R E S E A R C H A R T I C L E Open Access

Assessment of phytochemical content,

polyphenolic composition, antioxidant and

antibacterial activities of Leguminosae medicinal

plants in Peninsular Malaysia

Yik Ling Chew

1

, Elaine Wan Ling Chan

1

, Pei Ling Tan

1

, Yau Yan Lim

1

, Johnson Stanslas

2,3

, Joo Kheng Goh

1*

Abstract

Background: Many medicinal plants from Leguminosae family can be found easily in Malaysia. These plant s have been used as traditional medicines by local ethnic groups, where they are prepared as decoction, pastes for

wound infections, and some have been eaten as salad. This paper focused on the assessment of antioxidant potential, antibacterial activity and classes of phytochemicals of nine plants from the Leguminosae family.

Met hods : Acacia auriculiformis, Bauhinia kockiana, Bauhinia purpurea, Caesalpinia pulcherrima, Calliandra tergemina, Cassia surattensis, Leucaena leucocephala, Peltophorum pterocarpum, and Samanea saman were extracted with aqueous methanol and dichloromethane : methanol mixture to test for antioxidant and antibacterial activities. The

Folin-Ciocalteu assay was conducted to quantify the total phenolic content and 2, 2-diphenyl -1-picrylhydrazyl (DPPH) assay was used to determine the free radical quenching capacity. Antibacterial activity was assessed using disc diffusion (Kirby -Bauer) assay. Screening for major classes of phytochemical was done using standard chemical tests.

Results: B. kockiana flowers and C. pulcherrima leaves contained high total phenolic content (TPC) and strong DPPH

radical scavenging ability with TPC of 8280 ± 498 mg GAE/1 00 g, IC50 of 27.0 ± 5.0 μ g/mL and TPC of 5030 ± 602 mg GAE/1 00 g, IC 50 of 50.0 ± 5.0 μ g/mL respectively. Positive correlation was observed between TPC and free radical

scavenging ability. Most extracts showed antibacterial activity against Gram positive bacteria at 1 mg , while none showed activity against Gram negative bacteria at the same dose. All extracts (except Samanea saman flower ) showed antibacterial activity against two strains of methicillin resistant Staphylococcus aureus (MRSA) with MID values ranging between 100 μ g/disc and 500 μg/disc.

Conclusion: The potential source of antioxidant and antibacterial agents, especially for MRSA infection treatments were found in B. kockiana, C. pulcherrima , C. tergemina and P. pterocarpum . These preliminary results would be a guide in the selection of potential candidates for further pharmacological study and in search of new drug

candidate in treating MRSA infections.

Background

Natural products and secondary metabolites formed by living systems, notably from plant origin, have shown great potential in treating human diseases such as cancer, coronary heart diseases, diabetes and infectious diseases [1]. According to World Health Organisation,

65 - 80% of the world populations rely on traditional medicine to treat various diseases [2]. To date, many plants have been claimed to pose beneficial health

effects such as antioxidant and antimicrobial properties. With the emergence of multiple strains of antibiotic resistance microorganism , great interest has been generated in search for potential compounds from plants for therapeutic, medicinal, aromatic and aesthetic uses [2,3].

* Correspondence: goh.joo.kheng@sci.m onash.edu.my

1

School of Science, Monash Universi ty Sunway Campus, Bandar Sunway,

46150 Petaling Jaya, Selangor, Malaysia

Full list of author information is availabl e at the end of the article

Chew et al . BMC Complementary and Alternative Medicine2011, 11:12

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© 2 0 1 1 C he w e t al ; li ce ns e e B i oM e d C e nt ra l Lt d. T hi s i s a n O p e n A cc e s s ar ti cl e di st ri bu t e d u n de r t he t e rm s of th e C re a ti ve C o m m on s

At tr ib u ti on L ic e ns e ( h tt p: // c re a ti ve c om m o ns .o r g/ li ce ns e s /b y/ 2 .0 ), w hi ch p e rm i ts u nr e s tr i ct e d u s e , di s tr i bu t io n, an d re p ro d u ct io n in

a ny m e d iu m , p ro v id e d t he or ig in a l w or k i s p ro p e rl y ci te d .

