Effect of Noni (Morinda citrifolia Linn.) Fruit and Its Bioactive Principles Scopoletin and Rutin on Rat Vas Deferens Contractility: An Ex Vivo Study

Research ArticleEffect of Noni (Morinda citrifolia Linn) Fruit andIts Bioactive Principles Scopoletin and Rutin onRat Vas Deferens Contractility An Ex Vivo Study

Vijayapandi Pandy Megala Narasingam Thubasni KunasegaranDharmani Devi Murugan and Zahurin Mohamed

Department of Pharmacology Faculty of Medicine University of Malaya 50603 Kuala Lumpur Malaysia

Correspondence should be addressed to Vijayapandi Pandy pandiphdgmailcom

Received 18 February 2014 Revised 24 May 2014 Accepted 26 May 2014 Published 22 June 2014

Academic Editor Valdir Cechinel Filho

Copyright copy 2014 Vijayapandi Pandy et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

This study examined the effect of methanolic extract of Morinda citrifolia Linn (MMC) and its bioactive principles scopoletinand rutin on dopamine- and noradrenaline-evoked contractility in isolated rat vas deferens preparations MMC (1ndash40mgmL)scopoletin (1ndash200 120583gmL) and rutin hydrate (06ndash3126 120583gmL) dose-dependently inhibited the contractility evoked by submaximalconcentrations of both dopamine and noradrenaline respectively Haloperidol and prazosin reference dopamine D

2 and 120572

1-

adrenoceptors antagonists significantly reversed the dopamine- and noradrenaline-induced contractions respectively in a dose-dependent manner Interestingly MMC per se at higher doses (60ndash100mgmL) showed dose-dependent contractile response in ratvas deferens which was partially inhibited by high doses of haloperidol but not by prazosinThese results demonstrated the biphasiceffects of MMC on dopaminergic system that is antidopaminergic effect at lower concentrations (lt40mgmL) and dopaminergicagonistic effect at higher concentrations (gt60mgmL) However similar contractile response at high doses of scopoletin (05ndash5mgmL) and rutin hydrate (05ndash5mgmL) per se was not observed Therefore it can be concluded that the bioactive principles ofMMC scopoletin and rutin might be responsible for the antidopaminergic and antiadrenergic activities of MMC

1 Introduction

Morinda citrifolia Linn (family Rubiaceae) is also known asnoni Indian mulberry hog apple and cheese fruit The fruitflower leaves bark and root of M citrifolia have been usedfor diverse medicinal purposes as traditional plant-basedmedicine In fact the fruits of this plant have benefited thePolynesians over 2000 years as an alternative medicine forantibacterial antiviral antifungal antitumor anthelminticanalgesic hypotensive anti-inflammatory and immune-enhancing effects [1]

Thedecoction or infusions of roastedmature unripe fruitshave been recommended to relieve the symptoms of nauseaand vomiting [2] It has also been reported that noni fruitextract showed prokinetic and antiemetic activity on theintestinal transit being delayed by apomorphine (a potentagonist of dopamine D

2receptor) in mice and on the

apomorphine-induced emesis in dogs respectively Theseactions are attributed to antidopaminergic effect of nonifruit extract [3] In contrast Pu et al (2004) found thatgastric emptying was significantly inhibited in rats whichwere treated with noni juice (025 1 or 4mLkg) for 7 days[4] These opposing effects on gastric emptying observed fornoni by two different research groups might be due to thedifference in the type of fruit extract and the concentrationused in these experiments [5]

Our recent report revealed that the acute treatment ofmethanolic extract ofM citrifolia (MMC 1 3 5 and 10 gkgpo) significantly decreased the apomorphine-induced cageclimbing behaviour and methamphetamine-induced stereo-typy behaviour in mice in a dose-dependent manner whichdemonstrated the antidopaminergic effect of noni fruitextract [6] Furthermore it was also reported that the admin-istration of M citrifolia fruit extract significantly increased

Hindawi Publishing Corporatione Scientific World JournalVolume 2014 Article ID 909586 11 pageshttpdxdoiorg1011552014909586

2 The Scientific World Journal

the brain levels of serotonin dopamine and noradrenalinein rats [7] Similarly other neurochemical analysis of noni-treated rats (1mLday po) for 15 days revealed significantchanges in noradrenergic dopaminergic or serotonergicsystems in amygdala hippocampus and substantia nigra [8]Besides that in some preliminary studies nonirsquos capability inlowering blood pressure [9] and its vasodilatory properties[10] have been reported However the possible modes ofaction(s) of noni on the cardiovascular systemare lacking [11]

Studies have shown that noni fruit contains a number ofactive constituents including rutin (3310158404101584057-pentahydrox-yflavone-3-rhamnoglucoside) a flavonoid and scopoletin (6-methoxy-7-hydroxycoumarin) a coumarin derivative Rutinhas been reported to exert numerous pharmacological activ-ities including antioxidant anticarcinogenic neuroprotec-tive and anticonvulsant activity [12ndash15] Scopoletin hasbeen reported to exhibit antioxidant analgesic and anti-inflammatory effects [16 17] In addition scopoletin has beenrecommended as a marker constituent for the quality controland pharmacokinetic study of noni products [18] It has alsobeen demonstrated that scopoletin and rutin respectivelyshowed antidepressant activity mediated by its interactionwith 120572

1- and 120572

2-adrenoceptors [19 20] However there has

hitherto been no report demonstrating the interaction ofrutin and scopoletin on the dopaminergic system

Thus the present study aimed to investigate the inter-vention of noni (Morinda citrifolia Linn) and its bioactiveprinciples scopoletin and rutin on the dopaminergic andnoradrenergic systems using isolated rat vas deferens prepa-rations

2 Materials and Methods

21 Plant Collection and Identification Thefresh unripe fruits(dark green) of M citrifolia were collected in January 2012from Malacca Malaysia The plant material was taxonomi-cally identified and authenticated by Rimba Ilmu Institute ofBiological Sciences University of Malaya and the voucherspecimen (KLU 47738) was deposited at Rimba Ilmu forfuture reference The fruits were cut into thin slices andshade dried The shade dried fruit slices were pulverized ina mechanical grinder to obtain a coarse powder

22 Preparation of M citrifolia Fruit Extract Themethanolicextract of M citrifolia (MMC) was prepared using coldextraction with sonication The coarse fruit powder (18 kg)was soaked with 10L of methanol (Scharlau Spain isocraticHPLC grade) for 20 h followed by sonication using a waterbath sonicator at 30∘C for another 4 h The resultant solutionwas evaporated under vacuum in a rotary evaporator toobtain a dry mass extract (yield 1436 ww) The driedsolvent-free crude methanolic extract of M citrifolia wasstored at 4∘C in an air-tight labelled amber-coloured con-tainer until further use

23 Chromatography Conditions and Instrumentation MMCprepared as above was subjected to 1100 HPLC instrument(Agilent Technologies California USA) equipped with auto

sampler and a fraction collector Chromatographic sepa-rations were carried out on an Agilent Zorbax ODS C18(46 times 250mm 5 120583m 70A) and column temperature wasmaintained at 25∘CMobile phase composed ofmethanol and10 acetic acid in water was used Elution was performedwith a flow rate of 1mLminThe injection volume was 30 120583Land the detection wavelength was 365 nm Prior to injectionall samples were filtered through 020 120583m syringe filter intoHPLC sample vial to remove any remaining particles

