chapter 1 oct2012
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UEMK3233 Topic 1 8
1) Production of microbial cell (biomass)
Two main applications:-
a) as a source of protein Example: SCP
b) as a commercial inoculum (starter culture)
a) SCP production refer to microbial biomass used as food and feed
additives
either the isolated cell protein or the total cell materialmay be called SCP
great nutritional value high protein, vitamin, and lipid
content and the almost complete range of all essential
amino acids
Single Cell Protein
Microbial biomass or protein extracted there from
processes in which bacteria, yeast, fungi or algae are
cultivated in large quantitiesAs human or animal protein supplement in animal feed
or in human nutrition
As protein supplement (for infant as nutritional food
protein)
UEMK3233 Topic 1 10
Sample of the single cell protein
biosolids after the drum dryerUEMK3233 Topic 1 11
Component Alkane
yeast
Methanol
bacterium
Alga Soya
meal
Milk
powde
r
Raw protein 60 80 72.6 42 34
Fat 9 9.5 7.3 4 1
Mineral salts 6 9.5 4.7 6.5 8
moisture 4.5 2.8 3.6 10 5
Table 1: Composition (%) of SCP compared with soya meal
and milk powder
Protein content in bacteria 60 to 65%
Selected fungi and yeast 33 to 45%
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UEMK3233 Topic 1 12
Some psychological barriers:
microorganism involved must be safe & acceptable;
non-pathogenic; non-toxic forming (for example,
aflatoxins are produced by some fungi); geneticallystable
Toxic or carcinogenic substances used during the
production of microbial e.g., hydrocarbon, polycyclic
aromatic compounds .
may cause indigestion and allergic reactions
UEMK3233 Topic 1 13
b) as a commercial inoculants (or known as
starter cultures)
Major applications (as food starter cultures),
such as, in baking and dairy industries
Functions of food starter cultures:-
Texture modifications
Preservation
Flavor development
Nutritional improvement
UEMK3233 Topic 1 14
Examples:-
i. Bakers yeast bread making
ii. Cheese-starter cultures Streptococcus
cremoris, Streptococcus lactis
iii. Yoghurt manufacturing Stretococcusthermophilus, Lactobacillus bulgaricus, L.
acidophilus
Topic 1Topic 1 --ProkaryotesProkaryotes 1515
1 m 2 m 5 m
(a) Spherical (cocci) (b) Rod-shaped (bacilli) (c) Spiral
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UEMK3233 Topic 1 20
Examples of commercial applications of microbial enzymes
UEMK3233 Topic 1 21
c) Production of microbial metabolites
Growth of microbial divided into few stages:-
i. Lag phase
ii. Log orexponential
phase
iii. Stationary phase
iv. Death phase
Growth curve of a bacterial culture in
batch conditions
UEMK3233 Topic 1 22
Phases Processes
Lag Growth does not occur;
adaptation period
Log/exponential
Cells grow gradually at constantrate until reach maximum
growth rate
stationary Growth ceases and growth rate
reduced
death Viable cell number declinesUEMK3233 Topic 1 23
Product formation during microbial growth:-
a) Primary metabolites
referred asprimary products of metabolism
are essential for life and reproduction of cells
produced during log phase (trophophase)
e.g., amino acids, proteins, carbohydrates,lipids.
examples (refer Table 3 and 4)
also known as ????
produced by wild type OR genetically modifiedmicroorganism ???
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UEMK3233 Topic 1 24
some may be produced in concentrations far
higher than those required by the producing
strains
Table 3: Excessive production of certain primary metabolites
UEMK3233 Topic 1 25
Primary metabolite Commercial applications
Ethanol Active ingredient in alcoholic
beverages;
Citric acid Food industry
Acetone & butanol Solvents
Glutamic acid Flavour enhancer
Vitamins Feed supplements
Polysaccharides Food industry; oil industry
Table 4: Examples of primary product from microbial
metabolism and their commercial application
UEMK3233 Topic 1 26
UEMK3233 Topic 1 27
b) Secondary metabolites secondary products of metabolism produced during
stationary phase (idiophase) (Figure A)
are seemingly not essential for growth and reproduction
produced from the intermediates and products of primarymetabolism (refer Figure B) and their formation is extremely
dependent on environmental conditions every secondary metabolite is formed by only a few
organisms
Importance of secondary metabolites in fermentation
industries
a) Antimicrobial activity
b) Growth promoters
c) Pharmacological properties
Wild-type microbial ??? Genetically modified microbial??
also known as ?????
