
respectively;p<0.001)when comparedwithcontrol(160.173.62).
Similarly, animals treated with fluoxetine (20 mg/kg), as expected,
showed a significant decrease in the immobility time (73.332.11;
p<0.001).

Behavioraltests
Tailsuspensiontest(TST)
Tail suspension test commonly employed behavioral model for
screening antidepressantlike activity in mice, was first given by
Steru .,et.al. Animals were moved from their housing colony to
laboratoryintheirowncagesandallowedtoadapttothelaboratory
conditionsfor12hr.Eachmousewasindividuallysuspendedtothe
edge of a table, 50 cm above the floor, by adhesive tape placed
approximately1cmfromthetipofthetail.Eachanimalunder test
was both acoustically and visually isolated from other animals
during the test. The total period of immobility was recorded
manuallyfor6min.Animalwasconsideredtobeimmobilewhenit
didnt show any body movement, hung passively and completely
motionless.Thetestwasconductedinadimlightedroomandeach
mouse was used only once in the test. The observer, recording the
immobilityofanimals,wasblindtothedrugtreatmentsgiventothe
animalsunderstudy8,9. Fig.1:EffectsofMEPFanddiazepamondurationofimmobility
intheTST.ResultsareexpressedasmeanS.E.M(n=6).***P<
Forcedswimtest(FST) 0.001ascomparedtorespectivecontrolgroup.
Forced swim test, the most frequently used behavioral model for

screeningantidepressantlikeactivityinrodents,wasfirstproposed
by Porsolt., et. al. The procedure was same as followed previously.
Micewereindividuallyforcedtoswiminopenglasschamber(25
15 25cm) containing fresh water to a height of 15 cm and
maintained at 261C. At this height of water, animals were not
abletosupportthemselvesbytouchingthebottomorthesidewalls
of the chamber with their hindpaws or tail. Water in the chamber
was changed after subjecting each animal to FST because used
water has been shown to alter the behavior. Each animal showed
vigorous movement during initial 2 min period of the test. The
durationofimmobilitywasmanuallyrecordedduringthenext4min
Forcedswimtest(FST)
The possible antidepressant effect of MEPF after intraperitoneal
administrationwasstudiedintheforcedswimmingtest.Inthistest
(Fig.2),animalstreatedwiththreedosesofMEPF(100,200and300
mg/kg,i.p) showed decreasesin theirimmobility times, which was
significant (82.832.87, 60.671.98 and 47.672.46 respectively;
p<0.001) when compared with control (150.174.9). Similarly,
animalstreatedwithimipramine(15mg/kg),asexpected,showeda
significantdecreaseintheimmobilitytime(38.52.92;p<0.001).

ofthetotal6mintestingperiod.
Mice were considered to be immobile when they ceased struggling
and remained floating motionless in water, making only those
movements necessary to keep their head above water. Following
swimming session, mice were towel dried and returned to their
housingconditions8,9.
OpenField
This test utilizes behavioral changes in rodents exposed to novel
environments and is used to confirm that the observed
antidepressant effect is not due to stimulation of general motor
activity.VarioustypesofOpenfieldapparatushavebeenusedtotest
themice.Theopenfieldtestwascarriedoutonthedarkgrey floor
subdividedinto16equalpartsinawoodenbox(100cmx100cmx
30 cm).Respectivetreatmentwasgiventotheanimalsand 30min
later, the animals were individually placed in the corner square of
theopenfield.Thefollowingparameterswereobservedfor5min10: Fig.2:EffectsofMEPFanddiazepamondurationofimmobility
Activityinthecentre(numberofcentralsquarescrossed) intheFST.ResultsareexpressedasmeanS.E.M(n=6).***P<
Spontaneous ambulation (number of squares crossed at 0.001ascomparedtorespectivecontrolgroup.
periphery) [

Rearing(No.oftimestheanimalstandsontherearpaws).
STATISTICALANALYSIS
Allthe datarepresent meanS.E.M.values. The datawereanalyzed
by means of analysis of variance (ANOVA). Whenever ANOVA was
significant, further multiple comparisons were made using Tukeys
testastheposthoctest.AllanalyseswereperformedusingtheSPSS
statisticalsoftware.Thelevelsofstatisticalsignificancerangedfrom
p<0.05top<0.001.
RESULTS
Tailsuspensiontest(TST)
Inthistest(Fig.1),animalstreatedwiththreedosesofMEPF(100,
200and300mg/kg,i.p)showeddecreasesintheirimmobilitytimes,
Openfieldtest
Thoughtherewasslightincreaseinthenumberofsquarescrossed
(peripheral) by mice in MEPF treated groups (100,200 and
300mg/kg, i.p.) as compared to control, it was not statistically
significant(Fig.3&4).ThenumberofcentralSquarescrossedinthe
control and standard (Diazepam) group were 10.17 1.33 and
31.333.67secrespectively.Therewasasignificantincreaseinno.
of crossings in diazepam group as compared to control group. But
whendifferentdosesofMEPFwereusedalonetheincreaseinno.of
centralsquarecrossingswasnot statisticallysignificant.Therewas
significant increase in the rearing of animals with diazepam in
comparisontothecontrolgroup(p<0.01).Therewasalsoincreased
number of rearing in test drug treated groups which was not
statisticallysignificant.