HPLC-purified MMC was further subjected to LCMSanalysis The system comprised of Agilent 1290 Infinity LCsystem coupled to Agilent 6520 Accurate-Mass Q-TOF massspectrometer with dual ESI source used to analyse the sam-plesThe LC separation was carried out by an Agilent ZorbaxSB-C18 Narrow-Bore (21 times 150mm 35 120583m) LC parametersare autosampler temperature 25∘C injection volume 05 120583Lcolumn temperature 25∘C and flow rate of 021mLminA gradient system composed of solvents A (01 formicacid in water) to B (01 formic acid in 100 methanol)was employed Mass spectra data were acquired using thefollowing settings ESI capillary voltage was set at 4000V (+)ion mode and 4000V (minus) ion mode and fragmentor at 125VThe liquid nebulizerwas set to 45 psig and the nitrogen dryinggas was set to a flow rate of 10 Lmin Drying gas temperaturewas maintained at 300∘C The vaporizer temperature wasmaintained at 300∘C The ionization interface was operatedin both positive and negative modes Data was acquired ata rate of 103 spectras with a stored mass range of 100ndash3200mz for positive mode and 115ndash3200 mz for negative modeData was collected using Agilent MassHunter WorkstationData acquisition software LCMS data files were processedbyAgilentMassHunterQualitativeAnalysis B0500 softwareFeature finding was achieved using the molecular featureextraction (MFE) and correlation algorithms which locatesthe groups of covariant ions in each chromatogram Inpositive-ion mode this included adducts (H+ Na+ K+ and+NH4) and in negative ionmode it included adducts (minusHand

+Cl)

231 Tentative Compound Identifications The AgilentMETLIN (METLIN AM PCDL-N-130328cbd) databasewasused to make tentative identifications from the mass listscreated in Agilent Mass Profiler software This databaseincludes masses chemical formulas and structure infor-mation for various compounds All features were searchedagainst the METLIN database according to matchmz valueA list of identified compounds was generated

232 Quantification of Scopoletin and Rutin in MMC byLCMSMS Analysis Primary stock solution of rutin andscopoletin standard was prepared by dissolving the com-pounds in methanol to achieve the desired concentrationof 05mgmL Working standards solution was prepared byserial dilution of primary stock solution usingmethanol Peakarea was plotted against the corresponding concentrations toobtain calibration plot A MMC stock solution was preparedat the concentration of 10mgmL in methanol All thesamples prepared were filtered through a 022 nylon filter

The Scientific World Journal 3

(Bioflow Malaysia) and were transferred to autosampler vialfor LCMSMS quantification analysis The concentration ofeach constituent was expressed in 120583gmg of dry samples

The LCMSMS quantification analysis was performedusing Agilent 1290 infinity UHPLC coupled with Agilent6410 Triple quad LCMS The mass detector was equippedwith electrospray ionization (ESI) interface and controlledby MassHunter software 2120583L of all the samples preparedwas loaded on a 21mm (id) Narrow-BoreSB-C18 (length150mm) analytical column (particle size 35mM) used with aflow rate of 05mLmin in a solution A (01 formic acidin water) and solution B (100 acetonitrile with 01 formicacid) The total gradient time for the LCMS run was 11minThe ionization conditions were adjusted at 350∘C and 4000Vfor capillary temperature and voltage respectively The neb-ulizer pressure was 45 psi and the nitrogen flow rate was11 Lmin

24 Animals Sprague-Dawley male rats (250ndash300 g)obtained from the laboratory animal centre Universityof Malaya were used in this study The rats were housedin polycarbonate cages in a group of four animals understandard laboratory conditions at temperature of 22plusmn1∘C and1212 h lightdark cycle (light on from 8 am to 8 pm) Animalswere fed with standard laboratory food pellets and waterad libitum Animal Care and Use Committee University ofMalaya Kuala Lumpur approved the experimental protocol(ACUC Ethics number FAR27012012PV (R)) and care ofthe animals was taken as per guidelines of the Council forInternational Organization of Medical Sciences (CIOMS) onanimal experimentation [21]

25 Drugs and Chemicals Dopamine hydrochlorideL-noradrenaline prazosin hydrochloride haloperidolrutin hydrate dimethyl sulfoxide (Sigma-Aldrich) andscopoletin (Friendemann Schmidt Chemicals) were usedAll the drug solutions were prepared fresh in double distilledwater Dopamine hydrochloride and L-noradrenaline weredissolved in double distilled water containing ascorbic acid(0125wv) The stock solution of different concentrationsof MMC rutin hydrate and scopoletin was prepared usingDMSO (01 vv) All reagents were of isocratic HPLC gradeunless stated otherwise HPLC grade water was prepared byreverse osmosis and further purified

26 Effect of MMC Scopoletin and Rutin Hydrate on Dopam-ine or Noradrenaline-Evoked Contractile Response in IsolatedRat Vas Deferens The rats were sacrificed by CO

2inhalation

and pairs of vas deferens were dissected out and fixed ona wax plate with fine nail After carefully removing bloodvessels and connective tissues the epididymal portions of thevas deferens (length 1 cm) were cut from the preparationmounted with silk thread on to a hook in a 10mL organ bathand immersed in Krebs-Henseleit (K-H) solution gassedwith95 O

2plus 5 CO

2to achieve a pH of 73-74 the tem-

perature was 37∘C Preparations were stretched to a preloadtension of 500mg and equilibrated for 60min during whichtime the bath solution was changed every 15min Isometric

tensionwas recorded by a force transducer (Grass InstrumentCo Quincy MA USA) and the output was amplified andrecorded continuously using the PowerLab recording system(AD Instruments Sydney Australia) The K-H solutioncontained (mM) NaCl 119 KCl 47 CaCl

225 MgSO

410

NaHCO325 KH

2PO412 EDTA 003 and D-glucose 111 as

previously described [22] After the equilibration period thecontractile responses of vas deferens tissues were tested forviability by the repeated addition of high KCl solution (highK 80mM)Once reproducible contractions were obtainedwith high K a full noncumulative dose-response curve wasobtained by a stepwise increase in concentrations of theagonist (noradrenaline (NA) or dopamine (DA)) DA (04ndash1024 120583gmL) andNA (01ndash128120583gmL)were tested Each dosewas applied for 30s and thenwashed thoroughly (at least threetimes) before the next higher dose was added [23]

From the dose-response curves of DANA a submaximaldose of each agonist was selected for further studies In thepresence of DMSO (0001 vv) or the various concentra-tions of MMC (1ndash40mgmL 5min incubation) scopoletin(1ndash200120583gmL 5min incubation) and rutin hydrate (06ndash3126 120583gmL 5min incubation) in ascending order the con-tractile response of the chosen submaximal dose of DANAwas recorded Decrease in response with respect to thevehicle control (DMSO) represents inhibitory effect of theextractbioactive compounds on the corresponding agonistThe contraction of the chosen submaximal dose of DA(256 120583gmL) orNA (32 120583gmL) over highKCl solution (highK 80mM) in presence of DMSOMMCscopoletinrutinhydrate was calculated The same set of experiments wasalso repeated in presence of reference drugs haloperidol(dopamine D

