Fermentation Industry

Question 1

What is biofuel

Answer 1

It is a carbon neutral and a green energy source in 22nd century.
It is a liquid or gaseous fuel for internal combustion engines that are predominantly produced from biomass
Contribute to reduction of greenhouse gas emissions

Question 2

Examples of Biofuels

Answer 2

-Solid biofuels covers solid organic, non-fossil material of biological origin (also known as biomass) which may be used as fuel for heat production or electricity generation.

• Biogas is a gas composed principally of methane and carbon dioxide produced by anaerobic digestion of biomass or by thermal processes from biomass, including biomass in waste.

Question 3

What is synthetic biology and provide an example

Answer 3

Synthetic biology is to use organisms to produce recombinant proteins. It runs with system biology.

Bioremediation is highly involved in degradation, eradication, immobilization, or detoxification diverse chemical wastes and physical hazardous materials from the surrounding through the action of microorganisms. The main principle is degrading and converting pollutants to less toxic forms.
Slurry-phase bioremediation is a rapid process treatment processes. Contaminated soil is combined with water, nutrient and oxygen in the bioreactor to create the optimum environment for the microorganisms to degrade the contaminants which are present in soil. This processing involves the separation of stones and rubbles from the contaminated soil. After completion of this process the soil is removed and dried up by using vacuum filters, pressure filters and centrifuges. Then soil disposition occurs and resultant fluids are treated.

Question 4

Difference between recombination technology vs synthetic biology

Answer 4

The difference between recombinant technology and synthetic biology is that recombinant technology is the process of combining 2 or more DNA strands from different sources into 1 molecule to create a new set of genes, while synthetic biology is the design and construction of new biological parts, devices, and systems, and the re design of existing, natural biological systems from organisms for useful purposes.

Question 5

Define systems biology

Answer 5

Mathematical modelling combined with computational modelling of functional organism.
To design biological model of specific organism.
Important in bioengineering
Application to produce heterologous proteins.

Question 6

What is a pilot plant

Answer 6

A pilot plant is established before large scale plant with less quantities added compared to large scale. It is used for testing various parameters and for changing processes. It has enabled testing of new techniques on semi scale production scale and aid for process development.

Question 7

Define fed batch culture

Answer 7

It is the addition of specific carbon source or feed supplement to the culture at a specific interval to control growth of organism and suppressing limitations.

Question 8

What is batch culture

Answer 8

It is a closed system with a limited amount of nutrients added. Stopped at idiophase or trophophase to extract product. Can cause the accumulation of toxic wastes.

Question 9

What is continuous culture

Answer 9

Continuous culture is a process where nutrients are continually added to the bioreactor and the culture broth is removed at the same time.
The volume of the culture beoth is constant due to a constant feed in and feed out rate.
It is possible to set up the optimum conditions for maximum and long term product synthesis.

Question 10

Describe Penicillin production

Answer 10

1. The Penicillium chrysogenum contain its carbon source found in corn steep liquor and glucose.
2. The corn steep liquor and glucose are added to fermenter. Medium consists of essential ions required for fungus metabolic activity.
3. Medium is sterilized at high heat and high pressure with the fermentor.
4. Fermentation carried out fed batch mode (excessive glucose inhibit penicillin production)
5. Fermentation require 20 to 24 C temp, pH: 6.5
6. Culture medium mixed throughly around 200 rpm.
7. The seed culture developed in lab by addition of penicillium chrysogenum spores into liquid medium, after growth if is inoculated into fermenter.
8. Medium constantly aerated and agitated. Fed batch style, carbon and nitrogen added sparingly.
9. After 40 hrs, penicillin get secreted by fungus.
10. After 7 days, growth is completed, ph8 rise to cease penicillin production
11. Filtration by rotary vacuum filter to remove biomass (fungus and impurites)
12. To decrease pH phosphoricacid is introduced.
    13.organic solvent to dissolve penicillin present in filtrate.
13. Centrifugation carried out separating liquid (containing penicillin) amd solid waste.
14. Supernatant transferred downstream to continue extraction.