Phytochemicals are natural and non-nutritive bioactive compounds produced by plants that act as protective agents against external stress and pathogenic attack [4].

Secondary metabolite is crucial for plant defenses ( e.g. as an antioxidant or antimicrobial agent) which has enabled plants to survive . Based on their biosynthetic origin, phytochemicals can be divided into several categories: phenolics , alkaloids, steroids, terpenes, saponins,

etc. Phytochemicals could also exhibit other bioactivities such as antimutagenic, anticarcinogenic, antioxidant,

antimicrobial, and anti-inflammatory properties [5].

These plant-derived phytochemica ls with therapeutic

properties could b e used as single t herapeutic agent or

as combined formulations in drug development.

Phen olic is one of the majo r groups of phytoch emica l

th at ca n b e fo un d ub iq uit ou s ly i n ce r ta i n p la nt s. P he no -lic compounds are potent a nt ioxidants a nd free radical

scaven ge r w hich can act as hy drogen donors, r educing

age nts, m etal chela t ors and sin gl et o xygen q ue nc he rs

[4]. Studies h ave s hown that ph enol ic compounds such

as ca tec hi n and qu e rce tin we re ver y eff icient i n s tabilis-ing phospholipid b ilay ers against p eroxidation i nduced

by reactive oxygen species (ROS) [6,7].

In Malaysia, only 5 - 15% of more than 250000 species

of higher plants with therapeutic p ot ential have been

studied [8]. H ence, the re i s a vast potential t o r eveal

plant resources with useful phytochemicals [9].

Th e b ioactivities of nine medicinal plants from Legu-minosa e f amil y w ere a ssessed i n t his report. T he Le gu-minosae f am ily c omprises o f 65 0 gene ra a nd more than

18000 species [9]. Some of these plants wer e reported as

medicinal p lants, such as B. kock iana, B. pu rpu r ea,

C. pulcherrima, C. surattensi s which have been used tra-dit ion al l y to tre at v ari ou s di se ase s. Al so, t hes e me di ci nal

plants showed various bioac tivi ties such as anti-cancer,

anti-inflammatory, antimicrobial and antioxidant proper-ties. F or example, Glycyrrhiza u ralensis(kan cao ) root

ethano lic extr acts has chemo prevent ive proper ties where

it was f ound to induce apoptosis in MCF-7 breast can-cer cells [10];Trigonella fo enum-graecum(fenugreek)

seed extract ha s shown to have hypoglycaemic and

hypocholesterolemic p rope rties [ 11]; isoflavo noids iso-lated from Erythrin a v ariegata significantl y inhibi t the

gro wt h of MRSA [12] ; a nd Vicia f aba (broad bean)

showed antimicrobial activity a gainst bacteria such as

Escherichia c oli , Shigella sp., Bacillus s ubt ilis an d Sta-phylococcus aureus [13].

The main a ims o f t his report a re to ev aluate antioxi-dant capacity (total p henolic content and free radical

scavenging ability), antibacterial activity and to screen

for ph ytoch emica ls co nten t in flo wers and l eaves of ni ne

me di ci nal p la nt s fr o m Leg um i no sae fam il y, na me ly Aca-cia auriculiformis , Bauhinia kockia na, Bauhinia

purp urea, Ca esa lpi nia pulch er ri ma , Cal lia ndra terge-mina, Cassia surattensis, Leucaena leucocep hala, Pelto-p ho ru m p te ro c ar pu m ,andSa ma ne a s am a n.To the best

of our k nowledge, this is the first repo rt on a nt io xid a nt

an d an t i b ac t er i al ac t i v i ti es o f m os t f l ow er s an d l ea v e s o f

selected Leguminosae species.