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UEMK3233 Topic 1 28
Figure A: Production of antibiotics
(e.g.Penicillium spp. & Streptomycetes)
UEMK3233 Topic 1 29
d) Recombinant products
advent of recombinant DNA technology extend the range
of potential fermentation products
genes from higher organisms may be introduce into
microbial cells recipients are capable of synthesizingforeign proteins
hosts E. coli, S. cerevisiae and filamentous fungi
products from GMO interferon, insulin, epidermal growth
factor, chymosin etc
important factors to consider
i. secretion of the product
ii. minimization of the degradation of the product
iii. maximizing the expression of the foreign gene
UEMK3233 Topic 1 30
e) Microbial transformation / Bioconversions
to chemically modify a wide variety of organic
compound to convert a compound into a
structurally related, financially more valuable
compound
examples:
1. production of vinegar conversion of ethanol to acetic
acid
2. production of steroids, antibiotics and prostaglandins
reaction involved:- oxidation, dehydrogenation,
dehydration, condensation, amination,
isomerization,
UEMK3233 Topic 1 31
advantageous over the use of chemical
reagents:-
operate at low temperature
without requirement of any potential heavy metalpollution
disadvantages:-
require large amount of biomass
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UEMK3233 Topic 1 32
Oxidation of ethanol to acetic acid
UEMK3233 Topic 1 34
UEMK3233 Topic 1 35
Many words have been used to describe
engineering working with biotechnology:-
Bioengineering is a broad title include work
on medical and agricultural systems itspractitioners include agricultural, electrical,
mechanical, industrial, environmental
Biological engineering is similar but emphasizes
applications to plants and animals
UEMK3233 Topic 1 36
Biochemical engineering
extension of chemical engineering principles to systems using a
biological catalyst to bring about desired chemicaltransformations
often subdivided into bioreaction engineering and bioseparation
engineering
Biomedical engineering
the application of engineering techniques to the understanding ofbiological systems and to the development of therapeutictechnologies and devices.
kidney dialysis, synthetic skin, artificial joints.. etc are some
products of biomedical engineering.
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UEMK3233 Topic 1 39
Approach to research
biologists vs engineers
fundamental training distinctly different
For biologist,mathematical theories and quantitative methods play 2nd
role
very strong with respect to laboratory tools (interpretation oflaboratory data from complex systems) BUT they are often
have incomplete backgrounds in mathematics
usually better at the formation of testable hypotheses,
experimental design, and data interpretation from complex
systems results are qualitative and descriptive
UEMK3233 Topic 1 40
Engineerusually possess a very good background in the physical and
mathematical sciences
theory formulate mathematical equation (predictive model)
and then the validity of the theory is tested by comparingpredicted responses to those in experiments
typically unfamiliar with the experimental techniques andstrategies used by life sciences
UEMK3233 Topic 1 41
The skills of engineer and life sciences are
complementary integration of skills
To become a bioprocess engineer:
needs a solid understanding of biology (need to take further
courses in microbiology, biochemistry, cell biology.)
need to have improvements in experimental tools
learn more advanced work in biochemical engineering
Biochemical engineering
extension of chemical engineering principles to systems using a
biological catalyst to bring about desired chemical transformations
often subdivided into bioreaction engineering and bioseparation
engineering
UEMK3233 Topic 1 42
How biologists and engineers
work together
Figure 1: Steps in development of a complete
bioprocess for commercial manufacture of a new
recombinant-DNA-derived product
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UEMK3233 Topic 1 43
6 Plasmids
2 Animal tissue
3 Part of animalchromosome
4 Gene cut fromchromosome
5 Microoganism such
as E. coli
1 biochemicals
7 Cut plasmid
8 Recombinantplasmid
9 Insertion int omicroorganism
10 Plasmid
multiplication
and gene
expression
11 Celldivision
12 Small scale culture
13 Bench-top bioreactor
14 Pilot scale bioreactor
15 Industrial scale bioreacto r16 Product recovery
17 Packaging and marketingUEMK3233 Topic 1 44
Steps in bioprocess development
The interdisciplinary nature of bioprocessing is
evident if we look at the stages of development
required for a complete industrial process
as shown in Figure 1, several steps are required to
convert the idea of the product into commercially
reality
these stages involve different types of scientific
expertise
UEMK3233 Topic 1 45
6 Plasmids
2 Animal tissue
3 Part of animalchromosome
4 Gene cut from
chromosome
5 Microoganism suchas E. coli
1 biochemicals
7 Cut plasmid