2-receptor blocker) and prazosin HCl (120572

1-

adrenoceptor blocker) as positive controlsIn order to examine the nature of antagonism (reversible

or irreversible) by MMC on DANA-evoked contractileresponse in isolated rat vas deferens log dose-responsecurves of DANA in absence and presence of MMC (1ndash20mgmL 10min incubation) were carried out Similarlythe effect of MMC (1ndash40mgmL 10min incubation) on highK- (80mM) evoked contractility in rat vas deferens was alsostudied

In a separate experiment contractile response of MMC(20ndash100mgmL) per se was tested The contractile responseof MMC per se was recorded for 5min and washed thor-oughly with fresh K-H solution (at least three times in5min interval) before the next higher dose of MMC wasadded Following that contractile response of submaximaldose of MMC (80mgmL) was recorded in presence ofprazosin HCl (001 01 and 16 120583gmL) and haloperidol (128128 and 500 120583gmL) respectively Similarly the contractileresponses of scopoletin (05ndash5mgmL) and rutin hydrate(05ndash5mgmL) per se on rat vas deferens were also studied

27 Statistical Analysis The concentrations indicated in thetext or in the figures represent the final tissue-bath concen-tration of the respective drugs The responses were recordedas mean plusmn standard error of mean (SEM) and ldquo119899rdquo indicatesnumber of rats used for each set of data The statisticalsignificance of differences between treatments was evaluated


times104

8

6

4

2

0160 165 170 175 180 185 190 195 200 205 210 215 220 225

Counts versus mass-to-charge (mz)

H

H H

H HH

HH

O

O O O

1910342(MndashH)-

Cpd40 scopoletin minusESI MFE spectrum (14376ndash14827min) frag = 1250V MKU-5mgmL-MS-Neg

(a)

times104


Cpd

HO

HOHO

HO

41 rutin minusESI MFE spectrum (15713ndash1624 4min) frag = 1250V MKU-5mgmL-MS-Neg

O

O

OO O

O

OH

OH

OHOHOH

OH4

3

2

1

0580 585 590 595 600 605 610 615 620 625 630 635 640 645

6091472(MndashH)-

-30

CH3

(b)

Figure 1 MFE zoomed mass spectrum of (a) scopoletin and (b) rutin obtained fromMMC

by unpaired Studentrsquos 119905-test when comparing means of twogroups and one-way analysis of variance (ANOVA) andDunnett post hoc test for more than two group comparisonsA value of 119875 lt 005was considered as statistically significant

3 Results

31 Phytochemical Characterization of the MMC MS param-eters were optimized by individual standards of 10mgL oftarget compounds that were injected by infusion in full scanin positive and negative modes For scopoletin ESI negativeion mode provided the best response where unprotonatedmolecular ion [M-H]minus peak was observed at mz 1910342(Figure 1(a)) Rutin was detected only in negative modeThe unprotonated molecular ion [M-H]minus peak of rutin wasobserved atmz 6091472 (Figure 1(b))

The other components identified in full scan in positivemodes are monosaccharides D-glucoheptose fagomine andL-galactose disaccharides D-(+)-cellobiose and palatinosepolysaccharides levan and hyaluronic acid glycosides par-omomycin gentamicin C1a netilmicin and antibiotic JI-20A sugar acid isovalerylglucuronide sugar alcohol mesoe-rythritol erythritol and 14-dideoxy-14-imino-D-arabinitolquinone 4-bromo-35-cyclohexadiene-12-dione heterocy-cles naltrindole scopoletin and piperonyl butoxide aro-matic benzidine cyclic beta-23456-pentachlorocyclohex-anol polycyclic corrinoid fatty acids 4-hydroxy-valericacid 8S-hydroxy-hexadecanoic acid 23-dinor-TXB2 ara-chidic acid (d3) and 7-hydroxy tetranor iloprost fatty acylsn-valeryl acetic acid 12-hydroxy-10-octadecynoic acidand 910-dioxo-octadecanoic acid fatty alcohol 2299-tetramethyl-decan-110-diol and dolichyl phosphate sterollipids cholestan-3120572-ol31205726120572712057212120572-tetrahydroxy-5120573-cho-lan-24-oic acid alpha-tocopheronic acid and (24R)-112057224-dihydroxy-22-oxacholecalciferol glycerides 1-monopal-mitin terpenes and others tetrahydrocortisone cucurbitacinA and C253 67-epoxy highly branched isoprenoid amine1-naphthylacetylspermine amino acid Tyr Met Thr Ser AsnArg Ser N-acetyl-leucyl-leucine Ala His Ala His Arg AsnThr Gln Tyr Thr Tyr Tyr Gln His Gly and Leu Ala Argamide fenothiocarb sulfoxide

On the other hand adduct in LC-MS in negative modeprovided the following nucleoside azacitidine glucuronicacid isovalerylglucuronide carbohydrate acid arabinonicacid polysaccharides levan fatty acid 2-hydroxy-6-oxo-6-(2-hydroxyphenyl)-hexa-24-dienoate steroids fluocinolone

and dexamethasone phosphate fatty acyls 18-hydroxy-9S10R-epoxy-stearic acid 7-keto-stearic acid 24-dimethyl-tetradecanoic acid and DL-9-hydroxy stearic acid ter-penes antirrhinoside and monotropein heterocyclic com-pounds etanidazole 5-acetylamino-6-amino-3-methyluracilnifuradene 37-dimethyluric acid xanthine hypoxanthinescopoletin rutin and dioclein alkaloid atalanine amino acidderivatives fructoselysine 6-phosphate

32 Quantification of Scopoletin and Rutin in MMC byLCMSMS Analysis The concentration of scopoletin andrutin in MMC was found to be 1895 120583gmg and 166 120583gmgrespectively

33 Effect of MMC Scopoletin and Rutin Hydrate on DA-or NA-Evoked Contractile Response Dopamine and nora-drenaline produced concentration-dependent contractionsof epididymal segments of the isolated rat vas deferenspreparation are shown in Figures 2(a) and 2(b)

After incubation of the tissue preparation with MMC(1ndash40mgmL) scopoletin (1ndash200120583gmL) and rutin hydrate(06ndash3126 120583gmL) for 5min the contractile responses tothe selected submaximal dose of NA (32 120583gmL) and DA(256 120583gmL) were significantly reduced in dose-dependentmanner (Figures 3(a) 3(b) and 3(c) and Figures 4(a) 4(b)and 4(c))

34 Log Dose-Response Curve of NADA in Absence andin Presence of MMC As shown in Figures 5(a) 5(b) 6(a)and 6(b) the log dose-response curves of NA and DAwere dose-dependently shifted rightward in the presence ofMMC (1ndash20mgmL) The maxima of the agonists (NADA)curves were also progressively depressed by MMC in a dose-dependent manner The values of the calculated area underthe curve (AUC) of NA and DA were significantly decreasedby MMC dose-dependently

35 Effect of MMC on 80mM High K-Evoked ContractileResponse MMC (1ndash40mgmL) did not alter 80mMHigh K-evoked contractile response in rat vas deferens preparations(data not shown)

36 Effect of Haloperidol or PrazosinHCl onDA orNA-EvokedContractile Response While incubating the vas deferenspreparation with haloperidol (16ndash128 120583gmL) for 5min thecontractile responses to the selected submaximal dose of DA


Con

trac

tion

(ove

r hig

h K)

()

00 05 10 15 20 250

25

50

75

100

125

150

175

200

225

minus05

Log dose of dopamine (120583gmL)

(a)

00 05 10 150

10

20

30

40

Con

trac

tion

(ove

r hig

h K)

