Study of anti-microbial activity of

Hibiscus rosacinensis leaf extract.

-Arijit Chakraborty

Project done under supervision of

Biswanath Ghosh

Theory:

Anti-microbrial:

An anti-microbial is a substance that kills

or inhibits the growth of microorganisms[1] such

as bacteria, fungi, or protozoan's. Antimicrobial drugs

either kill microbes (microbiocidal) or prevent the growth

of microbes (microbiostatic). Disinfectants are

antimicrobial substances used on non-living objects or

outside the body.

Main classes:

There are mainly two classes of antimicrobial

drugs: Those obtained from natural sources:

Beta-lactam antibiotic (such as penicillins, cephalosporins)

Protein synthesis inhibitors (such

as aminoglycosides, macrolides, tetracyclines, chloramph

enicol, polypeptides)

• Synthetic agents:

– Sulphonamides, cotrimoxazole, quinolones

– Anti-virals

– Anti-fungals

– Anti-cancer drugs

– Anti-malarials

– Anti-tuberculosis drugs

– Anti-leprotics

• Anti-protozoals

Apparatus required:

i) Round bottom flask (RBF)

ii) Socks let apparatus.

iii) Beaker (1000 ml, 500 ml, 250 ml)

iv) spatula

v) Glass rod

vi) Funnel

Chemicals required:

i) Ethanol

ii) Chloroform

iii) Pet ether

iv) Phenol

v) Peptone, Nacl, Beef extract

vi) Distilled water

Procedure of Solubility chake:

8 test tubes are taken. 4 of them cold

water and rest are hot water.

Procedure of Thin Layer Chromatography:

At first TLC plate are prepared by silica gel

and water and dry it. Then TLC chamber are

prepared by organic materials like acetic

acid, methanol and water (3:2:1).

Procedure:

I. Leaves are collected and dried in room temperature up to one week.

II. Then leaves are crushed in mixing grinder. (Weight of leaf is 50 gm.)

III. Then the leaves are extract by using socks let apparatus.

IV. Four types of extraction are performed using Organic material.

I) Ethanol Extraction.

II) Chloroform Extraction.

III) Pet. Ether Extraction.

IV) Water Extraction.

V. In ethanol extraction ethanol is used 250 ml.

VI. In chloroform extraction chloroform is used 250 ml.

VII. In pet ether extraction pet ether is used 250 ml.

VIII. Extraction sample are taken in to petridishs.

IX. Then dried the Petridis.

X. Then chake the solubility of the extract in hot and cold

water.

XI. Then chake the phenol coefficient by thin layer

chomatography (TLC) method.

XII. Than redal walkar coefficient test are performed to

chake the anti microbial activity of Salmonella typhi in

nutrient broth.

XIII. Then chake the result.

Result

Observation table of solubility:

Sample Cold water Hot water

Ethanol extract Soluble Soluble

Chloroform extract Insoluble Insoluble

PET-Ether extract Insoluble Insoluble

Water extract Soluble Soluble

Observation table of TLC:

Sample Response

Ethanol extract + ve

Chloroform extract -Ve

PET-Ether extract -Ve

Water extract + ve

Observation table of R.W.C:

Concentration Time Of contact (gm)

2(1/2) 5 7(1/2) 10

1/100

(phenol)

+ + - -

1/100 (ethanol) + + - -

1/200

(ethanol)

+ - - -

1/400 (ethanol) + - - -

1/500 (ethanol) + - - -

Concentration Time Of Contact (gm)

2(1/2) 5 7(1/2) 10

1/100 (phenol) + + - -

1/100 (water) + + + -

1/200 (water) + + - -

1/400 (water) + - - -

1/500 (water) + - - -

Calculation:

Formula:

Rideal walker coefficient= (MIC of phenol/ MIC of

sample)

Ethanol extraction:

R.W.C = (MIC of phenol/MIC of ethanol)

= {(1/100)/(1/100)}

= 1

Water extraction:

R.W.C = (MIC of phenol/ MIC of water)

= {(1/100)/(1/200)}

= 2

Conclusion:

After performing the experiment, it

was found that Rideal walker coefficient of

ethanol extraction is 1 and water extraction is

2. So, 1/100 dilution of ethanol extraction is

equal to 1/100 dilution of phenol and 1/200

dilution of water extraction is effective as the

1/100 dilution of phenol.