This document discusses amylase production through submerged fermentation using Bacillus spp. It defines amylases as enzymes that break down starch and describes their classification. It explores the advantages of using microbes like Bacillus licheniformis and Bacillus amyloliquefaciens for producing amylases economically and to specification. The document outlines the materials and methods used, including culturing Bacillus spp., varying the fermentation temperature and pH, extracting and assaying the amylase. It concludes by reviewing industrial applications of alpha, beta, and gamma amylases.

1. Amylase Production on
Submerged Fermentation
by Bacillus spp.
Made by-
Miss Karishma Gangwani
MSc. Biotechnology sem II
Hislop School Of
Biotechnology

2. What Are Amylases ?
 enzymes that break down starch or glycogen.
 present in the saliva of humans and some other mammals, where it begins
the chemical process of digestion.
 All amylases are glycoside hydrolases and act on α-1,4-glycosidic bonds.
 produced by a variety of living organisms, ranging from bacteria to plants
and humans.

3. Classification Of Amylases
 based on how they break down starch molecules :
1. α-Amylase
2. β-Amylase
3. γ-Amylase

4. Advantages And Uses Of Microbial Amylase
 production of amylases is in economical bulk production capacity.
 microbes are also easy to manipulate to obtain enzymes of desired
characteristics.
 The microbial amylases meet industrial demands a large number of
them are available commercially.
 “BACILLUS LECHENIFORMIS”
“BACILLUS AMYLOLIQUIFACIENS”
“ASPERGILLUS NIGER”
 useful applications in food, brewing, textile, detergents and
pharmaceutical industries.
 In detergents production, they are applied to improve cleaning effect
and are also used for starch de-sizing in textile industry.
 mainly employed for starch liquefaction (a process of dispersion of
insoluble starch in aqueous solution) to reduce their viscosity,
production of maltose, oligosaccharide mixtures.

5. Materials And Methods For Production
 MICROORGANISM: Bacillus spp. was isolated from soil
and maintained on nutrient agar slants and subcultured for
every 10 days.
 INOCULUM AND FERMENTATION MEDIUM: The
inoculum was prepared by addition of sterile distilled water
into the freshly grown nutrient agar slants, from this 0.5ml of
cell suspension was inoculated into 100ml of sterilized
fermentation medium and incubated at 35°C for 10 hrs.
 The composition of fermentation medium are:
Bacteriological peptone –6gm
Magnesium sulfate –0.5gm
Potassium chlorate –0.5gm
Starch –1gm (pH -7)

6. Extraction Of Amylase From Fermentation
Medium:
 After incubation the fermentation medium was harvested by
centrifugation at 5000 rpm for 20 min at 4°C.The supernatant was
collected and subjected to estimate the amylase activity.
 EFFECT OF PH:
The fermentation medium was prepared by varying pH values(5.0, 6.0,
7.0, 8.0) for the production of amylase.
 EFFECT OF TEMPERATURE:
To study the effect of temperature on amylase production, the
submerged fermentation was carried out at different temperatures
(25°C, 30°C, 35°C and 40°C).

7. Assay of Amylase:
 The amylase activity is determined following the method of
Bernfeld. An assay mixture containing, enzyme extract, starch as a
substrate and DNSA as coupling reagent was used.
 One unit of amylase activity was defined as the number of micro
moles of maltose liberated by 1ml of enzyme solution per minute.

8. Applications Of Industrial Amylases:
ENZYMES SOURCE APPLICATIONS
Alpha- amylase Bacterial α amylase
(e.g., Bacillus subtilis
), Fungal α amylase
(e.g., Aspergillus niger
)
Textiles, starch syrups,
laundry and
dishwashing
detergents, paper
desizing , fermentation
ß- amylase From a strain of
Bacillus
Brewing, maltose
syrup
γ-Amylase Aspergillus niger Manufacture of
dextrose syrup and
high fructose syrup
production.

9. THANK YOU!!