inoculum development for fermentation.ppt

1. The Development of Inocula for
Industrial Fermentations

2. What criteria should fermentation
inoculum satisfy?
•It must be in a healthy and active state
•Available in sufficiently large volumes to provide an
inoculum of optimum size
•Suitable morphological form
•Free of contamination
•Retain its product-forming capabilities

3. What is difference between Inoculum
and Production medium?
•Inoculum grows in culture medium
•Design of production medium is determined
not only by nutritional requirements of
organism but also by requirements for
maximum product formation
•Both media can be of different composition

4. Examples of inoculum and production
media

5. How much quantity of inoculum is
used?
•Volume : Normally 3 – 10% is used
•Stock culture – Inoculum built up in number
of stages
•Risk of contamination and strain
degeneration

6. Criteria for transfer of inoculum
•Physiological condition of inoculum – major
effect on performance of fermentation
•Indicators of inoculum quality include
dissolved oxygen, pH and Co2
•Yeast, Bacteria and Fungi – common inocula

7. Preservation of Industrial
Microorganisms
•Stored – eliminate genetic change, protect
against contamination and retain viability
•Storage at reduced temperature – agar slopes
(5°) or freezer (-20°) – SC – 6 months
- Storage under liquid nitrogen – resuspension
in 10% glycerol , -150° to -196°C

8. Preservation of Industrial
Microorganisms
•Storage in a dehydrated form
-Dried soil cultures – fungi and actinomycetes
-Moist sterile soil – inoculate for several days
for some growth – dry at room temperatures
for 2 weeks – refrigerate samples
-Lyophilization or freeze drying – Bacteria
-Freezing culture followed by drying under
vacuum – resulting in sublimation of water.

9. How do Yeast multiply?
•Asexual – Fission or
Budding
-Takes place within a
single parent (haploid
or diploid)
-Splits nucleus – forms
daughter cells

10. How do Yeast multiply?
-Sexual – haploid cells-
Shmooing form –
Diploid cells
-Unfavorable conditions
– haploid cells die
-Diploid cells undergo
meiosis - into 4
daughter haploid
spores

11. Development of yeast inocula
•Wort – Batch of fermentation
•Crop – Harvested yeast from previous
fermentation
•Pitch – Inoculate

12. Development of yeast inocula
•Middle skimmings required
– Yeast cells float to the surface – most
flocculate
-Bottom cells – least flocculate
-Skimmings treated – reducing pH of slurry to
2.5 to 3, washing with water or ammonium
persulphate and treatment with antibiotics
like Polymixin, Penicillin and Neomycin

13. How do Bacteria multiply?
•Asexual reproduction –
Bacteria splits into two
cells – Binary Fission
•Sexual reproduction –
Joining of two parent
cells and exchange of
genetic material
- Conjugation,
Transformation and
Transduction

14. Development of bacterial inocula
•Produce active inoculum – gives short lag
phase
•Should be transferred in logarithmic phase of
growth – metabolically active
•Small amount of inocula – prevent
contamination and avoid an abnormal long
lag phase

15. How do Fungi multiply?
•Asexual reproduction – forms spores
•Sexual reproduction – Two parent cells unite
to form zygote

16. Development of mycelial inocula
•More difficult than yeast and bacteria
•Industrially important fungi and
streptomycetes – capable of asexual
sporulation – use spore as inocula

17. Development of mycelial inocula
•Three basic techniques – produce high
concentration of spores for use as inoculum
-Sporulation on solidified media
-Sporulation on solid media
-Sporulation in submerged culture

18. Development of mycelial inocula
•Sporulating on solidified media:

19. Development of mycelial inocula
•Sporulation on solid media
-Many filamentous organisms will sporulate on
surface of cereal grains
-Substrates – barley, hard wheat bran, ground
maize and rice
-Affected by amount of water added to cereal
before sterilization and relative humidity

20. Development of mycelial inocula
•Sporulation in submerged culture:
•More convenient than others – easy to
operate aseptically and done on large scale
•Suitable medium