Me thods

Plant materials and preparation of extracts

Flowers and leaves of plants of interest were collected

from Klang Valley in Peninsular Malaysia and w ere

identified by plant taxonomist, Mr. Anthonysamy Savari-muthu. The plant materials were collected on the day

when extrac tion was p erformed. Voucher specimens

(M UM- L EGU M-0 01 - MU M-L E GU M- 00 9) wer e de p os-ited in the h erbarium of School of Science, Monash

University Sunway campus.

Antioxidant activity

Extraction of flowers and l eaves was performed a s

descr ib e d b y C hew e t a l. [4] . 1 g o f each s am ple was

frozen with liquid nitrogen and was ground in a mor-tar. The powdered samples were then extracted with

50 mL 75% v/v metha nol a nd w ere sh ook continuously

for an h our at room temperature. The e xtracts w ere

f iltered unde r vac uum an d ke pt at -2 0°C u ntil f u rther

anal ysis.

Antibacterial activity and phytochemical screening

2 5 g o f f resh pla nt m ate r ia ls w ere c o lle ct ed a nd d ried

for 1 day using freeze dryer. The dried sam ples were

gro und into fine powder using h ousehold blender. The

powdered materials w ere e xtracted with 500 mL o f

dichloromethane:methanol (1:1) m ixture with c ontinu-ous stirring f or 3 d ays at room tempe rature. Then, the

extracts were vacuum filtered, and the plant residues

were extra cted again with 500 mL of methanol for 1 day

at room temperature, and t he extracts were filtered

again. Both extracts were c ombined a nd evaporated

under v acuum. T he crude ex tracts we re stored i n dark

at -20°C until further analysis.

Total phenolic content (TPC)

The t otal phenolic cont en t (TPC) of extrac ts wa s mea -sured using Folin- Ciocalteu method as described pre-viously [ 4]. 1.5 mL Folin-Ciocalteu’ s p h e nol r ea ge nt

(Sigma) ( 10% v/v) and 1.2 mL 7.5% w / v Na

2 CO3

were

added to 0 .3 mL sample extract. The reactio n mixture

was thoroughly mixed and was incubated in the dark for

30 minutes before the ab sorbance was measured at

765 nm. TPC was expressed in t erms of mg gallic acid

eq uiva l ent s (G AE ) pe r 100 g fr esh m ate ri al . T he cal ibra -tion equation fo r gallic acid w as y = 0.0111 x - 0.0148

(R

2

= 0.9998) , whe rex is the gallic acid concent ration in

mg/L and y is the absorbance reading at 765 nm.

Chew et al . BMC Complementary and Alternative Medicine2011, 11:12

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2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical

scavenging assay

D P PH r ad ic al sc av e ng i ng as s ay wa s me as u re d ac co r di ng

to Chew et al. [4]. 1 mL of extract with various dilutions

was a dded to 2 mL o f DPPH (Sigma) ( 0. 15 mM in

methanol). The reaction m ixtures w ere incubated for

3 0 mi nu te s a nd th e a bsorba nc e m ea su red a t 51 7 nm.

The DPP H sca veng i ng abi lity was calc ulat ed as IC50

an d

expressed in m g ascorbic acid (AA) e quivalents per

100 g of fresh material (AEAC) as follows:

AEAC (mg AA/100 g) =

IC

50(ascorbat e)

/IC

50(samp le)

×10

5

IC

50 (ascorbate)

of 0.00382 mg/mL was used in AEAC

calculation.

Antibacterial activity

Bacteria strains

Eight speci es (G ram positive and Gram negative) of bac-t er i a we r e us ed . T h e G r am p os i t i v e b ac t e ri a w er e Ba cil-lus cereus ATCC 14579, Micrococcus luteus ATCC

4698, methicillin-sensitive Staphylococcus aureus

(MSSA) ATCC 2 5923, a nd t wo strains o f m eth i cillin-resistant S. aureus(MRSA) (ATCC 33591 and clinical

strain). The G ram negative bacteria include Esch er ichia

coli ATCC 25922, Klebsiella pneumoniae ATCC 10031 ,

Pseudomonas aeruginosa ATCC 10145, and Enterobacter

aerogenes ( ob t ai ne d fr o m I ns t i t ut e fo r Me di ca l R es ea rc h

Mala ysia). All b ac teria w ere c ulture d o n nutrie nt agar

(Oxoid).

