Isolation and screening technique to

produce secondary metabollite from

microorganism
Nur Ida Panca N
2017
 Secondary metabolites do not play a role
in growth, development, and
reproduction, and are formed during the
end or near the stationary phase of
growth
 Many of the identified secondary metabolites
have a role in ecological function, including
defense mechanism(s), by serving as
antibiotics and by producing pigments.
 Examples of secondary metabolites include :

Erythromicyn

Bacitracin

Pigments
 Antibiotics
Most of the antibiotics in current use for the
treatment of various infectious diseases are
microbial products.
Studies on soil bacteria and fungi have shown
that these microorganisms are potentially rich
source of unique bioactive substances.
 Antibiotics
1. Isolation And Screening Of Soil Bacteria For
Potential Antimicrobial Activity
Antimicrobial agents are the most promising field
worldwide with the need of continuous search of
new ones.
 Materials & Methods
Source of soil sample :

Farm Soil Oil Spilled Soil

 Materials & Methods

Below 5-6 cm depth COLLECTED

stored in sterile plastic bags at 4C

 Materials & Methods
Isolation of antibiotic producing bacteria :

mixed thoroughly

0.5 g of soil Serial dilution

4.5ml sterile
sample saline tubes

0.1 ml of each aliquot was spread on NUTRIENT

AGAR (1%) (37C for 24 hr)
 Materials & Methods
Culture Identification and Preservation:

Preserved on agar slant

Cultures were purified

by streak plate method Preserved in 40%
glycerol at -70C
 Antibiotics
Screening of cultures for antimicrobial activity :
Standard clinical isolates; Escherichia coli,
Staphylococcus aureus, Pseudomonas vulgaris,
Cornybacterium pseudodiphtherieae, Proteus sp.
The screening of potential strains was done by
cross streak method
 Antibiotics
2. Isolation And Screening Of Antimicrobial
Producing Bacteria from Aquatic
Environment
Aquatic environment is souce of enormous
biological diversity which is scarcelly studied.
 Materials & Methods
Source of water sample :
 Materials & Methods
Contaminated water
 Materials & Methods

Isolation of bacteria :
Serial dilution

1 ml of water
sample

0.1 ml of each aliquot was

spread on NUTRIENT AGAR
 Materials & Methods
Growth and production of antimicrobial
The colony Inoculated in BHI broth (48 h at 31 1C
125 cycles/min)

centrifuge at
Supernatant 10,000 rpm for 15 minutes

Evaluate antimicrobial Filtrat

activity
 Antibiotics
Detection of antimicrobial activity
20 l Applied on cellulose disk on BHI agar plate
supernatant (previously inoculated with each indicator
strain suspension)

Clear zone
 Antibiotics
Treatment factors that can be used :

Effect of heat on antimicrobial

activity

Effect of enzyme on antimicrobial

activity
 Sources :
Source: Boundless. Primary and Secondary
Metabolites. Boundless Microbiology
Boundless, 26 May. 2016. Retrieved 22 Feb.
2017 from
https://www.boundless.com/microbiology/tex
tbooks/boundless-microbiology-
textbook/industrial-microbiology-
17/industrial-microbiology-198/primary-and-
secondary-metabolites-999-5345/
 Sources :
Hassan , et al. 2014. Isolation And Screening
Of Soil Bacteria For Potential Antimicrobial
Activity. FUUAST J. BIOL., 4(2): 217-219