The Study of the Antibacterial Activity of Asam Gelugor (Garcinia

Atroviridis) Against Methecillin-resistant Staphylococcus Aureus

(MRSA), Streptococcus Pneumoniae and Klebseilla Pneumoniae

May Florence Dela Cruz Bacayo

*, Nurul Shahira Sajali

, Wong Charng Choon

, Santosh Fattepur

Kiran Chanabasappa Nilugal

, Jiyauddin Khan

, Fadli Asmani

, Eddy Yusuf

School of Pharmacy, Management and Science University, 40100 Shah Alam Malaysia

Keywords: Antibacterial Activity, Garcinia Atroviridis Extracts, Zone of Inhibition.

Abstract: The curative ability of the endemic plants for different disorders has been described by traditional medicine

practitioners. Some herbs have been reported to have antibacterial activity such as Garcinia Atroviridis. The

antibacterial activity of Garcinia Atroviridis (Asam Gelugor) extracts was evaluated against Methicillin-resistant

Staphylococus aureus, Streptococcus pneumoniae and Klebseilla pneumoniae. Asam Gelugor fruits (600 grams)

were collected. The powder form were successively extracted with with 99.8 % of methanol. The extract was

filtered and dried using the rotatory evaporator at a temperature not exceeding 50ºC. The antibacterial activity

was determined by disc diffusion method for the zone of inhibition, minimum inhibitory concentration (MIC)

and minimum bactericidal concentration (MBC). The zone of inhibition were compared with that of ampicillin,

vancomycin, and gentamicin antibiotic disc. The test by using disc diffusion method shows highest inhibition

against the Klebsiella pneumoniae (15.33 ± 1.53) in 100% extract. Based on the result for MIC, inhibition is at

50mg/ml until 0.05 mg/ml against MRSA, while the MIC is positive in all concentration of extract against

Klebsiella pneumoniae but the MIC result is negative in all concentration of the extract against Streptococcus

pneumoniae. MBC result showed that there is bacterial growth in 500mg/ml of extract against MRSA while no

bacterial growth in extract against Klebsiella pneumonia, bacterial growth is positive in the extract against the

Streptococcus pneumoniae. The results shows that Asam Gelugor (Garcinia Atroviridis) may serve to the

development of a new antibacterial agent against these type of bacteria.

1 INTRODUCTION

Disease can be cure not only by a processed medicine

or medicine that can we can get at the market but

home remedies can also be used to treat the disease

furthermore, it is cheaper than the marketed drug.

Example of home remedies are ginger, turmeric,

garlic, black pepper, tamarind that can treat illness

such as infection or to reduce body temperature. Back

to ancient era, they use endemic plants to treat several

illness. Thus, plants play a major role in the treatment

of diseases. A very common example was morphine

from the opium poppy and most of the sources of the

drugs were from plant, animal and microorganism.

(Evans, W.C. Trease and Evans Pharmacognosy.,

2009)

Curative ability of many endemic plants has been

described by the practitioners of traditional medicine

for centuries. The increasing antibacterial activity of

various medicinal plants were reported from the

different parts of the world. The World Health

Organization estimates that plant extracts or their

active constituents are used as folk medicine in

traditional therapies of 80% of the world’s population.

(World Health Organization, WHO).

Herbs are widely exploited in the traditional

medicine and their curative potentials are well

documented. About 61 % of new drugs that were

developed between 1981 and 2002 were of natural

Bacayo, M., Sajali, N., Choon, W., Fattepur, S., Nilugal, K., Khan, J., Asmani, F. and Yusuf, E.

The Study of the Antibacterial Activity of Asam Gelugor (Garcinia atroviridis) Against Methecillin-resistant Staphylococcus Aureus (MRSA), Streptococcus Pneumoniae and Klebseilla

Pneumoniae.