Preparation of inoculums

Onesinglecolonyofeachtypeofmicroorganism(from

the nutrient agar stoc k culture) was i noc ulated with a

sterile l oop, a nd was transferred into 1 0 mL sterile

nutrient bro th ( Oxoid). The brot h cult ur es were incu-bated in a shaking incubator at 37°C for 16 - 20 hours.

Antibacterial susceptibility test: Disc diffusion assay

The antibacteri al a ssay was p erformed using d isc d iffu-sion (Kirby-Ba uer) m ethod [ 1]. T he density o f b acteria

was standardized using McFarland 0.5 turbidity standard

to 1 × 10

8

coliform units (c fu)/mL and wa s swabbed

onto Mueller Hinton Agar ( Oxoid) surface. 1 mg o f

cr ud e ex t r ac t wa s d i ss o lv e d i ni t i al l y i n 10 0 μ L m et ha no l

and loaded onto s ter ile Whatman No. 1 filter paper

discs (6 mm diameter) and t he discs were impregnated

onto in oc ulated agar. 10 μ g of streptomycin o r 30 μ g

vancom ycin (b ot h fro m Ox oid) and b lank d is c with out

extract were s erve d as positive and negative controls,

respectively. The plates we re l eft at 4 °C f or an hour to

allow t he dif fusion o f ex trac ts b efore t hey were i nc u-bated for 16 - 20 hours at 37°C. A nt ibacterial activity

w as i ndic ate d w he n cle a r in hi bition zone s w ere n oted

aroun d the discs. T he diameter of the i nhibition zone s

was measured a nd the r esults were expressed a s mean

of three independent experiments. The test was repeated

three times.

Determination of minimum inhibitory dosage (MID)

Mini mum inhibitory d osage ( MID), m inimum dose per

disc required to inhibit growt h of microorganism was

p er fo rm ed o n ex tr ac t s wh i c h sh o we d po s i ti v e ac t i v i t y i n

the p reliminary screening. MID w as determined as

described by H ab sah e t a l. [14]. Seri al d ilut ions o f

ex t r ac ts fr o m 0. 01 - 1 m g pe r di s c we r e lo ad ed o nt o fi l -ter paper discs according to disc diffusion method

described above.

Phytochemical analysis

Phytochemical screening for f lavonoids, tannins, sapo-nins and alkaloids wa s determined as described by Par-ek h a nd Chan da [15] a nd Aiy egoro a nd Okoh [1 6], b ut

the amount o f extra ct used w as inc reased for b etter

results visualisation. 1 g of extract was dissolved in

10 mL dH

2 O and was t hen filte red. 10 mg magnesium

tur nings were added into 1 mL of the f ilt rate, follo w ed

b y t he a dd i t i o n o f 0. 0 5 mL co n ce nt r at e d s ul p h u ri c a ci d .

The presence o f m ag enta red o bserved within t hree

m i nu t es c o nf i r m ed t h e pr e s en c e o f fl a v o no i ds . T he p r e-se n ce of b l ue - b l ac k pr ec i pi t at es re s ul ti ng fr o m t he ad di -t i o n o f f er r i c ch l o r i d e ( 0. 0 1 g / m L ) r e ag en t i nd i c at ed t h e

presence of tannins. Frothing test was used to determine

the presen ce of saponin s, where 2 mL of the extra ct was

shook v igorously for 4 minutes and the presence of

honey-comb froth i ndicated the e xistence of saponins.

The presence of alkaloids was determined by first dissol-ving 0.02 g of extract in 1 mL methanol and filtered, fol-lo wed by bo ili ng the e xtract wit h 2 mL of 1 %

hyd ro ch loric ac id fo r 5 minutes . 4 - 6 drops of Dragen-dorff’ s reagent was then added i nt o the extract, and the

formation o f o range p re cipitates i ndicated the p resen ce

of al ka loids. Screeni ng the pres ence of steroids and ter-penoids was performed as described by Kumar et al.

