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Research Article ​ISSN: 2395 -5775 ​PHYTOCHEMICAL PROFILING,
ANTIOXIDANT AND ANTIMICROBIAL ACTIVITY OF
METHANOL EXTRACT IN RAMBUTAN FRUIT (NEPHELIUM
LAPPACIUM) EPICARP AGAINST THE HUMAN PATHOGENS
Nethaji R, Thooyavan G, Mullai Nilla K and Ashok
K*

Abstract
A large number of the plants are claimed to possess the antibiotic properties in the traditional
system and are also used extensively by the tribal people throughout the world. It is now mostly
thought that nature has given the cure of every disease in one form or another. Plants have been
known to cure people from various diseases in Ayurveda. Effects of crude extract of the ​Nephelium

lappaceum ​epicarp were studied ​for the antioxidant and antibacterial activity. The research showed

that the methanolic extract were more effective.

​ The antioxidant activity of the methanol extract
were carried out using DPPH which showed better antioxidant properties which could be very useful

against reactive oxygen species that are formed ​during oxidative stress. Key
​ Words:
Phytochemical profiling, antimicrobial activity and antioxidant activity
Dur-India) as a health food supplement selling at
INTRODUCTION US$9 for a dozen bottles, each containing 63
ablets [1]. Several medicinal investigations on the
Rambutan fruit is one of the most interesting of all
validity of ​this belief have been conducted with
tropical fruits widely known and revered in South
conclusion [2-3].
East Asia as the” King of Fruits”. The fruits arevarying ​
distinctive for its large size, unique odor and a
The custard-like flesh has an exquisite flavor and is
formidable thorn covered hersk. The custard-like
at the same time aromatic and sweet with a
flesh has an exquisite flavor and is at the same
strange ​balsamic taste. Rambutan has been
time ​aromatic and sweet with a strange balsamic
taste.Rambutan has been claimed that the fruitclaimed that the fruit ​ possess great rejuvenating
possess great rejuvenating power. The Rambutanpower [4]. Traditional systems of
fruits from the genus D ​ urio ​belonging to themedicine,ayurveda were the basis of the health
Durionaceae family. In the 1920s, Rambutan fruit
care system in India until early ​years of twentieth
products, Inc.,of New York City launched a product
century.According to an estimation of ​ world health drugs are
​ prepared from excretory plant product
organization approximately 80% of people in such as gum, resin and latex. Not only, that
developing countries rely on traditional plant-derived drug offers a stable market worldwide,
medicinesfor primary health care needs but also plants continue to be an important source
[5].
for new ​drugs [8]. The scientific study of traditional
Rambutan fruit is one of the most interesting of all
medicines, derivation
​ of drugs through bio
tropical fruits widely known and revered in South
prospecting and systematic conservation of the
East ​Asia as the” King of Fruits”. The fruits areconcerned medicinal plants are thus of great
important. Many phytochemical found in fruits act
distinctive ​ for its large size, unique odor and a
formidable thorn covered hersk [6]. as powerful ​antioxidants that give them color,

protection against human diseases.

The presumption of world health organization is thatflavor, odor and ​
Rambutan fruits were used as good sex tonic and
the edibles, particularly of plant derived,its fruit sap is very useful in treating more than 50
empathetically indicates that these items have
common diseases. But ​there is no scientific study
medicinal values [7]. The drugs are derived from
the whole plant or from different organs, like leaves,
on the antimicrobial activity of
​ the extracts of the
Rambutan fruits [9].
stem, bark, root, flower, fruits, seed e ​ tc​. Some

Zoology, Presidency College, Chennai- 600 005, Tamil Nadu, India. ​Correpondence and Reprint Requests: ​Ashok K ​Received: ​Se p te mbe r 25 , 2015 |
Accepted: ​November 5, 2015 | ​Published Online: ​November 28 , 2015 This is an Open Access article distributed under the terms of the Creative
Commons Attribution License (creativecommons.org/licenses/by/3.0) ​Conflict of interest: ​None declared | ​Source of funding: ​Nil
Ashok K et al.,
Nephelium lappaceum ​Linn., commonly known as rambutan which belongs to the family of
Sapindaceae, is an attractive tropical fruit widely distributed in South-East Asia, especially in

Malaysia ​and Indonesia [10]. It is a potential fruit to be commercialized

​ since it is widely
planted all over Malaysia. Rambutan has been used as traditional medicine for centuries

especially as a remedy for diabetes and high blood pressure [11-12]. Rambutan ​contains a

large variety of substances possessing antioxidant

​ activity, such as vitamin C, vitamin E,
carotenes, xanthophylls, tannins and phenolics [13]. They also exhibit a wide range of

biological activities including antimicrobial, anticarcinogenic as ​well as antiproliferative, and

many other biological activities

​ have been attributed to their antioxidant properties [14].