()

minus10 minus05

Log dose of NA (120583gmL)

(b)

Figure 2 Log dose-response curve of (a) dopamine and (b) noradrenaline (NA) employing isolated rat vas deferens preparation (119899 = 5-6)

DMSO0001

1 25 5 10 15 20 25 30 4000102030405060708090

100110120130140

MMC (mgmL)

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

(a)

DMSO0001

0123456789

10111213141516

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowast

1 2 4 6 8 10 50 100 200

Scopoletin (120583gmL)

(b)

DMSO0001

00

25

50

75

100

125

150

175

200

225

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowastlowastlowast

lowastlowast

06 12 24 48 98 195 391

Rutin (120583gmL)

(c)

Figure 3 Effect of (a)MMC (1ndash40mgmL) (b) scopoletin (1ndash200 120583gmL) and (c) rutin hydrate (06ndash391 120583gmL) onNA- (32 120583gmL) evokedcontraction expressed as contraction over 80mMHigh K lowast119875 lt 005 and lowastlowast119875 lt 001 compared to DMSO (0001 vv) 119899 = 5


DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)

Figure 4 Effect of (a) MMC (1ndash30mgmL) (b) scopoletin (1ndash200 120583gmL) and (c) rutin hydrate (06ndash3126 120583gmL) on dopamine-(256 120583gmL) evoked contraction in isolated rat vas deferens preparations expressed as contraction over 80mM High K lowast119875 lt 005lowastlowast

119875 lt 001 and lowastlowastlowast119875 lt 0001 compared to DMSO (0001 vv) 119899 = 5

00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL

MMC 10mgmLMMC 20mgmL

(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)

Figure 5 Log dose-response curve of (a) noradrenaline (NA) and (b) area under curve in absence and in presence of MMC expressed as contraction over 80mMHigh K lowast119875 lt 005 and lowastlowastlowast119875 lt 0001 compared to DMSO (0001 vv) 119899 = 5


0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001

Log dose of DA (120583gmL)

MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)

Figure 6 Log dose-response curve of (a) dopamine (DA) and (b) area under curve in absence and in presence of MMC expressed as contraction over 80mMHigh K lowast119875 lt 005 and lowastlowastlowast119875 lt 0001 compared to DMSO (0001 vv) 119899 = 5

0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128

Haloperidol (120583gmL)

Figure 7 Effect of haloperidol (16ndash128 120583gmL) on dopamine-(256 120583gmL) evoked contraction on isolated rat vas deferens prepa-ration lowastlowast119875 lt 001 compared to DMSO (0001 vv) 119899 = 4

(256 120583gmL) were significantly reduced in dose-dependentmanner (Figure 7) Prazosin HCl (001 120583gmL) showed 100inhibition on NA response (data not shown)

37 Effect of High Doses of MMC Scopoletin and Rutin PerSe on Contractility of Rat Vas Deferens Remarkably MMCper se at higher doses (60 80 and 100mgmL) signifi-cantly increased the contractility of vas deferens in a dose-dependent manner (Figure 8) However the bioactive prin-ciples of MMC scopoletin and rutin respectively at higherdoses (05ndash5mgmL) did not show any contractile response(data not shown) in rat vas deferens

38 Effect of Haloperidol or Prazosin HCl onHigh DoseMMC-Evoked Contractile Response Haloperidol (128ndash500120583gmL)

0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001

Figure 8 Contraction evoked by MMC (20ndash100mgmL) per seexpressed as contraction over 80mM High K lowastlowast119875 lt 001 com-pared to DMSO (0001 vv) 119899 = 4

significantly decreased the contractility induced by high doseMMC (80mgmL) (Figure 9(a)) On the other hand prazosinHCl even at higher concentration (16120583gmL) pretreatmentcould not alter the contractile response of high doseMMC(Figure 9(b))

4 Discussion

The isolated rodent vas deferens is an established researchmodel for functional and molecular investigation of normaland altered autonomic (sympathetic) neuroeffector cou-pling [24] It has also been suggested that dopamine maybe a neurotransmitter in the rat vas deferens and thatexcitatory dopamine receptors are located on the smoothmuscle [25ndash27] Boadle-Biber and Roth (1975) reported that


128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast

DMSO0001 HAL (120583gmL)

(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)

DMSO0001 PRZ (120583gmL)

(b)

Figure 9 Effect of haloperidol (a) (HAL 128 128 and 500 120583gmL) and (b) prazosin (PRZ 001 01 and 16 120583gmL) on MMC (80mgmL) perse induced contractility expressed as contraction over 80mMHigh K lowast119875 lt 005 compared to DMSO (0001 vv) 119899 = 4

a high proportion of newly formed catecholamine in therat vas deferens was present as dopamine as compared tonoradrenaline [28] Relja et al (1982) reported that dopaminereceptors are present in rat vas deferens based on the bindingof [3H] haloperidol to the plasma membranes [29] Besidesthat Ohmura et al (1992) indicated the occurrence of twodistinct120572

1-adrenoceptor subtypes in the rat vas deferens [30]

The preliminary phytochemical analysis of the methanolextract of M citrifolia showed the presence of carbohy-drate alkaloids saponin glycosides tannins flavonoids andsteroids [31] Phytochemical studies using high performanceliquid chromatographic (HPLC) fingerprint profile of theMeOH extracts of noni fruit revealed three major peaksrepresenting scopoletin rutin and quercetin together withseveral minor peaks These are major bioactive constituentsof noni responsible for various pharmacological activities [3233]The present phytochemical analysis results are consistentwith earlier findings and confirmed the presence of scopo-letin and rutin (retention times 1451 and 1586min resp)Furthermore we estimated the concentration of these con-stituents in MMC using LCMSMS analysis and it has beenfound that MMC is enriched with scopoletin (1895 120583gmg)and rutin (166 120583gmg) Hence in the present study scopo-letin and rutin were used to establish its interaction withdopaminergic and noradrenergic systems

In the present study we observed that MMC(lt40mgmL) scopoletin (100 200120583gmL) and rutin hydrate(1563 3126 120583gmL) caused a significant concentration-dependent inhibition of contraction evoked by dopaminein the isolated rat vas deferens preparations thus indicatingantidopaminergic activity of MMC scopoletin and rutinhydrate (Figures 4(a) 4(b) and 4(c)) It has been observedthat MMC [1ndash20mgmL equivalent to scopoletin (1895ndash379 120583gmL) and rutin (166ndash332 120583gmL)] dose-dependentlydecrease the slope as well as maxima of the agonists(DANA) curves (Figures 5(a) 5(b) 6(a) and 6(b)) and

suggested that the antagonistic effect of MMC (lt40mgmL)on dopaminergic and noradrenergic system might beirreversible Furthermore MMC (lt40mgmL) did not alterthe contraction induced by high K (80mM KCl) in rat vasdeferens This observation excludes the possibility of theobserved inhibitory effect of MMC (lt40mgmL) on rat vasdeferens being due to direct muscle relaxing effect of MMCThe reference drug haloperidol (dopamine D