Question 11

Describe the different assays after colle tion from fermentor

Question 12

The advantages amd disadvantages of manufacturing of ethanol from sugarcane/ maize

Answer 12

Advantages:
Any plant can be used for production of bioethanol provided it contains sugar and starch.
Overall decrease in depletion of ozone layer, emissions produced are less reactive.
Reduces greenhouse gases through bioethanol blended fuel
Exhaust gases pf ethanol are cleaner
Bioethanol is biodegradable and less toxic than fossil fuels.

DISADVANTAGES:
Large amount of land is required to grow these crops
Sacrifice food crops for biofuel production
Expensive to set up biofuel laboratory
Demands strong technical knowledge for effective production amcavoid excess emissions.

Question 13

Why large scale fermentation is better than small scale fermentatiom

Answer 13

-Large scale with continuous production is more economic because of mass production.
-The initial implementation to produce 1000L and 10L are the same
- Harvest profit is larger in large scale

Question 14

What is most importance in large scale production in fermentation process

Answer 14

It is important to examine of recombinant cell products e protein by checking gene expression of DNA, mRNA ams protein

Question 15

Describe the acetone butanol fermentation process

Answer 15

Production of Inoculum:
Two species of Clostridium are employed and its spores are inoculated in medium in deep tubes. The tubes are heat shocked amd rapidly cooled to select the heat resistant spores. The tubes are incubated for growth and are successively transfered to larger media for increased growth.
At the stages of inoculum transfer, anaerobic conditions are produced by reducing condition of medium, and I’m,edited use of fresh and cooled medium before air becomes incorporated.
Medium for fermentation process is made.
Fermentation carried out anaerobically.
Phase2: rapid growth of bacterium and formation of large amounts of acetic acid, butyruc acid, CO2 and H2 gases. pH decrease amd remains constant.
Acidity increases ams adaptive enzymes produced to convert acids into neutral solvents.
2 phase: acid break km which more acids gets converted to acetone and butanol.
3 phase: rate or gas and solvent production decrease. Cells undergo autolysis and release riboflavin.
The fermentation yields several products which are industrially important.
Recovery process begins:
Acetone and butanol vaporises due to effect of steam.
Stea, and solvents are collected and condensed by cooling to get a solution.
The individual solvents in the solution are separated by fractional distillatikn

Question 16

Briefly describe the stage 1 of development in fermentation industry

Answer 16

Stage 1 (pre 1990):
Vinegar
Alcohol
Pasteur convinced the scientific world on the obligatory role of these microorganisms in fermentation.
Carlsberg brewery developed methods for isolating & propagating single yeast cells to produce pure cultures, established techniques to produce starter cultures.
The first aerobic fermenter developed for Vinegar generation.
Both were batch culture method
There was no proper quality control and no pilot plant facilities.

Question 17

Briefly describe the stage 2 of development in fermentation industry

Answer 17

1990 - 1940 Bakers yeast, glycerol, citric axis, lactic acid and acetone butanol
Glycerol biosynthesized by fermenting yeast Saccharomyces cerevisiae
Bakers yeast aerobic process, rapid growth resulted in ethanol production at the expense of biomass formation due to the oxygen depletion.
This was solved by fed batch system in which initial wort concentration was descresed so growth of cells are limited by carbon source rather than oxygen. Subsequent growth controlled by adding further wort on small increments.
Steam sterilisation under pressure was initiated to avoid contamination by aerobic bacteria.

Question 18

What are the 6 basic components of a fermentation process

Answer 18

1. Formulation of media to be used in culturing the process organism during the development of inoculum and in production fermenter.
2. Sterilization of medium, fermenters, and ancillary equipment.
   3.