DOI: 10.5220/0009843000002406

In Proceedings of BROMO Conference (BROMO 2018) - Symposium on Natural Product and Biodiversity, page 1

ISBN: 978-989-758-347-6

products focusing on infectious diseases and cancer

therapy. Unfortunately, the discovery rate of active

novel chemical entities is declining. Thus, natural

products from plants may give a new source of

antibacterial activity with possibly novel mechanism

of action. As the effects of plant extracts have been

studied by many researchers. (Nayan et al, 2011)

Antibacterial agents are the well-known weapons

to fight bacterial infections and helps to improve the

quality of human life since its introduction. But, due to

the emergence of drug-resistant bacteria, these

antibacterial agents have become less effective. It is

very essential to investigate newer drugs derived from

natural sources for the prevention and treatment of

bactrial infections as well as to combat antibiotic

resistance.

A large rainforest of Peninsular Malaysia is the

haven of Garcinia Artroviridis, known as Asam

Gelugur, Asam Gelugo or Asam Keping in Malay.

This species grows throughout in the rainforest and

widely cultivated especially in the northern states.

Asam Gelugor is commonly used for its weight

reduction properties (Ensiklopedia Pengubatan Herba,

2013). The tree grows for about 20 meters high and

has extended trunk, greyish smooth bark and hanging

branches. The floret are darkish red with a yellowish

to orange fruits borne singly on the ends of the twigs.

The plant contains fruit acids such as citric acid,

tartaric acid and ascorbic acid, that have antioxidant

properties. Phytochemical investigations have isolated

garcinia acid and its γ-lactone, atroviridin,

atrovirisidone, atrovirinone, and some organic acids.

(Amran et al, 2009). Hence, this study aims to

investigate the antibacterial activity of Garcinia

Atroviridis (Asam Gelugor) against MRSA,

Streptococcus pneumoniae and Klebsiella

pneumoniae.

2 METHODOLOGY

2.1 Extraction Process

Asam Gelugor fruits were collected and cut into small

pieces (600g). The oven dried fruits were grind until

its powder form which were extracted with 99.8%

methanol using a Soxhlet extractor. The solution was

filtered and dried using the rotatory evaporator at a

temperature not exceeding 50ºC.

2.2 Disc Diffusion Method for

Determination of Zone of Inhibition

The petriplates were inoculated with sterile swab

dipped into the inoculums. The excess inoculum from

the swab was removed by gently pressing and rotating

the swab firmly against the side of the tube in order to

remove excess fluid in the swab.

The sterile swab was streaked all over the surface

of the petriplate three times for a lawn of growth,

rotating the petriplate through an angle of 60° after

each application. The sterile swab was passed round

the edge of the agar surface. After the streaking, allow

the petriplates to dry in a room temperatures for a 5

minutes, with the petriplates lid closed. A narrow hole

was the bore in the petriplates and the extract were

added in the hole. The petriplates were placed in an

incubator at 37°C within 30 minutes

After 24 hours of incubation, use a metric ruler to

measure the diameter of the zone (including the

diameter disc) without opening the lid and record the

diameter in mm.

2.3 Minimum Inhibitory Concentration

(MIC)

Agar well diffusion method and the micro-broth

dilution technique were employed to determine the

minimum inhibitory concentration (MIC) for each

extract and test organism. A reconstituted extract of

500 mg/mL concentration was serially diluted in two-

fold up to 0.05 mg/mL. A 100 μL volume of each

dilution was introduced into duplicate wells in the

Mueller Hinton Agar plates that is pre‑inoculated with

test bacterial strain; and incubated at 37 °C for 24 h.

The minimum inhibitory concentration was taken and

recorded as the lowest concentration of the extract

showing measurable inhibition zone. For the

micro‑broth dilution technique, a 100 μL volume of

each dilution of the extract was introduced into

duplicate tubes of 2.0 mL Mueller Hinton broth

(MHB) seeded with 100 μL of the standardized

suspension of the test bacterial strain. Incubation was

at 37 °C for 24 hours and MIC was taken as the lowest

concentration of the extract that made the culture show

no visible growth.