2receptor

blocker 16ndash128 120583gmL) also showed dose-dependentinhibition on dopamine-induced contractile response in ratvas deferens Conversely MMC per se at higher doses (6080 and 100mgmL) exhibited significant dose-dependentcontractile response Interestingly contractility evoked byMMC per se was significantly inhibited by high doses ofhaloperidol (128ndash500120583gmL) Thus the present results havedemonstrated the biphasic effect of MMC on dopaminergicsystem with dopaminergic antagonistic effect at lowerconcentrations (lt40mgmL) and dopaminergic agonisticeffect at higher concentrations (gt60mgmL) However themain bioactive principles of noni scopoletin and rutinper se at higher doses (05ndash5mgmL) did not show anycontractile response in rat vas deferens Therefore it can bepostulated that the antidopaminergic effects of MMC mightbe mediated by its active principles scopoletin and rutinNevertheless these active principles might not be involvedin the high dose contractile response of MMC per se Thereis a possibility that some other bioactive phytoconstituentsof MMC could be involved in the high dose contractileresponse of MMC

M citrifolia has been used as an antiemetic agent inpatients considered with high risk for postoperative nauseaand vomiting (PONV) after various types of surgery The600mg dose of noni extract (equivalent to 20 g of driednoni fruit8712120583g of scopoletin) was the minimum dosethat could effectively reduce the incidence of postoperativenausea in the early postoperative period [34] However


these studies could not reveal the possible mechanism ofaction of M citrifolia for its antiemetic action In ourrecent published report we suggested the antipsychotic andantiemetic activities of M citrifolia might be mediated byantidopaminergic mechanism [6] Another recent reporton neurochemical analysis revealed the alterations in themonoaminergic system in the noni-treated rats (1mLdaypo for 15 days) compared to the control rats It was foundthat the reduction in these neurotransmitters including nora-drenaline in the amygdala and the hippocampus serotoninin the amygdala DOPAC in the hippocampus and thesubstantia nigra and HVA in the substantia nigra of thenoni groupmight be responsible for anxiolytic effects of nonijuice [8] Conversely it has been demonstrated that ethylacetate fraction of crude methanol extract of M citrifolia ata daily dose of 200 and 400mgkg which was administeredto the rats for 15 days significantly increased the brainlevels of serotonin dopamine and noradrenaline [7] Thisopposing effect of noni on monoamine levels in two differentstudies could be due to the difference in the doses of noniused for these studies Lower doses of noni might inhibitmonoaminergic system whereas higher concentrations couldstimulate monoaminergic systemThe enhancement in brainmonoamines after 15-day treatment of ethyl acetate fractionof crude methanol extract of M citrifolia might be due tothe presence of enrichment of active constituents in theethyl acetate fraction The higher concentration of activeconstituents in the ethyl acetate fraction when comparedto crude MMC could activate the monoaminergic systemThe present results are in good agreement with these earlierfindings that is MMC can act as a dopaminergic antagonistat lower concentrations (lt40mgmL) and as a dopaminer-gic agonist at higher dose (gt60mgmL) However furtherreceptor-ligand binding assays are necessary to confirm thebiphasic effects ofM citrifolia on dopaminergic system

In the present study MMC showed antiadrenergic effectat lower concentrations [lt30mgmL equivalent to scopo-letin (lt5685120583gmL) and rutin (lt498 120583gmL)] in the isolatedrat vas deferens preparation Besides that scopoletin (50100 200120583gmL) and rutin hydrate (98 195 391 120583gmL)caused a significant concentration-dependent inhibition ofcontraction evoked by noradrenaline (Figures 3(a) 3(b)and 3(c)) The reference drug prazosin HCl (adrenergic 120572

1

receptor blocker) completely reversed the noradrenaline-evoked contractility in the isolated rat vas deferens evenat lower concentration of 001 120583gmL A recent researchreport suggested that the spasmolytic and vasodilator effectsof M citrifolia root extract are mediated possibly throughblockade of voltage-dependent calcium channels and releaseof intracellular calcium and justified traditional medicinalusage of M citrifolia in diarrhoea and hypertension [11]These studies also demonstrated the vasorelaxant activity ofM citrifolia by inhibiting contractile response of 120572

1adreno-

ceptors agonist phenylephrine (1 120583M) in endothelium-intactrat aortic preparations [11] The present results are consistentwith these earlier findings and suggest that the antihyper-tensive effect of M citrifolia might be partially mediated by1205721adrenoceptors blocking effect The present study results

also revealed the antiadrenergic activity of scopoletin and

rutin respectively in rat vas deferens Therefore it can bepostulated that 120572

1adrenoceptors blocking effect of MMC

could be mediated by its bioactive principles scopoletinand rutin An earlier report revealed the predominanceof 120572-adrenoceptors in the testicular (epididymal) segmentof the vas deferens compared with the urethral (prostatic)segment [35] A nerve-stimulation evoked postjunctionalreceptor for NA is the 120572

1A-adrenoceptor acting via inositoltrisphosphate (IP

3) leading to increase in intracellular Ca2+

and the slow component of nerve-mediated contraction [36]However the possibility that there are differences betweenneurogenic and exogenously appliedNAhas been raised [37]They showed that contractions to exogenous NA involvedboth 120572

1A and 1205722AD adrenoceptors Hence the antiadrenergic

effect of MMC (lt30mgmL) and its bioactive principlesscopoletin and rutin could be mediated by both 120572

1A and1205722AD adrenoceptors The contractile response of high dose

of MMC (80mgmL) was not inhibited by 1205721-adrenoceptor

blocker prazosin even at higher concentration (16 120583gmL)Therefore the contractility response of high dose of MMCmight not be mediated through adrenergic mechanism Thebiphasic effect of MMC observed in the present study usingrat vas deferens preparation could be exhibited only at thedopaminergic system

5 Conclusion

In conclusion MMC showed biphasic effect on dopamin-ergic system that is antidopaminergic effect at lower dose(lt40mgmL) and dopaminergic agonistic effect at higherdose (gt60mgmL) in the isolated rat vas deferens prepa-ration Additionally MMC (lt30mgmL) showed the anti-adrenergic activity in the rat vas deferens Furthermoreantidopaminergic and antiadrenergic activities of scopoletin(lt200120583gmL) and rutin hydrate (lt3126 120583gmL) respec-tively have been established It has been postulated thatthe bioactive principles of noni scopoletin and rutin couldbe responsible for the antidopaminergic and antiadrenergicactivities of MMC However the mechanism of high dosecontractile response of MMC on rat vas deferens could notbe explained in the present study Further in vivo animalbehaviour studies arewarranted to confirm the biphasic effectof noni on dopaminergic system Studies in this direction arecurrently under way in our laboratory

Conflict of Interests

The authors declare that they have no competing interests

Acknowledgments

This study was supported by University of Malaya ResearchGrants (PG015-2012B and RG495-13HTM) and HIR MOHEProject no UMC6251HIRMOHEMED05 (H-20001-E000088)

References

[1] S Nayak and R Shettigar ldquoMorinda citrifolia a reviewrdquo Journalof Pharmacy Research vol 3 no 8 pp 1872ndash1874 2010


[2] W Ekpalakorn S Supjaroen P Keawkomol K ChompuwisetP Limangkoon and W Boonchui ldquoA clinical study ofMorindacitrifolia Linnin the treatment of nausea and vomitingrdquo inOffice of the Primary Health Care Ministry of Public HealthResearch Reports on Medicinal Plants Medicinal Plants and Pri-mary Health Care Project pp 40ndash41 Veteran AdministrationPrinting Bangkok Thailand 1987

[3] A Chuthaputti P N Pattaloong U Permpipat and Y Techad-amrongsin ldquoStudy on antiemetic activity of Morinda citrifoliafruitsrdquoThai Journal of Pharmaceutical Science vol 20 pp 195ndash202 1996