[17]. 0.2 g o f extract was d issolved in 10 mL methanol

an d f ilte re d. 1 mL of chloroform a nd 1 mL of conc en-t ra t ed su lp hu r i c ac id we re t he n ad de d i nt o 1 m L fi l t ra t e

by the side o f the tube and t he presen ce of yellow with

gre en f luorescenc e at the sulphuric acid lay er indicated

the presen ce of steroid s. For the detecti on of terp enoids,

1 mL of a cetic anhydr ide and 2 mL concentrated sul-phuric acid were added i nto 1 mL filtrate. Presence o f

reddish bro wn on interface i ndicated the presence o f

terpenoids.

Statistical analysis

A ll me as ur em ent s wer e car r ie d o ut i n tri pli cat e. S ta tis t i-ca l an al ys es we re pe r fo r me d us in g a o ne- wa y an al ys i s o f

variance ANOVA, and the significant difference between

m ea n s w as d et er m i ne d b y Du nc an’ s m u l ti pl e r an g e t e st .

Differences at P < 0 .05 were c onsidered st at istically

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sig ni fic ant . The re sul ts wer e pre se nte d as me an v al ues ±

SD (standard deviations).

Resu lts

TPC and DPPH scavenging activity

TPC of fl owers can b e ranked a s f ollows usi ng Fol in

Cio calt eu ass ay: B. koc kia na > C. sur att ensi s > C. p ulch e r-rim a> A . au ric ul ifor m is≥ C . t erg emin a > P. pte roc arp um

≥ L. leucocephala > S. saman > B. purpurea. Slightly dif-ferent ranking w as observed in TPC of the leaves:

C. pulch e rrima h a s t he high est TPC (503 0 ± 602 mg

GAE/100 g), followed by P. pterocarpum (4880 ±

27 5 m g G A E/ 10 0 g) , B. k o c k ia na ( 42 20 ± 10 4 mg GA E /

100 g), C. te rge mina (4200 ± 292 mg GAE/100 g),

C. sura ttensi s (3330 ± 309 mg GAE/100 g), A. a ur icul i-formis (22 80 ± 294 mg GAE/100 g), L. leucocephala

(1700 ± 277 mg/GAE/100 g), S. saman (1340 ± 22 mg

GAE/100 g) and B. purpurea (1310 ± 124 mg GAE/100 g).

Statistical analysis showed no significant difference

be tw een C. pu lch e rri maan d P. pt eroc arp um, B. ko ck ia na

and C. t er ge m ina, A. a ur iculi fo rmi sand L. leucocephala ,

S. saman and B. purpurea . It was observed that L. leucoce-pha la, S. s aman and B. pu rpur ea flo wer s and leave s wer e

ranked similar, with lowest TPC.

It was f ound that free radical sc avenging activity o f

the plants w as directly proport ional to TPC (F igure 1

and Table 1), where by the higher the TPC of the

extract, t he stronger the free radical scavenging activ-ity. B. kock iana flower displayed the highest TPC

(8280 ± 498 mg GAE/100 g) a nd A EAC ( 14 6 00 ±

23 60 mg A A/1 00 g) a nd stron ge st DPPH radic al

quenching a bility, i n which 27. 0 ± 5.0 μ g/mL of extract

could e ffect ively scaven ge 50% of the free D PPH radi-cals. Whereas, B. pu rp u r e a flower consists of much

lower T PC and AEAC ( 349 ± 26 mg GAE/100 g and

23 5 ± 35 mg AA/100 g respectively), a nd hi gher con-ce ntration of extrac t was nee de d to scave nge 50% of

the free r adica ls ( 1690 ± 280 mg/mL). S trong positive

associa tion was obse rve d be tw een TPC and AEAC

(R

2

= 0.9747), w h i le TPC a nd IC

50

were inversely p ro -portional (R

2

= 0 .9753) . This shows that radical

sc av e ngi ng a c tiv ity is c lose ly asso ci at ed w ith the p he -nolic c o mpounds in t h ese plan t s.

