INTRODUCTION OF BIOPROCESS ENGINEERING: HISTORY OF DEVELOPMENT AND STEPS

1. Define the concept of bioprocess engineering and its scope (lấy cái scope của gemini t thấy ổn
á)
- Bioprocess Engineering is a specialized field within biotechnology that focuses on the design,
development, and optimization of processes for manufacturing biological products
- Scope: It applies engineering principles to translate laboratory-scale biological discoveries into
industrial-scale production. This includes designing and operating bioreactors, separation and
purification systems, and other equipment necessary for biomanufacturing.
2. Describe the history of bioprocess engineering development with a focus on microbial products
- Bioprocess engineering started with ancient fermentation methods for making beer, bread, and
cheese. Early scientists like van Leeuwenhoek and Pasteur revealed that microbes drive these
processes. The breakthrough came with penicillin production during WWII, which led to
large-scale fermentation. In the 1970s, recombinant DNA technology enabled engineered
microbes to produce complex products like human insulin. Today, advanced bioreactors and
metabolic engineering optimize microbial systems to produce pharmaceuticals, biofuels, and
chemicals sustainably.
3. Describe the general steps for microbial product manufacturing

OVERVIEW ON IMPORTANT ASPECTS OF MICROBES, METHODS FOR MICROBIAL

CLASSIFICATION, AND CHARACTERISTICS OF INDUSTRIAL MICROORGANISMS

1. Describe important physiological characteristics of microbes

There are four requirement characteristics of industrial microbes including modes of nutrition and
nutrient, oxygen, temperature and pH.
- Modes of nutrition in microbes:
 - Nutrient requirements:
Macronutrients: C, and N sources and concentrations, P, S, K, Mg, Ca, Na
Micronutrients (trace elements): Fe, Cu, Mn, Se, Zn,…
Growth factors: vitamins (e.g. biotin, B1, riboflavin), amino acids, purines, and
pyrimidines.
- Oxygen requirements:
Aerobe: require oxygen
Microaerophile: grow only oxygen reduce from that present in air
Facultative: grow in either oxygen presence or absence
Anaerobe: inhibit oxygen, cannot use oxygen
Obligate anaerobe: cannot grow in the presence of oxygen

a. Aerobic
b. Anaerobic
c. Facultative
d. Microaerophilic
e. Aerotolerant anaerobe

- Temperature requirements:

Psychrophile Mesophile Thermophile Hyperthermophile

Growth ⩽ 15oC Between 20 and Between 45 and ≥ 80oC

temperature 45oC 80oC
optimum
Maximum < 20oC

Widespread in
nature
 - pH requirements:

Acidophiles Alkaliphiles

Growth - pH Typically below pH 6 Optima of 9 or higher

Sometime pH 11

Example Many fungi grow best at pH 5 or Alkaliphilic microorganisms

below, and a few grow well at found in soda lakes and high
pH values as low as 2. carbonate soils (highly alkaline
habitats)
Most well-studied Bacillus
species, such as Bacillus firmus.

2. Describe important features of industrial microbes and possible strategies for strain
enhancement
- Important features:
+ Microbial identification:
Morphological method: morphological characteristics (colony, cell, spore, spore bearer,
fruiting bodies, ect..)
Physiological method: taste, smell, pigment (for mushrooms), specific enzymes and
substances (for bacteria or fungi)
Molecular method: 16s rDNA (bacteria), 18s rDNA (algae & fungi), ITS (fungi), ect…
+ Industrial important species: traditionally bacteria and fungi
+ Criteria for selection of industrial microbes:
Suitable nutritional characteristics depend on the value of product
The optimum temperature, optimum above 40oC → reducing cooling-costs of a
large-scale fermentation
Suitability process and equipment for reaction of the organisms
The stability of the organism and its amenability to genetic manipulation.
The productivity of the organism, measured in its ability to convert substrate into product
and to give a high yield of product per unit time.
The ease of product recovery from the culture
+ Microbial isolation: from culture collection or natural environment
Culture collection Natural environment

Their cultural requirements can be anticipated cheaper, but more time consuming to get the
using whole descriptions of taxa and genome desired organisms
sequencing
Ex: National Collection of Type Cultures (NCTC)
website
National Collection of Yeast Cultures (NCYC)
website

- Possible strategies for strain enhancement:

+ Screening potential strains
+ Optimize culture medium (cultivate condition) and growth condition
+ Genome modification:
Natural variants: e.g. Mycelial organisms may be that most new isolates are probably
heterokaryons (contain more than one type of nucleus) and the selection of the progeny of
uninucleate spores results in the production of homokaryons (contain only one type of
nucleus) that may be superior producers.
 Mutation, UV light (can kill microorganisms), Radiation, Chemical (easy method) ,
Physical,...
+ Recombinant: protoplast fusion
+ Recombinant DNA technology → Cloning
+ Genome sequencing
3. List possible methods for microbial culture preservation
- Storage on agar slopes:
+ Stored in a refrigerator (5°C) or/and subculture at approximately 6-month intervals.
+ Sometimes, the slopes are covered with sterile medicinal-grade mineral oil.
- Cryopreservation - storage below -135oC
+ In liquid nitrogen or a freezer.
+ Cryoprotective agent: Glycerol
+ Microbial cultures for cryopreservation are grown into the stationary phase (to
induce the protective general stress response) whereas those of cell cultures are
harvested at the mid-exponential phase, to prevent the initial apoptosis
- Dried cultures:
+ Soil can be dried and kept at room temperature or in a refrigerator
+ Alternatives: Silica gel and porcelain beads
- Lyophilization:
+ Freezing of a culture followed by its drying under vacuum, which results in the
sublimation of the cell water.
+ Protective medium: milk, serum, or sodium glutamate.
+ Transferred to an ampoule → freeze → until sublimation complement → seal