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Table1:EffectsofdiazepamandMEPFonbehaviourofmiceinOpenfieldparadigm
No.ofsquarescrossed(MeanSEM) No.ofRearings
Treatments N (MeanSEM) Total
Centre periphery
Control 6 10.171.33 81.5010.15 91.6710.03 3.670.49
Diazepam(1mg/kg) 6 31.333.67*** 109.836.79ns 141.179.19** 7.170.65**
MEPF(100mg/kg) 6 15.002.08ns 88.177.16ns 103.178.55ns 4.000.58ns
MEPF(200mg/kg) 6 19.832.3* 90.174.13ns 1104.77ns 5.330.49ns
MEPF(300mg/kg) 6 20.501.31* 98.334.58ns 118.834.38ns 6.000.73ns
ValuesareexpressedasmeanSEM.ns=notsignificant,*P<0.05,**P<0.01,***P<0.001,comparedtorespectivevehicletreatedcontrolgroup.

DISCUSSION&CONCLUSION regulates the metabolic degradation of catecholamines, serotonin

The incidence of anxiety and depression in the community is very
high and is associated with lot of morbidity. Hence, it is very
important to address these problems and find effective remedies.
Though several drugs are available, all are associated with some
limitations and there is an urgent need for alternative medications
for these disorders. Despite the widely popular use of Passiflora
foetida for treating nervous disorders, there is an absence of
scientificreportsabouttheevaluationofitspharmacologicaleffects.
In this work, it was demonstrated that the administration of
different doses of the methanolic extract of P.foetida in mice was
abletoinduceantidepressanteffects.
On the basis of the clinical association of depressive episodes and
stressfullifeevents,manyoftheanimalmodelsfortheevaluationof
antidepressant drug activity assess stressprecipitated behaviors.
The two most widely used animal models for antidepressant
screeningaretheforcedswimmingandtailsuspensiontests.These
testsarequitesensitiveandrelativelyspecifictoallmajorclassesof
antidepressants11. In TST, immobility reflects a state of despair
which can be reduced by several agents which are therapeutically
effectiveinhumandepression.SimilarlyintheFST,miceareforced
to swim in restricted space from which they cannot escape. This
inducesastateofbehavioraldespairinanimals,whichisclaimedto
reproduce a condition similar to human depression12. It has been
seen that the TST is less stressful and has higher pharmacological
sensitivitythanFST13.
Results showed that the administration of the MEPF produced a
diminutionofimmobilitytime(aposturethoughttoreflectastateof
behavior despair in which animals have given up the hope to
escape) of mice exposed to the both forced swimming and tail
suspensiontests.Inthepresentstudy,methanolicextract(100,200
and 300 mg/kg, po) administered to mice, produced significant
antidepressantlike effect in both TST and FST and their efficacies
were found to be comparable to imipramine(15 mg/kg, po) and
fluoxetine(20mg/kg,po).
Data in the literature demonstrated that drugs that alter general
motoractivitymaygivefalsepositive/negativeresultsintheforced
swimmingtest.TheeffectsproducedbyMEPFandDZP(1.0mg/kg)
upon the open field test demonstrated that these products do not
modifythespontaneouslocomotoractivityofmice,whichindicates
that the plant extract exerts antidepressant effects without
modifyingsignificantlythisparameter.Therefore,itisprobablethat
these effects are not related to the stimulation of general motor
activity14.
Ithasbeenestablishedthattheshorteningofimmobilitytimeinthe
forced swimming and the tail suspension tests depends mainly on
the enhancement of central 5HT and catecholamine
neurotransmission15. Early evidence of a role for noradrenaline in
depression came from the discovery that drugs, either causing or
alleviatingdepression,actedtoalterthenoradrenalinemetabolism.
Furthermore,depletionstudiescarriedoutintreatedanduntreated
patients indicated a role for serotonin and noradrenaline in
depression16.
Harmaline alkaloids present in P.foetida act as reversible
monoamine oxidase inhibitors17 and in common with other beta
carboline binds to 5hydroxy tryptamine (HT) receptors18. MAO
andotherendogenousaminesincentralnervoussystem.Inhibition
of this enzyme causes a reduction in metabolism and subsequent
increaseintheconcentrationofbiogenicamines.Alsotheflavonoid
components of MEPF might be interacting with adrenergic and
serotonergic systems in mediating the antidepressant effects of
MEPF. However, the precise mechanisms by which the extract
producedantidepressantlikeeffectarenot completelyunderstood.
Further studies would be necessary to evaluate the contribution of
activechemicalconstituentsfortheobservedantidepressantactivity
as it still remains to be determined which components were
responsiblefortheseeffects.
ACKNOWLEDGEMENT
TheauthorsaregratefultoDr.(Mrs.)C.C.Barua,VeterinaryCollege,
AAU, Guwahati, Assam, India, for her valuable suggestions during
theprojectwork.
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