Statistical anal ysis s howed t hat T PC and a ntio xidant

activi ty of flowers a nd leav es of the same speci es are

significantl y different. Ho wever, exceptions w er e see n

in A. auri culiformis, C. surattensis, L. leucocephalaan d

S. saman. On t he other hand, TPC o f both parts of

Figure 1 Total phenolic content (TPC) of flowe rs (F) and leaves (L) of Legumi nosae species . Result s are expresse d a s means ± SD (n = 3).

A.a. = A. auriculiformis; B.k.=B. kockiana; B.p. = B. purpurea; C.p. = C. pulcherrima ; C.t.= C. tergemina; C.s.=C. surattensis; L.l. = L. leucocephala ;

P.p. = P. pterocarpum ; S.s. = S. saman .

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S. saman we re shown to be statistically signific ant but

th ey w ere not s ignif ica nt in terms o f an t ioxi da nt

activity.

Antibacterial activity and minimum inhibitory dosage

(MID) of extracts

The susceptibility of bacteria towar ds the plant e xtract s

wa s asse ssed. Al l nin e e xtrac t s w ere t este d o n s ele cted

Gram positive and negative b ac te ria . The sc re ening

show ed most e xtra cts displa yed a b road spectrum o f

antib a ct erial a ctivity towards Gram positive but som e

had selective a ct ivity towards the bacteria, i.e. B. pu r-pur ea flower exhibited inhib itio n activity towards anti-biotic r esistant S. aureus (MRSA), but not to t he other

three Gram p ositive b acteria ( Table 2). No inhibition

was o bserved in Gram negative bacteria at the same

dosage for all the plant extracts.

Minimum i nh ibitory dosa ge (MID) was evalu ated to

assess the potency of the e xtracts i n i nh ibiting the

gro wt h of b act er ia . T he an tib ac te ria l ac ti vi ty of B. k oc k i-ana flower an d C. surattensis le af extracts w ere strong.

Most of the antibacterial activity o f t he other extrac ts

ex hib i ted me diu m to wea k in hib i ti on act ivi ty . It i s in te r-esting to observe thatB. purpurea flower exhibited med-ium inhibi tio n acti vit y towards MR SA, but i nactiv e

against the antibiotic sensitive strain (MSSA) a nd other

Gram positive bacteria. Besides, i t w as noted that

S. saman flower did not exhibit any antibacterial activities

towar ds both G ram positive an d ne gat i ve bac ter ia tested

in this study.

Phytochemical screening

It has b een repo rted that bi ologi cal activ iti es in the

selected plants were exhibited b y different class of phy-tochemicals [18-24]. T here fore , it i s important to

screen for phy tochemical group in these plants. Phyto-chem ical screening on the 18 extracts showed t he pre-sence of diffe rent type s of c he mica l constituents.

Ta nni n w as dete cted i n most of the extrac t s ( 14

extracts), followed by t erpe noid (13 extracts), s teroid

(13 extracts), f lav ono id (12 e xtracts) , a nd saponin ( 10

extracts). Alkaloid was only p resent in 4 extracts

(Table 3).

Disc ussion

Antioxidant activity

Plants p roduc e dive rse arrays of phytochemicals which

ar e us ef u l i n th e de v el o pme n t o f ne w dr ug s . T he s e p hy -toc hem ic als ar e mos tly sec onda ry me tabo lite s con st ant ly

synthesized b y t he plant for defens ive purposes [ 4] . F or

Instance , a ntioxidant s a re biologically produced as

defensive m echanism t o prevent tissues destruction

caused by highly reactive chemical species, which are

formed from various biochemical reactions.