Medicinal plants represent a rich source of ​antimicrobial agents. There is also an urgent need

to
search for new antimicrobial compounds with novel mechanisms of action because there has
been an alarming increase in the incidence of new and re- emerging infectious diseases [15].

A numeral ​commercial variety of rambutan (​Nephelium lappaceum ​Linn.) exists. In Malaysia,

red and yellow rambutan

​ fruits are available in the market. The fruits are ovoid, with a red or

yellow pericarp covered with soft spines that vary in colouring from yellow ​and red. They are

different in taste. However, so far there

​ is no comparative study have been reported in

rambutan varieties, though various extracts of ​Nephelium lappaceum ​have been reported for
antibacterial activity [16-17].
The main target of our study is finding of phytoconstituents from epicarp methanol extract and
Table1 Phytochemical analysis of ​Nephelium lappaceum ​epicarp methanolic extract

to determine the antioxidant and antimicrobial ​efficacy of ​Nephelium lappaceum ​epicarp

Nephelium lappaceum
Phytochemicals ​
methanol S.No.
​
​
extract against the human pathogens. ​epicarp methanolic extract 1 Carbohydrates
​
Present
Molisch’s test ​

MATERIALS AND METHODS ​2 Alkaloids

​
Dragendorff’s test Steroids and sterols
Present

Collection and identification of Nephelium lappaceum ​3

Liebermann Burchard’s and Salkowski ​test Fresh
​ Rambutan fruits (​Nephelium lappaceum​) were

purchased from Koyambedu fruit market, Chennai, ​Tamilnadu, India and were authentically

identified by Prof.
​ P. Jayaraman, Institute of Herbal Science, Plant Anatomy Research Centre,
West Tambarm, Chennai, India, as Sapindaceae with voucher specimen no:
PARC/2015/2126.
Extraction
Nephelium lappaceum ​epicarp were cut in 1⁄2 inch pieces and dried in air seven days and
finally in mechanical drier at 60-70oC. The dried samples were ​grinded to course powder with

a mechanical grinder and

​ extracted with methanol for 7 days with occasional shaking in a
beaker. The extract was filtered and the filtrate was dried at 50 to 60°C and the yielded
percentage was calculated. ​Present
Glycosides
4​
Present ​ Flavonoids ​ Present ​ Triterpenoids ​ Present ​ Tannins ​
Baljet’s test ​ 5​ Shinoda test. ​ 6​ Tin and thionyl chloride test ​ 7​ Lead acetate
Present ​ Proteins and aminoacids ​ Present
test and gelatin test ​ 8​ Biuret test and Ninhydrin ​
Table 2 Antibacterial activity of ​Nephelium lappaceum ​epicarp methanolic extract
 Name of the
S.No. ​
Name of the bacteria
extract ​

Zone of inhibition in mm 10 ​μg/ml 15

​ ​μg/ml 20
​ ​μg/ml 25
​ ​μg/ml 10
​ μg/ml ​Norflaxin 10
​ μg/ml ​Penicillin ​1
Metanol
E.coli ​9mm 10mm 12mm 14mm 18mm 11mm 2 ​Klebsiella pneumonia ​8mm 9mm 11mm 13mm 18mm 0mm 3 ​Pseudomonas
auerogenousa ​0mm 0mm 0mm 0mm 18mm 13mm ​4 ​Proteus vulgarius ​9mm 11mm 13mm 15mm 19mm 11mm
​ picarp methanolic extract
Table 3 Antifungal activity of ​Nephelium lappaceum e
Name of ​
S.No. ​ the
extract

Zone of inhibition in mm 10 ​μg/ml 15

​ ​μg/ml 20
​ ​μg/ml 25
​ ​μg/ml Ketconazole
​ ​10 μg/ml Nyastin
​ ​ Methanol ​A. fumigatus ​10mm
​10μg/ml 5
11mm 13mm 15mm 7mm 0mm

Table 4 Percentage of scavenging activity of ​Nephelium ​lappaceum ​epicarp methanolic extract Concentration
​
(μg/ml) Name
​ of the ​fungi