[4] H-F Pu W-J Huang W-M Tseng et al ldquoEffects of juice fromMorinda citrifolia (Noni) on gastric emptying in male ratsrdquoChinese Journal of Physiology vol 47 no 4 pp 169ndash174 2004

[5] S Mahattanadul W Ridtitid S Nima N Phdoongsombut PRatanasuwon and S Kasiwong ldquoEffects of Morinda citrifoliaaqueous fruit extract and its biomarker scopoletin on refluxesophagitis and gastric ulcer in ratsrdquo Journal of Ethnopharma-cology vol 134 no 2 pp 243ndash250 2011

[6] V Pandy M Narasingam and Z Mohamed ldquoAntipsychotic-like activity of Noni (Morinda citrifolia Linn) in micerdquo BMCComplementary and Alternative Medicine vol 12 article 1862012

[7] P Muralidharan and J Srikanth ldquoAnti epileptic activity ofMorinda citrifolia linn fruit extractrdquo E-Journal of Chemistry vol7 no 2 pp 612ndash616 2010

[8] S Kalandakanond-Thongsong and J CharoenphandhuldquoAnxiolytic-like effects of noni juice (Morinda citrifolia L) Onthe respective changes of neurotransmitters in rat brain in theelevated plus-maze testrdquo Thai Journal of Veterinary Medicinevol 42 no 3 pp 275ndash280 2012

[9] Dang-Van-Ho ldquoTreatment and prevention of hypertension andits cerebral complications by total root extracts of Morindacitrifoliardquo Pressemedicale vol 63 no 72 p 1478 1955 (French)

[10] I Runnie M N Salleh S Mohamed R J Head and MY Abeywardena ldquoVasorelaxation induced by common edibletropical plant extracts in isolated rat aorta and mesentericvascular bedrdquo Journal of Ethnopharmacology vol 92 no 2-3pp 311ndash316 2004

[11] A H Gilani S-U Mandukhail J Iqbal et al ldquoAntispasmodicand vasodilator activities of Morinda citrifolia root extractare mediated through blockade of voltage dependent calciumchannelsrdquo BMC Complementary and Alternative Medicine vol10 article 2 2010

[12] C la Casa I Villegas C Alarcon de la Lastra V Motilva andM J Martın Calero ldquoEvidence for protective and antioxidantproperties of rutin a natural flavone against ethanol inducedgastric lesionsrdquo Journal of Ethnopharmacology vol 71 no 1-2pp 45ndash53 2000

[13] K H Janbaz S A Saeed and A H Gilani ldquoProtective effectof rutin on paracetamol- and CCl4-induced hepatotoxicity inrodentsrdquo Fitoterapia vol 73 no 7-8 pp 557ndash563 2002

[14] F Pu K Mishima K Irie et al ldquoNeuroprotective effects ofquercetin and rutin on spatial memory impairment in an 8-arm radial maze task and neuronal death induced by repeatedcerebral ischemia in ratsrdquo Journal of Pharmacological Sciencesvol 104 no 4 pp 329ndash334 2007

[15] M Nassiri-Asl S Shariati-Rad and F Zamansoltani ldquoAnti-convulsive effects of intracerebroventricular administrationof rutin in ratsrdquo Progress in Neuro-Psychopharmacology andBiological Psychiatry vol 32 no 4 pp 989ndash993 2008

[16] C-Y Shaw C-H Chen C-C Hsu C-C Chen and Y-C Tsai ldquoAntioxidant properties of scopoletin isolated fromSinomonium acutumrdquo Phytotherapy Research vol 17 no 7 pp823ndash825 2003

[17] T-N Chang J-S Deng Y-C Chang et al ldquoAmeliorativeeffects of scopoletin from Crossostephium chinensis againstinflammation pain and its mechanisms in micerdquo Evidence-Based Complementary and Alternative Medicine vol 2012Article ID 595603 10 pages 2012

[18] B F Issell A Franke and R M Fielding ldquoPharmacokineticstudy of noni fruit extractrdquo Journal of Dietary Supplements vol5 no 4 pp 373ndash382 2008

[19] J C Capra M P Cunha D G Machado et al ldquoAntidepressant-like effect of scopoletin a coumarin isolated from Polygalasabulosa (Polygalaceae) in mice evidence for the involvementofmonoaminergic systemsrdquo European Journal of Pharmacologyvol 643 no 2-3 pp 232ndash238 2010

[20] D G Machado L E B Bettio M P Cunha et al ldquoAntidepress-ant-like effect of rutin isolated from the ethanolic extract fromSchinus molle L in mice evidence for the involvement of theserotonergic and noradrenergic systemsrdquo European Journal ofPharmacology vol 587 no 1ndash3 pp 163ndash168 2008

[21] N Howard-Jones ldquoA CIOMS ethical code for animal experi-mentationrdquoWHO Chronicle vol 39 no 2 pp 51ndash56 1985

[22] Y Huang ldquoInhibition of contractions by tricyclic antidepres-sants and xylamine in rat vas deferensrdquo European Journal ofPharmacology vol 327 no 1 pp 41ndash47 1997

[23] P Vijayapandi A Vamsi Krishna B Srikanth et al ldquoIn vitroanticholinergic and antihistaminic activities of Acorus calamusLinn leaves extractsrdquo African Journal of Traditional Comple-mentary and Alternative Medicines vol 10 no 1 pp 95ndash1012013

[24] T D Westfall and D P Westfall ldquoPharmacological techniquesfor the in vitro study of the vas deferensrdquo Journal of Pharma-cological and Toxicological Methods vol 45 no 2 pp 109ndash1222001

[25] A Simon and E F Van Maanen ldquoDopamine receptors anddopaminergic nerves in the vas deferens of the ratrdquo ArchivesInternationales de Pharmacodynamie et deTherapie vol 222 no1 pp 4ndash15 1976

[26] F M Tayo ldquoOccurrence of excitatory dopaminoceptors in therat and guinea-pig vas deferensrdquo Clinical and ExperimentalPharmacology and Physiology vol 6 no 3 pp 275ndash279 1979

[27] A Badia P Bermejo and F Jane ldquoPre- and postsynaptic effectsof sulpiride in the rat isolated vas deferensrdquo Journal of Pharmacyand Pharmacology vol 34 no 4 pp 266ndash268 1982

[28] M C Boadle-Biber and R H Roth ldquoFormation of dopamineand noradrenaline in rat vas deferens comparison with guineapig vas deferensrdquo British Journal of Pharmacology vol 55 no 1pp 73ndash78 1975

[29] M Relja Z Lackovic andNHNeff ldquoEvidence for the presenceof dopaminergic receptors in vas deferensrdquo Life Sciences vol 31no 23 pp 2571ndash2575 1982

[30] T Ohmura M Oshita S Kigoshi and I Muramatsu ldquoIdentifi-cation of1205721-adrenoceptor subtypes in the rat vas deferens bind-ing and functional studiesrdquoBritish Journal of Pharmacology vol107 no 3 pp 697ndash704 1992

[31] S Ramesh M Radhakrishnan R Anburaj R Elangomatha-van and S Patharajan ldquoPhysicochemical phytochemical andantimicrobial studies on Morinda citrifolia L fruits at differentmaturity stagesrdquo International Journal of Pharmacy and Phar-maceutical Sciences vol 4 no 5 pp 473ndash476 2012


