Abag, Mary Eisle E.

Acosta, Charrisma A.
Beron, Danica D.
Camsol, Kristina C.
Biochemistry is the study of a chemical
process in living things.

A process that uses living cells or
biomolecules to carry out a chemical
transformation leading to the production and
ultimate recovery of valuable products.

Biochemists study microorganisms like
proteins and carbohydrates.
The category fine chemicals within the context
of biotechnology broadly interpreted to
bioproduction of high value molecules and
bioconversion of high value starting materials
to yet more valuable products.

Some products of biotechnology:

1. Enzyme
2. Antibiotics
3. Recombinant DNA Technology

Enzymes are usually proteins of high
molecular weight that act as catalysts.
Enzymes are specific, versatile and very
effective biological catalysts, resulting in
much higher reaction rates as compared to
chemically catalyzed reactions under
ambient conditions.
More than 2000 enzymes are known.

Enzymes lower the activation energy of the
reaction catalyzed by binding the substrate
and forming an enzyme substrate complex.
Enzymes do not affect the free energy
change or the equilibrium constant.
The interaction between the enzyme and its
substrate is usually by weak forces. In most
cases, van der waals forces and hydrogen
bonding are responsible for the formation of
ES complexes.
Among various enzymes produced at large
scale are proteases, isomerases and oxidases.
These enzymes are produced using
overproducing strains of certain organisms.
Separation and purification of an enzyme
from an organism require disruption of cell,
removal of cell debris and nucleic acid,
precipitation of proteins, ultrafiltration of
the desired enzyme, chromatographic
separations (optional), crystallization and
drying.
The first step in the large scale production of
enzyme is to cultivate the organisms
producing the desired enzyme.
Proteases are produced by using overproducing
strains of Bacillus aspergillus , Rhizopus and
Mucor;
Pectinaeses are produced by Aspergillus niger;
Lactases are produced by yeast and aspergillus
Lipases are produced by certain strains of yeast
and fungi;
Glucose isomerase is produced by Flavobacterium
arborescens or Bacillus coagulans
After the cultivation step, cells are
separated from the media usually by
filtration or sometimes by centrifugation.
Depending od intracellular or extracellular
nature of the enzyme either the cells or the
fermentation broth is further processed to
separate and purify the enzyme.

Proteases are used in:
food processing such as cheese making (rennet),
baking, meat tenderization (papain, trypsin), and
brewing (trypsin, pepsin)
in detergents for the hydrolysis of protein stains
(subtilisin carlsberg)
tanning and the medical treatment of wounds


Enzymes are commonly used in medicine for
diagnosis, therapy, and treatment purposes.
Trypsin and Streptokinase can be used as an anti-
inflammatory agent.
Lysozyme which hydrolyses cell wall of gram-positive
bacteria, is used as an antibacterial agent
Urokinase is used in dissolving and preventing blood clots

They are specific in their action and are
therefore less likely to produce unwanted
byproducts.
They are biodegradable and therefore cause
less environmental pollution.
They work in mild conditions, i.e. low
temperatures, neutral pH and normal
atmospheric pressure, and therefore are
energy saving.
They are highly sensitive to changes in
physical and chemical conditions surrounding
them.
They are easily denatured by even a small
increase in temperature and are highly
susceptible to poisons and changes in pH.
Therefore the conditions in which they work
must be tightly controlled.
The enzyme substrate mixture must be
uncontaminated with other substances that
might affect the reaction.
Enzymes are used in other many other
industries including textile, paper and
leather manufacturers. Experts state that
harsh chemical treatments used in these
industries' production can present potential
hazards to staffers.
The U.S. Department of Agriculture reports
that using commercial enzymes to produce
grain ethanol can reduce greenhouse gas
emissions associated with the manufacture of
ethanol.


There are significant municipal concerns
relating to the use of surfactants and
enzymes in the process of wastewater
treatment.
Many municipalities across the country are
banning the use of surfactants and enzymes
entering wastewater treatment because of
the harmful effects these products have on
the sewer lines and the damage is costing
cities much money in repair.
The Greek word anti means "against", and
the Greek word bios means "life" (bacteria
are life forms).
An antibiotic is a drug that kills or slows the
growth of bacteria.
These are used to treat infections caused by
bacteria.
Although there are a number of different
types of antibiotic they all work in one of
two ways:
A bactericidal antibiotic kills the bacteria.

A bactericidal usually either interferes with the
formation of the bacterium's cell wall or its cell
contents.

Penicillin is a bactericidal.

A bacteriostatic stops bacteria from multiplying.

An antibiotic is given for the treatment of an
infection caused by bacteria. Antibiotics
target microorganisms such as bacteria, fungi
and parasites. However, they are not
effective against viruses. If you have an
infection it is important to know whether it
is caused by bacteria or a virus.