% of scavenging activity ​Ascrobic acid Methanolic extract 0

​ 0 0 ​1 35.26 34.789 10 41.632 38.061 20
​ 45.98 44.162 ​30 55.252 57.509 40

56.292 55.87 50 60.59 62.49

202 International Journal of Current Innovation Research, Vol. 1, Issue 9, pp 201-206, November 2015
Phytochemical Profiling, Antioxidant And Antimicrobial A
Phytochemical analysis
The phytochemical analysis of ​Actinidia deliciosa ​the fruit extracts were carried out to identify
constituents, using standard pytochemical method described by Gomathi et al., 2012 [18].

screening involves detection of second metabolities such as carbohydrates, alkal ​steroids and

sterols, glycosides, flavon triterpenoids,

​ tannins, proteins and aminoacids.
Fig1 Antimicrobial activity of ​Nephelium lappaceum ​methanolic extract (1A- E
​ .coli​, B- ​Klebsiella pneumonia ​Pseudomonas

auerogenousa​, D- ​Proteus vulgarius 10μg/ml,

​ b- 15μg/ml, c-20 μg/ml, d-25 μg/ml, e

μg/ml), f- Nyastin (10 μg/ml) ​Test microorganisms

The test microorganisms used for antimicro analysis were clinical isolates of ​pneumonia, E.

coli, Pseudomonas auerogeno Proteus vulgaris​, and ​Aspergillus fumigates bacterial

​
strains were maintained on Nutrient A and fungi on Sabouraud Dextrose Agar at Medical

Education Department, King Institute ​Preventive Medicine and Research, Guindy, Chen India.
​
Fig 2 ​Radical scavenging activity of ​Nephelium lappaceum
methanolic extract ​In vitro antimicrobial activity

Determination of zone of inhibition by disc diffusio ​method

In vitro ​antibacterial and antifungal activities wer
International Journal of Cur

oxidant And Antimicrobial Activity of Methanol Extract In Rambutan Fruit (Nephelium L ​Against The Human

Pathogens
 Actinidia deliciosa ​of ​carried out to identify the ard
​ pytochemical methods as i et al., 2012

[18]. The detection of secondary carbohydrates, alkaloids, ​s, glycosides, flavonoids, oteins
​
and aminoacids.

ms used for antimicrobial al isolates of ​Klebsiella ​Pseudomonas auerogenousa​, ​Aspergillus

fumigates​. The maintained

​ on Nutrient Agar aud Dextrose Agar at the partment, King Institute

of ​d Research, Guindy, Chennai,

f inhibition by disc diffusion
antifungal activities were

examined for methanol extract ​lappaceum ​epicarp. Antibacterial activities

​ of methanol extract

lappaceum ​epicarp against patho (Gram-positive

​ and negative investigated by the agar disk

diffusi Antimicrobial activity testing was carr agar cup method. Purified methano ​dissolved in

dimethyl sulfoxide, sterili using

​ sintered glass filter, and stored determination of zone of

inhibitio positive, Gram-negative, and fung ​taken as a standard antibiotic for co results.
​ All the

extracts were scre antibacterial and antifu the ​Escherichia coli, ​Pseudomonas aeruginosa,
fungi ​Aspergillus fumigatus ​(10, 15, 20 and 25​μ​g/ml) of epicarp methanol and standard
prepared in double-distilled water ​ephelium lappaceum ​epicarp ​Klebsiella pneumonia​, C- ​teus vulgarius​, E- ​A.

fumigatus​), (a- ​agar tubes. Mueller-Hinton sterile ag seeded

​ with indicator bacterial strai allowed to
stay at 37°C for 3 ​25 μg/ml, e- Ketconazole (10 tin
​ (10 μg/ml)

experiments were carried out under by

​ using norfloxacin and penicillin f activity and
Ketconazole and Nysati activity as standard drugs. The zo ​inhibition around the disks were

​ hours of in incubation at 37°C f ​48 to 96 hours for fungi at 28°C. Th the

meas 24 ​
microorganism species to the met ​Nephelium lappaceum ​epicarp measuring the sizes of

inhibitory zon diameter

​ of disk) on the agar surf disks, and values < 8 mm were co active
against microorganisms.
Antioxidant Activity using radical

