CHE323

Introduction to Biochemical Engineering 1

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 Definitions
• Biochemical Engineering involves the application of chemical engineering principles and
approaches to biological based systems and processes.
-It is central to the areas of environmental engineering, and to biotechnology processes
that produce pharmaceuticals, fine chemicals and genetically engineered products.
-Study of the biological and biochemical principles supports the field of biochemical
engineering.
-A Biochemical Engineer is someone who when with engineers, talks biology, when with
biologists, talks engineering and when with biochemical engineers, talk politics.
• Bioprocessing is any process in which microbes or living organisms or enzymes play a vital role
in getting transformation of the feed into useful products.
-Examples are converting milk to curds, fruit juices into wines, sugars into alcohols.
-It’s a branch of chemical/process engineering dealing with design and development of
equipment and processes that use biological components such as enzymes or
microorganisms
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 Biochemical Engineering is a multidisciplinary field:

Material science
Chemical engineering

Biology:
microbiology,
molecular biology
Biochemical
Engineering
Pharmacology
Biochemistry

Chemistry Genetics

Medicine

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 See Table in the lecture note
for some stages of development
in bioprocessing (fermentation)

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 Related disciplines/areas of specialization
• Metabolic engineering including metabolic modeling, bioprocess
engineering, process control, bio-separation, bio-informatics, bio-material
engineering, tissue engineering, manufacture engineer, reactor design.
• Biochemical engineering covers research at the scale of the biocatalyst
development (microbe, insect cell, mammalian cell, plant cell, enzyme)
• Biotechnology is a study about biological systems or living organisms to
make or modify products or processes for specific use, such as in agriculture,
food production, and medicine.

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Traditional and modern application of biotechnology
• They are in the areas of food, bakery products and alcoholic
beverages.
• Earliest application was in the production of wine from fruit juices.
• Areas of health and hygiene for the production of vaccines, enzymes,
various fermented foods, organic acids etc.
• Bio-processing or biotechnology operations are interrelated with a
large number of faculties in science.
• Alcoholic fermentation was considered to involve both chemical and
microorganisms.
• The biochemical engineers were considered to provide sterile
environments in most bio-processing.
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 Unit operations in bioprocesses
The whole bio-processing operations can be broadly classified into two
classes/types of operations.
 Fermentation
 Product recovery and effluent treatment.
• Fermentation is biochemical operation and can be classified as unit process.
• Operations carried out on the fermentation broth for product recovery and
downstream operations including effluent treatment may be classified as unit
operation.
• Some of the operations carried out on the fermenter such as fluid dynamics,
mixing, air sparging, heat transfer to the fluids may also be categorized as unit
operations

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• Product recovery and downstream processing lines involve mixing and separations of
various phases, streams and fluids.
We can broadly comprehend the systems encountered as
 Solid-liquid
 Liquid-liquid
 Liquid-liquid-solid
 Liquid-gas
 Solid-liquid-gas
 Solid-solid
Mixing is also one of the most important unit operations in bio-processing. It could be for
• G-L, L-L, L-S, G-L-S systems.
• Design o various impellers/mixers for mixing is a challenge for biochemical engineer.
• Efficient mixing of the broth decides the progress of reaction.
- for example, mixing of gas in a fermentation broth is useful for the utilization of microbial
cells to respire, grow, increase in size/multiply, and perform well.

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Outline of integrated bioprocess
Integrated bio-processing consists of various steps:

Step 1: Isolation of the strain

• Identification and isolation of the microorganism which brings out
the desired bioconversion.
-This is generally the task of a microbiologist.
-It involves some shake flask experimentation, etc.
• The techno-economic viability of the process will be taken up
subsequently if the results of this step 1 are encouraging.

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Step 2: Preservation of the strain
• The strain isolated in step 1 will be preserved for future use.
• Its ability to perform and yield the products should also be preserved.
The strain can therefore be stored ;
a) Refrigerated temperature (2 – 6 oC)
b) At frozen storage temperature (-18 to -80 oC)
c) By freeze drying (lyophilization)

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 Step 3: Growth of inoculums
• Before using the strain in a fermenter/bioreactor, the inoculum (cells
added) is cultured
for growth.
• The preserved strain/culture is revived by growth in shake flasks or by
solid-state fermentation on the solid surfaces.
• Solid state fermentation(SSF) is a method of growing microorganisms in an
environment of limited moisture without having free flow of water.
-In other words, microorganisms grow on a solid surface which is moisten
bed and which has also got free access to air.