[32] S Deng B J West A K Palu B-N Zhou and C J JensenldquoNoni as an anxiolytic and sedative a mechanism involving itsgamma-aminobutyric acidergic effectsrdquo Phytomedicine vol 14no 7-8 pp 517ndash522 2007

[33] S D Pachauri S Tota K Khandelwal et al ldquoProtective effect offruits ofMorinda citrifolia L on scopolamine induced memoryimpairment in mice a behavioral biochemical and cerebralblood flow studyrdquo Journal of Ethnopharmacology vol 139 no1 pp 34ndash41 2012

[34] S Prapaitrakool and A Itharat ldquoMorinda citrifolia Linn forprevention of postoperative nausea and vomitingrdquo Journal of theMedical Association of Thailand vol 93 no 7 pp S204ndashS2092010

[35] L Vardolov and J N Pennefather ldquoRegional variation in thedistribution of 120572 adrenoreceptors in the vas deferens of the ratrdquoArchives Internationales de Pharmacodynamie et de Therapievol 221 no 2 pp 212ndash222 1976

[36] V Honner and J R Docherty ldquoInvestigation of the subtypesof 1205721-adrenoceptor mediating contractions of rat vas deferensrdquoBritish Journal of Pharmacology vol 128 no 6 pp 1323ndash13311999

[37] L Cleary C Vandeputte and J R Docherty ldquoInvestigation ofpostjunctional 120572

1- and 120572

2-adrenoceptor subtypes in vas def-

erens from wild-type and 1205722AD-adrenoceptor knockout micerdquoBritish Journal of Pharmacology vol 138 no 6 pp 1069ndash10762003

Submit your manuscripts athttpwwwhindawicom

PainResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom

Volume 2014

ToxinsJournal of

VaccinesJournal of

Hindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

AntibioticsInternational Journal of

ToxicologyJournal of


StrokeResearch and TreatmentHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Drug DeliveryJournal of



Advances in Pharmacological Sciences

Tropical MedicineJournal of


Medicinal ChemistryInternational Journal of


AddictionJournal of



BioMed Research International

Emergency Medicine InternationalHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Autoimmune Diseases


Anesthesiology Research and Practice

ScientificaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of


Pharmaceutics


MEDIATORSINFLAMMATION

of


the brain levels of serotonin dopamine and noradrenalinein rats [7] Similarly other neurochemical analysis of noni-treated rats (1mLday po) for 15 days revealed significantchanges in noradrenergic dopaminergic or serotonergicsystems in amygdala hippocampus and substantia nigra [8]Besides that in some preliminary studies nonirsquos capability inlowering blood pressure [9] and its vasodilatory properties[10] have been reported However the possible modes ofaction(s) of noni on the cardiovascular systemare lacking [11]

Studies have shown that noni fruit contains a number ofactive constituents including rutin (3310158404101584057-pentahydrox-yflavone-3-rhamnoglucoside) a flavonoid and scopoletin (6-methoxy-7-hydroxycoumarin) a coumarin derivative Rutinhas been reported to exert numerous pharmacological activ-ities including antioxidant anticarcinogenic neuroprotec-tive and anticonvulsant activity [12ndash15] Scopoletin hasbeen reported to exhibit antioxidant analgesic and anti-inflammatory effects [16 17] In addition scopoletin has beenrecommended as a marker constituent for the quality controland pharmacokinetic study of noni products [18] It has alsobeen demonstrated that scopoletin and rutin respectivelyshowed antidepressant activity mediated by its interactionwith 120572

1- and 120572

2-adrenoceptors [19 20] However there has

hitherto been no report demonstrating the interaction ofrutin and scopoletin on the dopaminergic system

Thus the present study aimed to investigate the inter-vention of noni (Morinda citrifolia Linn) and its bioactiveprinciples scopoletin and rutin on the dopaminergic andnoradrenergic systems using isolated rat vas deferens prepa-rations

2 Materials and Methods

21 Plant Collection and Identification Thefresh unripe fruits(dark green) of M citrifolia were collected in January 2012from Malacca Malaysia The plant material was taxonomi-cally identified and authenticated by Rimba Ilmu Institute ofBiological Sciences University of Malaya and the voucherspecimen (KLU 47738) was deposited at Rimba Ilmu forfuture reference The fruits were cut into thin slices andshade dried The shade dried fruit slices were pulverized ina mechanical grinder to obtain a coarse powder

22 Preparation of M citrifolia Fruit Extract Themethanolicextract of M citrifolia (MMC) was prepared using coldextraction with sonication The coarse fruit powder (18 kg)was soaked with 10L of methanol (Scharlau Spain isocraticHPLC grade) for 20 h followed by sonication using a waterbath sonicator at 30∘C for another 4 h The resultant solutionwas evaporated under vacuum in a rotary evaporator toobtain a dry mass extract (yield 1436 ww) The driedsolvent-free crude methanolic extract of M citrifolia wasstored at 4∘C in an air-tight labelled amber-coloured con-tainer until further use

23 Chromatography Conditions and Instrumentation MMCprepared as above was subjected to 1100 HPLC instrument(Agilent Technologies California USA) equipped with auto

sampler and a fraction collector Chromatographic sepa-rations were carried out on an Agilent Zorbax ODS C18(46 times 250mm 5 120583m 70A) and column temperature wasmaintained at 25∘CMobile phase composed ofmethanol and10 acetic acid in water was used Elution was performedwith a flow rate of 1mLminThe injection volume was 30 120583Land the detection wavelength was 365 nm Prior to injectionall samples were filtered through 020 120583m syringe filter intoHPLC sample vial to remove any remaining particles

HPLC-purified MMC was further subjected to LCMSanalysis The system comprised of Agilent 1290 Infinity LCsystem coupled to Agilent 6520 Accurate-Mass Q-TOF massspectrometer with dual ESI source used to analyse the sam-plesThe LC separation was carried out by an Agilent ZorbaxSB-C18 Narrow-Bore (21 times 150mm 35 120583m) LC parametersare autosampler temperature 25∘C injection volume 05 120583Lcolumn temperature 25∘C and flow rate of 021mLminA gradient system composed of solvents A (01 formicacid in water) to B (01 formic acid in 100 methanol)was employed Mass spectra data were acquired using thefollowing settings ESI capillary voltage was set at 4000V (+)ion mode and 4000V (minus) ion mode and fragmentor at 125VThe liquid nebulizerwas set to 45 psig and the nitrogen dryinggas was set to a flow rate of 10 Lmin Drying gas temperaturewas maintained at 300∘C The vaporizer temperature wasmaintained at 300∘C The ionization interface was operatedin both positive and negative modes Data was acquired ata rate of 103 spectras with a stored mass range of 100ndash3200mz for positive mode and 115ndash3200 mz for negative modeData was collected using Agilent MassHunter WorkstationData acquisition software LCMS data files were processedbyAgilentMassHunterQualitativeAnalysis B0500 softwareFeature finding was achieved using the molecular featureextraction (MFE) and correlation algorithms which locatesthe groups of covariant ions in each chromatogram Inpositive-ion mode this included adducts (H+ Na+ K+ and+NH4) and in negative ionmode it included adducts (minusHand

+Cl)

231 Tentative Compound Identifications The AgilentMETLIN (METLIN AM PCDL-N-130328cbd) databasewasused to make tentative identifications from the mass listscreated in Agilent Mass Profiler software This databaseincludes masses chemical formulas and structure infor-mation for various compounds All features were searchedagainst the METLIN database according to matchmz valueA list of identified compounds was generated