The antioxidant activity of ​Nephelium lappaceum antioxidant

​ ascorbic acid was assess of the

radical scavenging effect of ​diphenyl-1-picrylhydrazyl (DPPH) activity

​ according to the method
Chanwitheesuk ​et al​., 2005 [20] extract with different concentrations 40 and 50 ​μ​g/ml) were
 prepared Ascorbic acid was used as the sta ​μ​g/ml solution. 0.004 % of DPP prepared in

methanol and 5 ml of t mixed with 5 ml of methanol extra ​standard solution distinctly. These s
ournal of Current Innovation Research, Vol. 1, Issue 9, pp 201-206,
Nephelium lappaceum e​ picarp
ambutan Fruit (Nephelium Lappacium) Epicarp
for methanol extract of ​Nephelium ​Antibacterial and antifungal f methanol extract of
Nephelium ​against pathogenic bacterias ive and negative) and fungi were by the agar disk
diffusion method [19]. l activity testing was carried out by using ethod. Purified methanol

extracts were ​dimethyl sulfoxide, sterilized by filtration lter,

​ and stored at 4°C. For the n of

zone of inhibition, pure Gram- negative, and fungal strains were ​tandard antibiotic for

comparison of the the

​ extracts were screened for their l and antifungal activities against ​hia

coli, Klebsiella pneumonia, s aeruginosa, Proteus vulgaris ​and the ​gillus fumigatus​. The

sets of four dilutions g/ml)

​ of ​Nephelium lappaceum ​ethanol and standard drugs were

distilled water using nutrient ​Hinton sterile agar plates were indicator
​ bacterial strains (10​8​cfu)

and stay at 37°C for 3 hours. Control ​were carried out under similar condition rfloxacin
​ and
penicillin for antibacterial Ketconazole and Nysatin for antifungal standard drugs. The zones of

growth ​und the disks were measured after 18 to in

​ incubation at 37°C for bacteria and ​urs for

fungi at 28°C. The sensitivities of anism

​ species to the methanol extract of epicarp were

determined by ​e sizes of inhibitory zones (including the disk)

​ on the agar surface around the
alues < 8 mm were considered as not st microorganisms.
Activity using radical-scavenging method
idant activity of methanolic extract epicarp and the standard ​ascorbic acid was assessed on

​ scavenging effect of the stable 2, 2- ​icrylhydrazyl (DPPH) free radical ording

the basis al ​ to the
method described by ., 2005 [20]. The methanol different concentrations (1, 10, 20, 30, g/ml)
were prepared using methanol. id was used as the standard in 1-50 ion. 0.004 % of DPPH
solution was methanol and 5 ml of this solution was 5 ml of methanol extract solution and

lution distinctly. These solution mixtures

November 2015 203
Ashok K et al.,
absorbance of the combination was determined
were kept in dark for 30 min. The degree of DPPH
purple decolourization to DPPH yellow indicatedat517 nm using UV-Visible Spectrophotometer and
the scavenging effectiveness of the extract. Theascorbic acid was served as a positive control.
Lower absorbance of the reaction mixture indicated
combination with ​sugar molecules. They are
higher free radical scavenging activity.
mportant in medicine because
​ of their action on
% DPPH radical scavenging = (Absorbance of
heart and are used in cardiac insufficiency [26].
control -Absorbance of test Sample) / (Absorbance
Thus, cardiac glycosides are drugs and can be
Of control) X 100
used in the treatment of congestive heart failure

RESULT AND and cardiac arrhythmia. ​They work by inhibiting the

DISCUSSION
Na​+​/Na​+ ​pump, resulting in
​ an increase in the levels
Phytochemical of sodium ions in the myocytes, which then leads to
screening a rise in calcium ions level. This inhibition raises

he amount of Ca2​+ ​ions ​available for concentration

The phytochemical screening of the methanolic
increases cardiac output and
extracts of ​Nephelium lappaceum ​epicarp showedof the heart muscle, ​
that the leaves are rich in, carbohydrates, proteins educes distension of the heart [27-28]. These
and aminoacids, steroids, alkaloids flavonoids,glycosides are found as secondary metabolites in
tannins, triterpenoid and glycosides. Flavonoids areseveral plants and animals [29]. However, some
most commonly known for their antioxidant activity.glycosides, such as ouabain, are ​
toxic as it inhibits
They are transformers which modify the body‘sactive transport of Na​+ ​in cardiac
reactions to carcinogens, viruses, and allergens. muscle (sodium pump inhibitor), which results in
They show anticancer, anti-inflammatory, inhibition of translocases during electron transport
antimicrobial and anti-allergic activity [21-22], and chain, and leading to death [30]. Also phloridzin

may be useful in ​therapeutic roles [23]. Alkaloids (toxic glycosides) blocks the transport of sugar