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Step 4: Pre-fermentation culturing
• Before the cells are admitted into the fermenter for performing
bioconversion, they are initially grown separately with nutrients and the
substrate so that the cells can multiply and proliferate.
• This will help increase the cell density. The optimal cell concentrations in a
fermenter are as follows:
1) Bacterial: 0.1 – 3.0%
2) Actinomycetes: 5 – 10%
3) Fungi: 5 – 10%

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Step 5: Fermentation
• Major activity in industrial operation in the whole bioprocess is FERMENTATION.
• Fermentation is the heart of the bio-processing operation.
• Fermenter sizes vary from 1 to 450 m3, depending upon the type of
fermentation process.
Based on capacity, tonnage and nature of the
fermentation product, fermenters can be
classified;
1. Batch fermenter.
2. Fed-batch fermenter.
3. Continuous fermenter.
The conditions could be aerobic (with bubbling
of air) or anaerobic (in the absence of air).

A Fermenter
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Step 6: Recovery and purification of the product
• Recovery and purification sometimes decide the economic viability of the
process.
• The cost of recovery can vary anywhere between 20 and 60% of the total
manufacturing costs.
The methodology depends on;
-Nature of the cells.
How to hold the products : is it intracellular (product is held within the cells) or
extracellular (the product is excreted from the cells).
-Solids can be removed by simple filtration or centrifugation techniques.
-The intracellular material is obtained by cell disruption techniques for example
intracellular enzyme extraction
-Purification and isolation of the final products are carried out using different
unit operations.
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Step 7: Treatment of effluents
• Effluents are released during fermentation processes.
• The effluents may be rich in organic matter and may be a potential hazard for
the environment if released like that.
• Ability to economically be able to treat effluents before release to the
environment makes or mars the viability of the product manufacture.
• Various effluent-treatment techniques can be classified as:
-Physical
-Chemical
-Biological
• The final choice depends upon the individual cases and local
circumstances.

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 A process for converting biomass to ethanol
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 Microbiology
• Microbiology is the study of microorganism which are not only microscopic and exist as
single cells, but also ultramicroscopic organism which are not cellular and hence cannot exist
independently e.g. viruses
-It deals with study of functioning of cells, their diversity and evolution, their interaction
with environment, other living organism and man.
• Microbiology forms the basis for the complimentary development in cell biology,
biochemistry, molecular biology and genetics.
• Microbiology has evoled as an important branch of science, and is studied with respect to 2
major aspects:
-As basic biological science by providing a system to understand the nature of life
processes, the principle behind it, and the genetics which is involved
-As applied biological science, microbiology deals with the study of useful
microorganisms as well as that of pathogenic organisms.
***See the lecture notes for different past discoveries
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 Biochemistry
• It is the study of life cyclic processes in terms of chemicals.
• It is not the study of chemistry of living organisms.
• It obtains information from the basic microorganisms viz., bacteria, viruses,
algae and fungi.
• It draws information on energetics from basic thermodynamics.
• Some of the chemical/biochemical reactions in the living organisms are
facilitated by another type of compounds known as enzymes. This facilitation is
known as catalysis.
• Enzymes are therefore known as biocatalysts or biological catalysts.
• Cells themselves contain some of the enzymes.
• Cells regulate the type of enzyme and its number by which it regulates both the
type of biochemical reaction and its rate.
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• Genetic engineering deals with the information flow of the living organisms.
• Biochemistry, coupled with genetic engineering principles play a vital role in
devising techniques for manipulating most of the biochemical
transformations.
• Living organisms contain various biomolecules which are the building blocks of
the cell and also help in storing and releasing energy for biotransformations.
• Biomolecules include carbohydrates, lipids, proteins, nucleic acids, vitamins,
etc.

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 Introduction to Microbiology
Branches of Microbiology
1. Medical microbiology
• Study of pathogenic (disease causing) organisms and ways to eliminate them. Common diseases such
as typhoid fever, tuberculosis (bacterial), hepatitis C (viral), malaria (parasitic), dandruff, candida
(fungal infections)
2. Agricultural microbiology
• Study of plant disease, understanding beneficial interactions with plant systems, like soil fertility, crop
protection, and increasing yield.
3. Environmental microbiology
• Study of the relationship of microorganisms with its habitat, pollution effect, and its impact on
environment from the stand point of ecological balance and health
4. Food & Dairy microbiology
• Study of microorganisms that produce various food and dairy products; study of control of
microorganisms in food and transmission of food-borne diseases.
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 Microscopy
• Microscopy is an important tool for revealing the secrets of the world of
small creatures.
• Microorganisms and their structural components are measured in smaller
units such as micrometers or microns (10-6 m), nanometers or millimicrons
(10-9 m), and angstroms (10-10 m).
• Microscopic resolutions are limited by the size/type of the microbe. Some
microbes are unicellular, algae, fungi, protozoa, viruses, proteins, lipids, small
molecule.
• Operation ranges could vary from unaided eye, to light microscope, and to
electron microscope.