2.4 Minimum Bactericidal

Concentration (MBC)

BROMO 2018 - Bromo Conference, Symposium on Natural Products and Biodiversity

A modified agar well diffusion technique was

employed in the determination of the minimum

bactericidal concentration (MBC). A 2 mm diameter

agar disc cut out from the inhibition zone of the last

three consecutive wells in each dilution showing

inhibition was inoculated into a fresh sterile nutrient

broth medium. The broth cultures were incubated at 37

°C for 24 hours after which 100 μL was spread over a

fresh sterile MHA. The MHA culture was in turn

incubated at 37 °C for 24 hours and the least

concentration of the extract showing no growth was

taken as the MBC. An MBC which coincided with or

was next to the MIC value was considered bactericidal

while those that differed markedly were considered

bacteriostatic.

3 RESULT AND DISCUSSIONS

The antibacterial activity of Garcinia Atroviridis has

been measured by the different test employed on it.

The disc diffusion method was used to measure the

zone of inhibition of the extract shows highest

inhibition against Klebsiella pneumoniae (15.33 ±

1.53) in 100% concentration of the extract followed by

inhibition against MRSA (10.76 ± 4.13) then

Streptococcus pneumoniae (4.88 ± 0.44). The negative

result is shown the inhibition of 25% concentration of

the extract against Streptococcus pneumoniae.

The statistical tool one-way ANOVA showed that

the antibacterial activity of the fruit extracts is

significant when compared with positive control with

the P value (p< 0.05) against MRSA, S. pneumoniae

and K. pneumoniae.

MIC results showed at 50mg/ml until 0.05 mg/ml

against MRSA, while the MIC is positive in all

concentration for inhibition against Klebsiella

pneumoniae but the MIC result is negative in all

concentration against Streptococcus pneumoniae.

The MBC result showed that there is bacterial

growth in 500mg/ml of extract against MRSA while

no bacterial growth in the lowest concentration of

extract against Klebsiella pneumoniae. The bacterial

growth is positive in the lowest concentration of

extract against the Streptococcus pneumoniae.

The antibacterial activity of Asam Gelugor fruit

extract which was shown in this study is attributed to

the presence of the different active constituents of the

plant. (Zakaria, 2011)

Table 1: Zone of inhibition vs bacterial species with different

concentration of G. Atroviridis plant extract

Figure 1: Comparison of zone of inhibition of plant extract

vs gentamicin in Klebsiella pneumoniae

Figure 2: Comparison of zone of inhibition of plant extract

vs ampicillin in Streptococcus pneumoniae

Figure 3: Comparison of zone of inhibition of plant extract

vs ampicillin in Streptococcus pneumoniae

The Study of the Antibacterial Activity of Asam Gelugor (Garcinia atroviridis) Against Methecillin-resistant Staphylococcus Aureus

(MRSA), Streptococcus Pneumoniae and Klebseilla Pneumoniae

Table 2: Summary of G. Atrovidis plant extract zone of

inhibition against the 3 strains of bacteria

BACTERIA

MRSA

K.PNEUMONIAE

S.PNEUMONIAE

SIG

CONCENTRATION

MEAN (mm) ± SD

100%

10.76

± 4.1

15.33 ± 1.53

4.8 ± 0.44

0.05

50%

5.66 ±

1.57

7.00 ± 1.00

2.43 ± 0.40

25%

4.53 ±

0.50

5.33 ± 0.58

4 CONCLUSION

The in-vitro antibacterial study of Garcinia Atrovidis

against the selected gram-negative and gram-positive

organisms conclude that Garcinia Atrovidis extract

strongly inhibit the growth of Klebsiella pneumoniae

as well as the standard antibiotic (Gentamicin). The

plant extract has the potential to treat infection caused

by Klebsiella pneumoniae.

ACKNOWLEDGMENT

The author is very grateful and thankful to the research

committee and lecturers of Management and Science

University, Malaysia for providing all the needed

equipment and materials as well as the continuous

guidance and support until the completion of this

research project.

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The Study of the Antibacterial Activity of Asam Gelugor (Garcinia atroviridis) Against Methecillin-resistant Staphylococcus Aureus

(MRSA), Streptococcus Pneumoniae and Klebseilla Pneumoniae