are organic compounds

​ that contain nitrogen, and across ​the mucosal cells of small intestine and also
are physiologically active with sedative and epithelium; it displaces Na​+ ​from the
renal tubular
​
analgesic properties. They are used in relieving binding sites of carrier protein and prevents the
pains, anxiety and depression [24]. binding of sugar molecule and produces glycosuria
[31-32].
Alkaloids are toxic due to their stimulatory effects,
leading to excitation of cells and neurological
In vitro antimicrobial
dysfunction [25]. Glycosides are compounds
activity
containing a carbohydrate and non-carbohydrate
Antibacterial activity of the methanol extract of
residue in the same molecule. In these

compounds, the carbohydrate

​ moiety is attached Nephelium lappaceum ​epicarp was tested against
by an acetal linkage of carbon-1 to the aglycone. the different test microorganisms are shown in the
They all contain steroid as aglycone component in
table 2 and3. The methanolic extract of ​Nephelium
 dosage determined for proper administration.
lappaceum ​epicarp showed higher antibacterial
property against ​Escherichia coli​, ​KlebsellaIn vitro antioxidant activity

pneumoniae ​and ​Proteus vulgaris ​but no activityDPPH test is based upon the ability of DPPH, a
stable free radical, to decolourize from purple in the
was observed in ​Pseudomonas auerogonesa ​andpresence of antioxidants. It is a direct and

in antifungal
​ activity methanol extract showed
dependable method for determining the radical
moderate zone of inhibition against A ​ spergillus
scavenging action. Ascorbic acid was chosen as
the standard antioxidant for this test. The DPPH
fumigatus ​as shown in the figure 1A,B,C and D.
radical contains an odd electron, which is
From the
​ primary phytochemical screening, weresponsible for the absorbance at 517 nm and also

have been ​able to identify that methanol extract offor a noticeable ​deep purple color. When DPPH

Nephelium lappaceum ​epicarp has gotaccepts an electron donated

​ by an antioxidant
compound the DPPH becomes colourless, which is
phytomedical property it may
​ be due to the nature
quantitatively measured from the changes in
of biologically active compounds present in it whose
absorbance. Highest scavenging ​was observed
activity are enhanced ​in the presence on methanol

stronger extractionwith N​ ephelium lappaceum ​epicarp methanolic

​
and also methanol has an ​ extract followed by Ascrobic acid known antioxidant
capacity which could have produced greater
standard.
number of active constituents responsible for
antibacterial activity aswell as antifungal activity.Scavenging activity of DPPH radical was found to

The antimicrobial activities can be ​enhanced if the

rise with increasing concentration of the methanolic
active components are purified and adequate
​

204 International Journal of Current Innovation Research, Vol. 1, Issue 9, pp 201-206, November 2015
Phytochemical Profiling, Antioxidant And Antimicrobial Activity of Methanol Extract In Rambutan Fruit

(Nephelium Lappacium) Epicarp ​Against The Human Pathogens extract

​ of ​Nephelium lappaceum
epicarp. Additionally, it has been determined that the antioxidant effect of plant products is
mainly due to radical scavenging activity of phenolic compounds such as flavonoids,

polyphenols and tannins [33]. The antioxidant activity of phenolic compounds is mainly ​due to

their oxidation reduction properties, which can play

​ an important role in adsorbing and

neutralising free radicals, reducing singlet and triplet oxygen, or ​decomposing peroxides [34].

Oxidative injury now appears

​ as the fundamental mechanism causing a ​number of human
neurologic and other disorders such as
​ autoimmune pathologies, inflammation, viral infections
and digestive system disorders including ​gastrointestinal inflammation and ulcer [35]. The
present results suggest that all the tested plant extracts have moderate to potent antioxidant

activity. Subsequently a variety of constituents are ​known from the four crude extracts which

we studied therefore,
​ it becomes very difficult to ascribe the ​antioxidant properties selectively

to any one group of

​ constituents without further studies it is impossible. Thus further thorough
investigations are necessary. ​With the increase in absorbance there is increase in reducing
​
antioxidant power. So with high amount of ​extract there is increase in absorbance and with the
decrease in extract quantity the absorbance is lowered.
CONCLUSION

In conclusion, the above results showed that ​methanolic extract of rambutan fruit epicarp are

good candidate for further investigation of ​antimicrobial and antioxidant potentials.

Furthermore, the
​ isolation of active constituents which are responsible for the activity is

necessary for broad ​investigation.

Acknowledgment
The authors are thankfull to Sterline Bioscience, Chennai for the technical support.
Conflicts of Interests
The authors declare no conflict of interest
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