Excessive free radicals which a re produced from var-ious biochemical reactions such as triplet chlorophyll,

single t o xy gen and hydroxyl ra di ca ls a r e l ethal t o

plant s. One of the m aj or cl asses of na t ural antioxidan ts

found in p lants t hat r emove such free r adica ls is p oly-phenols. P o lyphenol s are able to neutrali ze free radi-cals, scavenge singlet and triplet oxygen, and t o b reak

down pe roxides. Total p olyphenol c ontent (TPC) of

plants was evaluated using Folin-Ciocalteu method,

which measured the r edox properties of polyphenols i n

extracts [4].

The amount o f T PC of se lected plan ts could be cate-gorised i nto 4 classes: high (>5000 mg GAE/100 g),

mediu m hi gh (3 000 - 5 00 0 m g G AE/1 00 g), me d ium

low (1000 - 3 00 0 mg GAE/100 g) and l ow (<1000 mg

GAE/10 0 g ). Ac cordin g t o K ue te an d Eff erth [2 5], fre e

radical scavenging ability was also ranked according to

IC

50

: hig h (< 50 μ g/m L ); mod er at e ( 50 - 100 μ g/ mL) and

low (>50 μ g /mL). Most o f t he e xtrac t s s howed medium

level of TPC (med ium high: 5 ex trac ts; med ium low: 1 0

extrac ts). It w as notic ed tha t most l eav es had hi gher

TPC than the flowers of the same species, i.e. TPC of

B. pu rpurea, C. pulche rrima, C. terg emina, P. pterocar-pum an d S. saman lea ves were significant ly higher than

their respective f lower s (Figure 1 ). Although TPC o f

C. surattensisflower w as found to be higher than its

leaves, t hey we re s tatistica lly not significant. From this

interesting o bservation, i t c ould be explained b y t he

Table 1 Free radical scavenging activity of flowers (F)

and leaves (L) of Leguminosae species

Plants Part Antioxidant activity

IC

50

( μg/mL) AEAC (mg AA/100 g)

A. auriculiformis F 152 ± 13

a

2540 ± 221

a

L 161 ± 30

a

2430 ± 457

a

B. kockiana F 27.0 ± 5.0

a

14 600 ± 2360

a

L 61.0 ± 10.0

b

6410 ± 985

b

B. purpurea F 1690 ± 280

a

235 ± 35

a

L 384 ± 45

b

1010 ± 122

b

C. pulcherrima F 118 ± 24

a

3350 ± 618

a

L 50.0 ± 5.0

b

7690 ± 735

b

C. tergemina F 179 ± 12

a

2170 ± 130

a

L 61.0 ± 6.0

b

6300 ± 610

b

C. surattensis F 96.0 ± 16.0

a

4080 ± 728

a

L 94.0 ± 11.0

a

4130 ± 463

a

L. leucocephala F 289 ± 17

a

1340 ± 78

a

L 187 ± 15

b

2070 ± 156

b

P. pterocarpum F 189 ± 10

a

2040 ± 120

a

L 51.0 ± 0.0

b

7630 ± 331

b

S. saman F 343 ± 43

a

1130 ± 153

a

L 384 ± 39

a

1000 ± 97

a

Results are expr essed as mea ns ± SD (n = 3). For each col umn, valu es

foll owed by the same lette r (a-b) are statis tica lly ins ignifican t ( P > 0.0 5) as

dete rmine d using ANO VA, and this does not apply for differe nt plant s.

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Table 3 Classes of phytochemicals present in the plant extracts

Plants Part Tannin Terpenoid Alkaloid Steroid Saponin Flavonoid

A. auriculiformis F- + + - + - L- + - + + - B. kockiana F+ - - + - + L+ - - + - + B. purpurea F- + + + + + L- + - + + - C. pulcherrima F+ + - + - + L+ + - + - + C. tergemina F+ + - + + + L+ + - + + + C. surattensis F+ + - - - + L+ - - - - + L. leucocephala F+ - + - - + L+ - - + + + P. pterocarpum F+ + - - + - L+ + - + + - S. saman F+ + + + + + L+ + - + - -Note:“+ ” = pr esent; “-” = abs ent.