FERMENTOR

1. Describe the components and main functions of the fermentor

a. Functions
- Provide operation free from contamination
- Maintain a specific temperature
- Provide adequate mixing and aeration
- Control pH of the culture
- Allow monitoring and/or control of dissolved oxygen
- Allow feeding of nutrient solutions and reagents
- Provide access points for inoculation and sampling
- Minimize liquid loss from the vessel
- Facilitate the growth of a wide range of organisms
b. Components

Vessel (container): contain culture medium, microbes’

growth
Motor: provide stirring power for mixing
pH probe: measure pH and control of pH
O2 probe: identify O2 and control of dissolve O2
Impellers: stirring to disperse the incoming air
effectively
Temperature sensor and control unit (Platinum
thermometer sensor) – for setting desired temperature
Cooling jacket: cover vessel and control temperature
Air filter: take air inside (O2 for aerobe, N2 for
anaerobe)
Sample line: take sample to check concentration,
parameter, size
Harvest line: collect production or connect with other vessel for continuous fermentation
 i. Basic components include motor, pump, gas control, etc.
ii. Vessel and accessories
1. Vessel
- General used: glass, stainless steel
- Accessories: Gas pump, impellers, platinum thermometer sensor and heater pad or coil,
pH probe, foam probe
iii. Peripheral equipment such as reagent bottles
- Reagent pumps (pH and antifoam control)
- Medium feed
- Rotameter/ gas supply
- Sampling device
iv. Instrumentation and sensors
- Antiform control
- Speed control
- Temperature control

TYPICAL FERMENTATION PROCESS AND MODES OF FERMENTATION

1. Describe the typical fermentation process

2. Compare different modes of fermentation and when to use them

Purpose Advantages Factors Problem Application of

Affecting SSF associated with SSF
SSF

Solid-state - occurring in - The process - Selection of Microbial - Agro-food

Fermentati the is simple microorganisms activity industry:
on (SSF) absence/near - Cost-effective : filamentous produces heat, - Agriculture
absence of - Less effluent fungi have but the material - Industrial
water released, shown better they grow in fermentation
- Employing reduces growths in doesn’t transfer
natural pollution solid substrates heat well,
substrates such - High product -Substrate: causing slow
 as rice bran, yields provides both cooling. Too
sugarcane - Simple physical much heat can
bagasse, fermentation support and damage the
sawdust equipment nutrients (rice product, slow
- Techniques - Easier bran, sawdust, growth, and
involving fungi downstream sugarcane lower yield
and processing bagasse) for and quality.
microorganisms the microbes
require less - Process
moisture optimization
content

Purpose Advantages Factors Types of SmF Application

Affecting SmF

Submerged - utilizes free - Purification of Substrates: Batch Penicillin

Fermentation flowing liquid products is soluble sugars, fermentation production
(SmF) substrate for easier Molasses, Continuous Citric acid
fermentation Liquid media, Semi-continuous production
- best for Fruit and Feed batch
microbes e.g. vegetable
bacteria which juices,
require high Sewage/waste
moisture water
content Microbes:
- substrates require high
are utilized moisture
more rapidly content
- used in the (bacteria,..)
extraction of
secondary
metabolites
that need to be
used in liquid
form.
-Agitation is
always
essential

Batch Continuous
Fresh medium Not add Add
Closed culture system
Contamination Low High
Productivity Non-productive down time High and efficiently
productivity
Genetically engineered Degeneration of biocatalyst
biocatalyst (Control genetic
stability)
Product quality Different types of product Same products from time to
time
Setup Simple Complex
Accumulation of inhibitory Yes No
products
(Tích tụ của các sp ức chế)
Cost Cheap Expensive

Batch Fed-batch Continuous Semi-continuou

s

Definition Closed culture The nutrient is Open system, Simultaneous

system continuously nutrient addition
contains added and outflow
limited nutrient intermittently to Once system in withdrawal are
medium initial a batch culture equilibrium: carried out
and then chemostat inermittently
without the volume, cell
inoculated
removal of number, 2 types:
cells.
culture fluid. nutrient remain
constant Cyclic-continuo
us;

Cell reuse

In and out Once According to Constant Cyclic-continu

fresh operation cẻtain feed volumn ~ fresh ous (theo chu
medium started: no strategy medium added kỳ) complete/
inlet or outlet = removed near complete
Generally, two culture medium fermentor à
If fermentation basic types of at the same harvest = fresh
aerobic fed batch are rate medium add,
system should variable (continuously) but reduce time
be gas inlet volume, and => until
(aeration) & fixed volume eventually the
gas outlet intermittent
(exhaust) feeding
becomes
continuous

Cell reuse: cell

centrifuge from
the fermentation
broth and used
to reinoculate
fresh medium (It
is continuous
only in the
sense that cells
are reused; in
essence it is a
batch
fermentation)