232 Quantification of Scopoletin and Rutin in MMC byLCMSMS Analysis Primary stock solution of rutin andscopoletin standard was prepared by dissolving the com-pounds in methanol to achieve the desired concentrationof 05mgmL Working standards solution was prepared byserial dilution of primary stock solution usingmethanol Peakarea was plotted against the corresponding concentrations toobtain calibration plot A MMC stock solution was preparedat the concentration of 10mgmL in methanol All thesamples prepared were filtered through a 022 nylon filter







2inhalation


2plus 5 CO




225 MgSO

410

NaHCO325 KH





1-






times104

8

6

4

2

0160 165 170 175 180 185 190 195 200 205 210 215 220 225


H

H H

H HH

HH

O

O O O

1910342(MndashH)-


(a)

times104


Cpd

HO

HOHO

HO


O

O

OO O

O

OH

OH

OHOHOH

OH4

3

2

1

0580 585 590 595 600 605 610 615 620 625 630 635 640 645

6091472(MndashH)-

-30

CH3

(b)



3 Results












Con

trac

tion

(ove

r hig

h K)

()

00 05 10 15 20 250

25

50

75

100

125

150

175

200

225

minus05


(a)

00 05 10 150

10

20

30

40

Con

trac

tion

(ove

r hig

h K)

()

minus10 minus05


(b)


DMSO0001

1 25 5 10 15 20 25 30 4000102030405060708090

100110120130140

MMC (mgmL)

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

(a)

DMSO0001

0123456789

10111213141516

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowast

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

00

25

50

75

100

125

150

175

200

225

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()


lowastlowast

06 12 24 48 98 195 391

Rutin (120583gmL)

(c)



DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)



00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL


(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)



0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of







2inhalation


2plus 5 CO




225 MgSO

410

NaHCO325 KH





1-






times104

8

6

4

2

0160 165 170 175 180 185 190 195 200 205 210 215 220 225


H

H H

H HH

HH

O

O O O

1910342(MndashH)-


(a)

times104


Cpd

HO

HOHO

HO


O

O

OO O

O

OH

OH

OHOHOH

OH4

3

2

1

0580 585 590 595 600 605 610 615 620 625 630 635 640 645

6091472(MndashH)-

-30

CH3

(b)



3 Results












Con

trac

tion

(ove

r hig

h K)

()

00 05 10 15 20 250

25

50

75

100

125

150

175

200

225

minus05


(a)

00 05 10 150

10

20

30

40

Con

trac

tion

(ove

r hig

h K)

()

minus10 minus05


(b)


DMSO0001

1 25 5 10 15 20 25 30 4000102030405060708090

100110120130140

MMC (mgmL)

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

(a)

DMSO0001

0123456789

10111213141516

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowast

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

00

25

50

75

100

125

150

175

200

225

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()


lowastlowast

06 12 24 48 98 195 391

Rutin (120583gmL)

(c)



DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)



00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL


(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)



0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of


times104

8

6

4

2

0160 165 170 175 180 185 190 195 200 205 210 215 220 225


H

H H

H HH

HH

O

O O O

1910342(MndashH)-


(a)

times104


Cpd

HO

HOHO

HO


O

O

OO O

O

OH

OH

OHOHOH

OH4

3

2

1

0580 585 590 595 600 605 610 615 620 625 630 635 640 645

6091472(MndashH)-

-30

CH3

(b)



3 Results












Con

trac

tion

(ove

r hig

h K)

()

00 05 10 15 20 250

25

50

75

100

125

150

175

200

225

minus05


(a)

00 05 10 150

10

20

30

40

Con

trac

tion

(ove

r hig

h K)

()

minus10 minus05


(b)


DMSO0001

1 25 5 10 15 20 25 30 4000102030405060708090

100110120130140

MMC (mgmL)

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

(a)

DMSO0001

0123456789

10111213141516

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowast

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

00

25

50

75

100

125

150

175

200

225

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()


lowastlowast

06 12 24 48 98 195 391

Rutin (120583gmL)

(c)



DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)



00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL


(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)



0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of


Con

trac

tion

(ove

r hig

h K)

()

00 05 10 15 20 250

25

50

75

100

125

150

175

200

225

minus05


(a)

00 05 10 150

10

20

30

40

Con

trac

tion

(ove

r hig

h K)

()

minus10 minus05


(b)


DMSO0001

1 25 5 10 15 20 25 30 4000102030405060708090

100110120130140

MMC (mgmL)

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

(a)

DMSO0001

0123456789

10111213141516

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowast

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

00

25

50

75

100

125

150

175

200

225

Con

trac

tion

of32120583

gm

L of

NA

(ove

r hig

h K)

()


lowastlowast

06 12 24 48 98 195 391

Rutin (120583gmL)

(c)



DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)



00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL


(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)



0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of


DMSO0001

1 25 50 100 15 20 25 300

25

50

75

100

125

150

MMC (mgmL)

lowast

lowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

(a)

0

50

100

150

200

lowastlowast

lowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

1 2 4 6 8 10 50 100 200


(b)

DMSO0001

0

50

100

150

200

lowastlowast

lowastlowastlowast

Con

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

Rutin (120583gmL)

06 12 24 48 98 195 391 781 15633126

(c)



00 05 10 150

20

40

60

80

100

DMSO 0001

Con

trac

tion

(ove

r hig

h K)

()

lowastlowastlowast

lowastlowastlowast

minus10 minus05


MMC 1mgmLMMC 5mgmL


(a)

1 5 10 200

100

200

300

400

MMC (mgmL)

AUC

of N

A (o

ver h

igh

K)

lowast

DMSO0001

(b)



0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of


0 1 20

20

40

60

80

100C

ontr

actio

n (o

ver h

igh

K) (

)

lowastlowastlowast

lowastlowastlowast

minus1

DMSO 0001


MMC 1mgmLMMC 5mgmL


(a)

0

100

200

300

400

AUC

of D

A (o

ver h

igh

K)

lowast

1 5 10 20

MMC (mgmL)

DMSO0001

(b)


0

25

50

75

100

125

lowastlowast

lowastlowastCon

trac

tion

of256120583

gm

L of

DA

(ove

r hig

h K)

()

DMSO0001

16 32 64 128






0102030405060708090

100110120

Con

trac

tion

of M

MC

(ove

r hig

h K)

()

lowastlowast

lowastlowast

lowastlowast

806040 10020

MMC (mgmL)

DMSO0001



4 Discussion



128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of


128 128 5000

25

50

75

100C

ontr

actio

n of

80 m

gm

L M

MC

(ove

r hig

h K)

()

lowast

lowast


(a)

001 01 160

25

50

75

100

Con

trac

tion

of 8

0 mg

mL

MM

C (o

ver h

igh

K) (

)


(b)







2receptor






1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of




1


1adreno-














5 Conclusion




Acknowledgments


References








































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of







































1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of








1- and 120572







Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of





Volume 2014

ToxinsJournal of

VaccinesJournal of















AddictionJournal of






Autoimmune Diseases




Journal of


Pharmaceutics



of

Documents

Effect of Noni (Morinda citrifolia Linn.) Fruit and Its Bioactive Principles Scopoletin and Rutin on Rat Vas Deferens Contractility: An Ex Vivo Study