Table 2 Antibacterial activity of flowers (F) and leaves (L) of Leguminosae species

a

evaluated using disc diffusion

assay at 1 mg per disc

Bacterial strains

Diameter of zone of inhibition (mm)

Plants Part B. cereus ATCC

14579

M. luteus ATCC

4698

MSSA ATCC

25923

MRSA ATCC

33591

MRSA (clinical isolate)

b

A. auriculiformis F 10.0 ± 0.0 11.8 ± 0.4 11.8 ± 0.4 15.3 ± 0.4 13.3 ± 0.4

L 9.50 ± 0.7 12.0 ± 0.0 12.0 ± 0.0 15.3 ± 1.1 13.5 ± 0.7

B. kockiana F 10.0 ± 0.0 12.0 ± 0.0 12.5 ±0.7 11.0 ± 0.0 11.5 ± 0.7

L 8.00 ±0.00 10.0 ± 0.0 9.50 ± 0.70 9.00 ± 0.00 10.0 ± 0.0

B. purpurea F - - - 12.3 ± 0.4 10.0 ± 0.0

L 7.00 ± 0.00 8.30 ± 0.35 7.00 ± 0.00 8.00 ± 0.00 -C. pulcherrima F 7.80 ± 0.40 10.0 ± 0.0 10.0 ± 0.0 9.50 ± 0.70 9.50 ± 0.70

L 7.80 ± 0.40 10.0 ± 0.0 10.0 ± 0.0 10.5 ± 0.7 9.50 ± 0.70

C. tergemina F 7.00 ± 0.00 7.50 ± 0.70 8.50 ± 0.00 9.30 ± 0.35 8.50 ± 0.70

L 10.3 ± 0.4 11.0 ± 0.0 12.0 ± 0.0 13.0 ± 0.0 11.0 ± 0.0

C. surattensis F 7.50 ± 0.70 10.0 ± 0.0 10.0 ± 0.0 11.0 ± 0.0 10.0 ± 0.0

L 7.50 ± 0.70 10.0 ± 0.0 10.0 ± 0.0 10.0 ± 0.0 10.0 ± 0.0

L. leucocephala F - 9.30 ± 0.40 8.00 ± 0.00 11.0 ± 0.0 8.80 ± 1.10

L - 9.00 ± 0.00 9.30 ± 0.40 9.30 ± 0.40 10.0 ± 0.0

P. pterocarpum F 8.00 ± 0.0 11.3 ± 0.4 9.80 ± 0.40 10.8 ± 0.4 10.3 ± 0.4

L 9.80 ± 0.40 12.0 ± 0.0 11.5 ± 0.7 11.8 ± 0.4 10.8 ± 1.1

S. saman F- - - - - L - 21.3 ± 0.4 8.50 ± 0.70 9.00 ± 0.00 9.30 ± 0.35

Streptomycin (10 μ g/

disc)

24.0 ± 0.0 27.0 ± 0.0 14.0 ± 0.0 n.t n.t

Vancomycin (30 μg/disc) n.t n.t n.t 21.0 ± 0.0 21.0 ± 0.0

Note: diam eter of the antib acter ial disc is 6 mm. Results are expres sed as mea ns ± SD ob tained from 3 indep endent experi ments.

a

All extrac ts wer e inact ive

agai nst Gram negat ive bacteria ( Esc herichia coli ATCC 25922 , Kle bsiell a pneumon iaeATCC 10031 , Pseudo monas aerugin osa ATCC 10145 , Enter obacte r aer ogenes

obtai ned from Institut e for Medi cal Resea rch Mala ysia) tested.

b

Obtained from Fac ulty of Medici ne and He alth Scien ces UPM.“- ” indic ates no inhib ition act ivity.

“n.t” rep resents not tested.

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diff erenc es i n antioxidant distribution in the l eaves and

flowers. Similar observation was discovered in Cassia fis-tula [26] and Cyn ara card uncu lus [27] . Si ddhu raj u et al .

[26] report ed t he pro duct ion of a nt hraquinones,