The industrial production of penicillin was
broadly classified in to two processes
namely:

Upstream processing
Downstream processing


UPSTREAM PROCESSING:
Upstream processing encompasses any
technology that leads to the synthesis of a
product. Upstream includes the exploration,
development and production.

SELECTION & IMPROVEMENT OF STRAIN
INOCULUM PREPARATION

DOWNSTREAM PROCESSING:
The extraction and purification of a
biotechnological product from fermentation
is referred to as downstream processing.



CARBON SOURCES:
Lactose acts as a very satisfactory carbon
compound, provided that is used in a
concentration of 6%. Others such as glucose &
sucrose may be used.

NITROGEN SOURCES:
Ammonium sulphate and ammonium acetate can
be used as nitrogenous sources.

MINERAL SOURCES:
Elements namely potassium, phosphorus,
magnesium, sulphur, zinc and copper are
essential for penicillin production. Some of these
are applied by corn steep liquor.

FERMENTATION PROCESS:

The fermentation process is usually carried out
in a large fermenter. The fermentation
medium is formulated and fed in to the
fermenter. The inoculum is also maintained
properly.
SOLID LIQUID SEPARATION:
The first step in product recovery is the
separation of whole cells and other insoluble
ingredients from the culture broth. Several
methods such as floatation, flocculation etc, are
used to serve this purpose.
CONCENTRATION:
This is a step followed to concentrate the
desired product. The methods used to
concentrate includes adsorption, precipitation
etc.
PURIFICATION:
Chromatographic techniques are generally used
to purify the product. As concerned with the
production of antibiotics ion exchange
chromatography seems to be the better option.
EXTRACTION AND PURIFICATION OF PENICILLIN:
There are some special techniques used for the
extraction of penicillin from the fermentation
medium. They are as follow:

REMOVAL OF MYCELIUM
COUNTERCURRENT SOLVENT EXTRACTION OF
PENICILLIN

TREATMENT OF CRUDE EXTRACT:
The solvent extract recovered in the previous
stage is carefully extracted back with aqueous
sodium hydroxide. This is followed by charcoal
treatment to eliminate pyrogens and by
sterilization





Researchers describe an approach that is
more efficient--and environmentally friendly-
-in developing new antibiotics, those needed
to kill the increasing number of infections
resistant to multiple drugs.
One way to increase the number of
antibiotics for fighting infections is to start
where nature stopped and engineer the
enzymes to produce new molecules, and thus
new antibiotics.
The interdisciplinary team of scientists,
including research professors David H.
Sherman and Janet L. Smith from the
University of Michigan's Life Sciences
Institute and Fecik of University of Minnesota
College of Pharmacy, is the first to crystallize
an enzyme in the process of closing the
antibiotic ring, which illustrates exactly how
the ring is formed.
Researchers describe a method that can be
used to get the bacteria itself to produce
new compounds that turn into the ring
structure and may be useful as new drugs.

Typical drug development involves chemical
manipulations that result in chemical waste,
which can be difficult to dispose of and is
hazardous to the environment.
This research implies it is realistic to develop
a more environmentally friendly way to
discover more potential drug compounds
with less chemical manipulation, and thus
less chemical waste.

Benefit to Animal Welfare

Banning or severely restricting the use of
antimicrobials in animals may negatively
impact a veterinarians ability to
protect animal health and prevent
suffering from disease, which can lead
to poor animal welfare.


Food safety begins with healthy animals,
continues in the harvesting process with good
hygiene and application of hazard
analysis/critical control point regulations,
and extends to the packing and handling of
the food product in both the market and the
home. All of these steps work in tandem, and
skipping any one of them increases the risk
of foodborne illness.


Antibiotics are used in animal feeds to increase
animal weight, increase efficiency of feed
utilization, increase reproductive efficiency
and decrease morbidity and mortality. These
benefits to animals and animal producers are
reflected in decreases in food costs to humans.
There are also benefits to human health from
use of antibiotics in food animals. By reducing
the incidence of animal health problems, use
of antibiotics in food animals reduce the
transference of animal infections to humans.

The contention that the effectiveness of
penicillin and tetracycline for use in human
medicine is rapidly diminishing as a result of
the proliferation of resistant bacteria caused
by subtherapeutic use of antibiotics in animal
production is not supported by experimental
data.

Rather, the evidence suggests that a fairly stable
level of resistance of the intestinal bacteria in
humans has long since been established to
penicillin and tetracycline as it has been in
animals.
Heavy indiscriminate use of antibiotics in
poultry (meat, dairy products) production
may result in the transfer of residual
antibiotics to human food and the transfer of
resistant bacteria to humans.

The " resistant gene " of mutant may be
transferred to another pathogenic species
thus spreading a health hazard.