Production of an active, pure culture in sufficient quantity to inoculate the production vessel.

4. Growth of organism in production fermenter under optimum conditions for product formation
5. Extraction of product and its purification
   6.
   Disposal of effluents produced by the process

Question 19

Briefly describe the stage 3 of development in fermentation industry

Answer 19

1940s
A major advancement was the development of large scale Extraction process for recovery of Penicillin
Penicillin production required establishment of strain improvement programs which became a dominant feature.
The introduction of pilot plant facilities aided process development which enabled testing pf new techniques on semi production scale.
Penicillin fermentation technology provided basis for new process development such as antibiotics and vitamins.
Microbial products were screened for different activities which lead to miniaturised culture systems, robotic automation and assays.
Initiated programs for manufacturing ethanol for motor fuel

Question 20

Briefly describe the stage 4 of development in fermentation industry

Answer 20

Use of hydrocarbons which are potential carbon sources result in increased oxygen demands and high heat outputs by these fermentations which led to develop pressure jet and pressure cycle fermenters that eliminated mechanical stirring requirement.
Batch and fed-batch fermentations were common in the industry but the technique of growing an organism continuously by adding fresh medium to the vessel and removing culture fluid had been applied only to a very limited extent on a large scale.
Vessel size continued getting larger but extremely large continuous fermenter for 100 day operation caused large aseptic problem.
Hence, high standards of fermenter construction, aseptic operation and continuous sterilization of feed streams were established.
To minimise human error, computer systems to control sterilization am operation cycles was established.

Question 21

Briefly describe the stage 5 of development in fermentation industry

Answer 21

Low value high volume product
“New biotechnology”
In vitro recombinant DNA technology enabled the expression of human and mammalian genes in cultured animal cells and microorganisms.
This lead to the development of large-scale fermentation processes for the production of therapeutic human.
These products have been classified as biologicals and are commonly referred as biopharmaceuticals.

Exploitation of genetic engineering - production of monoclonal antibodies (mabs), major development in biotechnology, influenced fermentation industrial progress

Availability of monoclonals – improved analytical techniques, increased expectations as therapeutic agents.

Establishment of recombinant proteins

Question 22

During 2005-12 , approvals were dominated by antibody based products and engineered proteins which have been modified post synthesis. What are these modifications?

Answer 22

• Antibody drug conjugates which bind to the tumor directing the drug to its target.
• Modifications to extend the half life of the therapeutic protein in the body.
• Modifications to alter pharmacokinetic properties of therapeutic protein.

Question 23

Briefly describe the stage 6 of development in fermentation industry

Answer 23

Commercial use of recombinant proteins triggered the construction of production facilities for engineered producing organisms.

Recombinant proteins classified as biologicals
To produce biological must be precisely specified and carried out in a facility that has been inspected and licensed by the regulatory authority.
Any changes must receive regulatory approval.
For drugs, only major changes require prior approval.
The cost of developing recombinant protein process is extremely high.
Complete genome sequence enabled Comparison of genomes and visualisation of gene expression.
Adoption of systems biology and synthetic biology

Question 24

Components of a fermentation process

Answer 24

1. Formulation of media to be used in culturing the process organism during the development of inoculum and in production fermenter.
   2.

Sterilization of medium, fermenters, and ancillary equipment.

3. Production of an active, pure culture in sufficient quantity to inoculate the production vessel.
4. Growth of organism in production fermenter under optimum conditions for product formation
5. Extraction of product and its purification
6. Disposal of effluents produced by the process

Question 25

Before establishment of fermentation process begins, what must be carried out

Answer 25

1. a producer organism must be isolated
2. modified such that it produces the desired product in commercial quantities
3. its cultural requirements determined
4. design of the plant is optimized
5. extraction process must be established

Question 26

What is metabolic flux analysis

Answer 26

Examines flux of intermediates through pathways to construct, athematical mode, micking the metabolic networks of the cell