8 Recombinantplasmid
9 Insertion int o
microorganism
10 Plasmidmultiplication
and gene
expression
11 Celldivision
12 Small scale culture
13 Bench-top bioreactor
14 Pilot scale bioreactor
15 Industrial scale bioreacto r16 Product recovery
17 Packaging and marketingUEMK3233 Topic 1 46
Step 1 to 11
concerned with genetic manipulation of host organism a
gene from animal DNA is cloned into Escherichia coli
genetic engineering
done in laboratories on a small scale by scientists trained in
molecular biology and biochemistry experiment tools: petri dishes, micropipettes,
microcentrifuges, nano- or microgram quantities of
restriction enzymes, electrophoresis gels for DNA, protein
fractionation
Parameters of major importance stability of the
constructed strains and level of expression of the desired
productbioprocess development
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UEMK3233 Topic 1 47
Step 12 Small-scale cultivation (lab-scale) is mostly carried out
using shake flasks (250-mL to 1-L) to study the growth
and production characteristics of cells
culture environment (medium composition, pH,temperature..and other environmental condition) allow
optimal growth of microorganism and productivity of
product
practical skills microbiology, fermentation
calculated parameters cell growth rate, specific
productivity and product yield used to describe
performance of the organism
UEMK3233 Topic 1 48
Step 13 to 14 (scaling up)
once the culture conditions for production are known, scale-
up of the process starts
Bench-top bioreactor (cultures can be more closely
monitored in bioreactor than in shake flasks better
control over the process)
Lab scale 1- or 2-litre
Equipped with instruments for measuring and adjusting temperature,
pH, DO, stirrer speed, and many other process variables
Information collected: oxygen requirements of the cells, shear
sensitivity, foaming characteristics .
Identify the limitation imposed by the reactor on the activity of the
organismwhether or not the reactor can provide conditions for
optimal activity of the cells is of prime concern!!
UEMK3233 Topic 1 49
Step 14 Pilot-scale bioreactor
Engineers trained in bioprocessing are normally involved in
pilot-scale operations a vessel of capacity 100 to 1000 L
is built according to specifications determined from the
bench-scale prototype
Aim of pilot-scale studies to examine the response of
cells to scale-up
Changing the size of the equipment seems relatively trivial; however, loss or
variation of performance often occurs
Even though the geometry of the reactor, method of aeration and mixing,
impeller design and other features may be similar in small and large
fermenters, the effect on activity of cells can be great!
Loss of productivity following scale-up may or may not be recovered
Economic projections often need to be re-assessed as a result of pilot-scale
findings
UEMK3233 Topic 1 50
Step 15 (Industrial-scale operation)
This part of process development is clearly in the
territory of bioprocess engineering
As well as the reactor itself, all of the auxiliary
service facilities (air supply and sterilisation
equipment; steam generator; supply lines; medium
preparation; sterilisation facilities; cooling-water
supply. etc) must be designed and tested
Need to ensure the fermentation to be carried out
aseptically
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UEMK3233 Topic 1 51
Step 16 (Product recovery or downstreamprocessing)
After the production process, raw broth is treated in a seriesof steps to obtain pure product
Product recovery is often difficult and expensive
accounts for 60% or 80 90% of the total processing cost Actual procedures used depend on the nature of the product
and the broth physical, chemical or biological methods
Techniques applied industrially for downstream processingare first developed and tested using small-scale apparatus many operations which are standard in the laboratorybecome uneconomic or impractical on an industrial scale
Scientists trained in chemistry, biochemistry, chemicalengineering and industrial chemistry play important rolesin designing product recovery and purification systems
UEMK3233 Topic 1 52
Step 17 (Packaging and marketing)
After the product has been purified in sufficient purity, it
can be packaged and marketed
New pharmaceuticals products require medical and
clinical trials to test the efficacy of the product
Bioprocess engineers with a detailed knowledge of the
production process are often involved in documenting
manufacturing procedure for submission to regulatory
authorities manufacturing standards must be met,
particularly in the case for recombinant products where a
greater number of safety and precautionary measures is
required!
UEMK3233 Topic 1 53
As shown in this example, a broad range of
disciplines is involved in bioprocessing
Scientists working in this area are constantly
confronted with biological, chemical,
physical, engineering and sometimes
medical questions!!
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