Most compounds or at least most groups of
compounds acting via the same mechanisms are
found in hospital effluent and in some cases
even in municipal sewage in concentrations that
are high enough to warrant further risk
assessment and risk management.

The emission of antibiotics into the environment
should be reduced as an important part of the
risk management.

For this reason, unused therapeutic drugs should
not be flushed down the drain and physicians
must be made aware that antibiotics are not
completely metabolized by patients.

The environmental significance of
therapeutic drugs, disinfectants and
diagnostics should be included in the
undergraduate curricula of medical students
and pharmacists.

Antibiotics act as an ecological factor in the
environment that could potentially affect
microbial communities.
The effects include phylogenetic structure
alteration, resistance expansion, and
ecological function disturbance in the micro-
ecosystem.
Numerous studies have detected changes of
microbial community structure upon addition
of antibiotics in soil and water environment.
Effects of antibiotics on ecological functions
have also been discovered, including
nitrogen transformation, methanogenesis,
and sulfate reduction.

Also known as in vitro recombination, is a
technique involved in creating and purifying
desired genes.
The process of in vitro recombination makes
it possible to cut different strands of DNA
with a restriction enzyme and join the DNA
molecules together via complementary base
pairing.
Emerged as a response to the need for
specific DNA segments in amounts sufficient
for biochemical analysis.

Recombinant DNA and biotechnology can be
used to form proteins not normally produced in
a cell. In addition, bacteria that carry
recombinant DNA can be released into the
environment to increase the fertility of the
soil, serve as an insecticide, or relieve
pollution.

The technology of recombinant DNA has been
made possible in part by extensive research
on microorganisms during the last century.
One important microorganism in recombinant
DNA research is Escherichia coli (E. coli).
The biochemistry and genetics of E. coli are
well known, and its DNA has been isolated
and made to accept new genes.
The DNA can then be forced into fresh cells
of E. coli, and the bacteria will begin to
produce the proteins specified by the foreign
genes. Such altered bacteria are said to have
been transformed.
Arthur Kornberg
Replication of viral DNA in a breakthrough that
served as a proof-of-concept for cloning.
Werner Arber
Discovery of restriction enzymes in a bacteria
that degrade foreign viral DNA molecules while
sparing their own DNA.
Ligase could be used to glue DNA molecules.
Herbert Boyer and Stanley Cohen
founded the first company, Genetech, to work
with rDNA in its drug development labs
Insertion of rDNA into bacteria in such a way
that the foreign DNA would replicate naturally


Some of the molecular biology techniques
utilized during recombinant DNA include:

The study and/or modification of gene
expression patterns
Gene cloning
DNA sequencing
Creation of transgenic plants and animals
A summary of the process of making recombinant DNA:
Treat the DNA taken from both sources with
the same restriction enzyme, which cuts
both molecules at the same site.
The overhanging single-stranded ends of the
cut are called sticky ends, which pair with
its complementary sticky end, when mixed.
DNA ligase is used to covalently link the two
strands into a molecule of recombinant DNA.
The DNA is replicated many times.

Transformation
Host cells are bacteria
Non-bacterial transformation
Host cells are not bacteria
Phage Introduction
Host cells are phage
Development of cure for genetic and
acquired diseases
Development of DNA vaccines
Development of cure for infertility
Development of genetically modified plants
Has the potential to provide more food to
the world
Improved medicines
Improved livestock
Lowering the cost of medicines
Safety concerns
Environmental concerns
Ethical dilemmas
Potential for experimental abuse
Germline treatment going from treating
diseases to a method for picking the traits
you want in a child.

The sharp bands give the wrong impression
that the molecules are perfectly separated.
The sticky ends can degrade.
It is possible that the plasmids werent cut
completely
A small portion of the plasmid was not even
cut at all.


Economic and Access Concerns
Environmental and Monitoring Concerns
Allergenicity
Antibiotic Resistance Transfer
Concerns with Naturally Occurring Toxicants

Recombinant vaccines
Prevention and cure of sickle cell anemia
Prevention and cure of cystic fibrosis
Production of clotting factors
Production of insulin
Production of recombinant pharmaceuticals
Plants
Germ line and somatic cell gene therapy

Agriculture
Genetically modified crops, pesticide resistant
crops
Pharmacology
Artificial insulin production, drug delivery to
target sites
Medicine
Gene therapy, antiviral therapy, vaccination,
synthesizing clotting factors
Others:
Fluorescent fishes, glowing plants, etc.
Pathogenicity
The potential ability of living organisms and
viruses to cause disease in man, animals and
plants.
Safe handling of Bulk Quantities of Micro-
organisms
Safety of Biologically Active Products

Altered plants pollinating nearby plants. It
could change some characteristics of said
plants
Insulin
Human growth hormone
Tissue plasminogen activator
Interferon
Vaccines