***see the lecture notes for different ranges of microscopic resolutions

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 Microbial taxonomy
• Taxonomy is the subject of classification of biological existence of life species.
• Earlier, all living organisms were classified under two main kingdoms, namely, Plantae and
Animalia by Swedish botanist Carolus Linnaeus in the 18th century.
• In 1969, Robert H. Whittaker proposed a 5-kingdom classification:
1. Monera
2. Protista
3. Fungi
4. Plantae
1. Animalia
A term for generalizing the taxonomical classification of microorganisms is UNIVERSAL
ANCESTOR which can also be referred to as Universal Phylogenetic Tree (see next slide).
It is referred to as Last Universal Common Ancestor (LUCA), it’s the most recent population of
organisms from which all organisms now living on earth have common descent

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***see the lecture notes for the simplified classification of universal ancestor

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Monera: true bacteria and blue green algae.
Prostita: eucaryotes such as protozoa which are unicellular, contains chlorophyll
(plant like) and possess movement (animal like).
Fungi: include non-green eucaryotic organisms like slime moulds, bread moulds,
sac fungi.
-Composition of their cell wall differed from that of plants, and had adjacent
cells without separation, hence are not considered as true multicellular
organism
Plantae: include algae, all mosses, ferns, cornifers, and flowering plants.
Animalia: Sponges (are primitive multicellular animals, incapable of moving),
worms, insects, and vertebrates

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Microbial diversity
• Evolution roots of cell form a basis for understanding the microbial
diversity.
• Microbial diversity can be seen in terms of variations:
-in cell size,
-morphology (internal structure),
-metabolism (chemical reactions in the body's cells that change
food into energy),
-adaptation (adjusting or changing to become more suited to an
environment),
-growth (increase in size or concentration) etc.

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 Microbial Nomenclature
• Basic taxonomic unit is species which can be defined as a collection of similar
strains.
• Groups of species are collected as genera. They share major properties.
• Groups are classified into families, families into order, and orders into divisions.
Following the binomial system of nomenclature, microorganisms are given genius
and species name, which are either Latin or Greek derivations.
Example;
• Bacillus subtilius or Bacillus meaning “rod shape”, which indicates its slender nature;
• B. cereus meaning “waxen”(shape can easily change);
• B. stearothermophilus meaning “heat loving”;
• B. acidoceldarius meaning “acid-thermal”
They are written in italics with the genus name stating with capital letter and the species name
with a small letter.
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There are many ways of grouping (classification) of procaryotes.
• The characteristics of taxonomic approach include morphology,
-Gram reaction (Gram reaction is a classical technique to identify the basic difference in
the cell wall structure of the bacteria.
-The cells are stained with crystal violet dye, and then treated with iodine solution,
and then cells washed with alcohol.
• Those which retain the blue crystal violet colour are classified as gram positive, and
those which do not retain the colour are called as gram negative species),
• Other approaches for group classifications are:
 nutrition, cell wall chemistry, pigment, storage products, ability to use various substrates,
fermentation products, gaseous needs, temperature, pH requirement, tolerances, sensitivity.

Molecular taxonomy includes guanine and cytosine ratio (GC ratio) in organisms, the extent of
DNA: DNA hybridization, amino acid sequencing and protein analysis.

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 Chemical composition
• Cells are composed of small molecules as well as macromolecules.
• They are made up of 4 basic elements, viz. Carbon, Oxygen, Hydrogen and
Nitrogen.
-Phosphorus, Iron, Sulphur, Zinc, Manganese, Copper, Molybdenum, and Cobalt
are also present apart from the 4 basic elements.
• Water accounts for 90% of the weight of the cells.
• Of the macromolecules, proteins are most abundant by weight (55%).
• Most procaryotes require an organic compound of some sort as their source of
carbon.
• After carbon, the most abundant element in cell is nitrogen (12% by dry
weight), which is a major constituent of protein and nucleic acids.
• Nitrogen is found in organic and inorganic forms, and is mostly assimilated as
ammonia NH3, nitrates NO3 or N2.
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• Organisms assimilate various organic compounds, and use them to make new cell wall
materials, which may be amino acids, fatty acids, organic acids, sugars, nitrogen bases
and aromatic compounds.
• Macronutrients required in nature and media for organisms include Carbon, Hydrogen,
Oxygen, Nitrogen, phosphorus, Sulphur, Potassium, Magnesium, Sodium, Calcium, Iron.
• Micronutrients required by cells include; Chromium, Cobalt, Copper, Manganese,
Molybdenum, Nickel, Selenium, Tungsten, Vanadium, Zinc, Iron.
• GROWTH FACTORS
-They are the organic compounds required in small amounts for growth and
metabolism, and these include vitamins, amino acids, purines, pyrimidines.
-Vitamins are not only needed for growth, but also play an important role as co-
enzymes. Most of the vitamins are synthesized by organisms themselves, but some
depends on external sources.
***see the lecture notes for more macronutrients and the chemical forms in which they are
supplied.

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