Antimicrobial activity of Annona Muricata L. aqueous extract

Supervisor : Sir Muhammad Norzein bin Abd Rahim Name: Muhammad Fattah bin FazelI.D Number: 12171211070Course: Diploma of Biomedical Laboratory Technology

Introduction

Annona Muricata L.

• AnnonaceaeFamily

• 5-10 m in height and 15 cm in diameter

• Trunk straight; bark smooth, dull grey or grey-brown, rough and fissured with age

Characteristics

• Leaves: Suppurative, febrifuge • Bark: Tonic• Roots: Antispasmodic,

parasiticidal

Traditional usage

(Orwa et al.2009)

(B. Neela et al., 2010)

Antimicrobial resistance

Resistance of a microorganism to an antimicrobial medicine to which it was originally sensitive.

(WHO, 2013)

- Infections caused by resistant microorganisms often fail to respond to the standard treatment- Resulting in prolonged illness and greater risk of death

(WHO, 2013)

- Become a serious problem for treatment of gonorrhea- Involving even "last-line" oral cephalosporins, and is increasing in prevalence worldwide

(WHO, 2013)

- Enzymes produced by the bacteria destroy last generation antibiotics.- Render powerful antibiotics to be ineffective.- Often the last defense against multi-resistant strains of bacteria

Antimicrobial

Resistance

Antimicrobial activity of Annona Muricata L.

• Seen against to the several organisms:PseudomonasStaphylococcusKlebsiellaBacillusE. coli.

(C. Vijayameena et al, 2010)

Antimicrobial activity of Annona Muricata L.

(C. Vijayameena et al, 2010)

Antimicrobial activity of Annona Muricata L.

• The antibacterial effect of the leaves of Annona muricata was evaluated on: Staphylococcus aureus

• Escherichia coli • Proteus vulgaris • Streptococcus pyogenes• Bacillus subtilis, Salmonella typhimurium,• Klebsiella pneumonia • Enterobacter aerogenes (M. Shaily et al., 2013)

Antimicrobial activity of Annona Muricata L.

• The most susceptible Gram-positive bacteria was B. subtilis and S.aureus while the most susceptible Gram-negative bacteria was K. pneumoniae and P. vulgaris.• The significant antibacterial activity of active extracts was

compared with the standard antibiotic, streptomycin.

(M. Shaily et al., 2013)

Objectives• General objective: • To study the antimicrobial property of Annona Muricata L. aqueous

extract. • Specific objectives• 1. To determine the inhibition zone of bacteria for each

concentration of Annona Muricata L. aqueous extract. • 2. To compare the inhibition zone of bacteria between Disc

Diffusion method and Well Diffusion method using Annona Muricata L. aqueous extract.

Hypothesis

• Null hypothesis: • Annona Muricata L. aqueous extract will not show any antimicrobial

effect towards Bacteroides fragilis, Clostridium perfringens, MRSA and Streptococcus pyogenes.

• Alternate hypothesis• Annona Muricata L. aqueous extract will show antimicrobial effect

towards Bacteroides fragilis, Clostridium perfringens, MRSA and Streptococcus pyogenes by inhibition of colony on the agar.

Problem statement

• Lots of antimicrobial medicine has been developed to fight pathogens. • Have evolve and become resistant to many antibiotics that

previously made.• Researchers started to study the traditional plants.• This study is conducted to investigate the antimicrobial activity

of Annona Muricata L. aqueous extract towards certain bacteria

Methodology

Methodology• Study design• Annona Muricata L. has been extracted into aqueous solution

to detect the antimicrobial activity by well diffusion and disc diffusion• The size of zone inhibition is measured at the end of the

experiment. • The main purpose of this experiment is to investigate whether

the Annona Muricata L. aqueous extract has antimicrobial activity against Bacteroides fragilis, Clostridium perfringens, MRSA and Streptococcus pyogenes or it does not

Project flowchart

Preparation of Annona Muricata L. extract

Preparation of Mueller-Hinton agar Make bacteria inoculum Culture the bacteria on

the Mueller-Hinton agar

Start the antibacterial activity of Annona

Muricata L. experiment

Observe and measure the inhibition zone Analysis of data

Extraction method

Well diffusion method

Well diffusion method

Well on MHA agar

Making well using Durham

tube

Disc diffusion method

Results

Results

• No inhibition zone appeared on Mueller-Hinton agar• For both well diffusion method and disc diffusion method

Results

Well diffusion method, MRSA no inhibition zone Well diffusion method, Streptococcus pyogenes no inhibition zone

Results

Disc diffusion method, Clostridium perfringens no inhibition zone

Disc diffusion method, Bacteroides fragilis no inhibition zone

Results  Diameter of Inhibition (mm)

Bacteria Gentamicin Vancomycin Amoxicillin/Clav Erythromycin

Streptococcuspyogenes 23 - - 12

        

MRSA 22 16 13 14

Bacteroides fragilis 21 - - -

Clostridium perfringens 22 - - -

Discussions

Discussions

• In this study, Annona muricata L. aqueous extract is used on MRSA, Streptococcus pyogenes, Bacteroides fragilis and Clostridium perfringens to test its antimicrobial activity. • However, no zone of inhibition showed on Mueller-Hinton agar

Discussions

Disc diffusion- retains the active component -does not allow it to diffuse in agar

-Dilution with normal saline -Extract compound not properly diffuse into the agar

Compound exist in small quantities- destroyed during dry under sun

Concentration of the stock

solution- not enough

to kill or inhibit the bacteria

Conclusions

Conclusion

• The antimicrobial activity of Annona muricata L. was not shown in this study.• Due to several factors during carrying out the experiment,

there was no inhibition zone appear on Mueller-Hinton agar for Bacteroides fragilis, Clostridium perfringens, Streptococcus pyogenes and MRSA.

• This study is important in finding new antimicrobial drugs because nowadays, there are too many antibiotic-resistance bacteria emerge around the world• Annona muricata L. extract might be the next successful

antibiotic and can help in curing disease cause by pathogenic microorganism. • Therefore, null hypothesis is accepted.

Recommendation

• Further studies can be done by increasing the concentration of the stock solution. • Also using different part of Annona muricata such as bark or

fruit.

Reference1.C. Vijayameena, G. Subhashini, M. Loganayagi and B. Ramesh,

2013. Phytochemical screening and assessment of antibacterial activity for the bioactive compounds in Annona muricata. International Journal of Current microbiology and Applied Sciences, 2(1): 1-8

2.Shaily Mishra, Shamim Ahmad, Nilesh Kumar, Brijesh Kumar Sharma, 2013. Annona muricata (The cancer killer): A review. The Global Journal of Pharmaceutical Research, 2(1): 1613-1618

3.Bipul Biswas, Kimberly Rogers, Fredrick McLaughlin, Dwayne Daniels, Anand Yadav, 2013. Antimicrobial Activities of Leaf Extracts of Guava (Psidium guajava L.) on Two Gram-Negative and Gram- Positive Bacteria, Hindawi Publishing Corporation

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