BIOTECHNOLOGY INDUSTRIAL AND APPLIED MICROBIOLOGY

workkinkajouΒιοτεχνολογία

5 Δεκ 2012 (πριν από 4 χρόνια και 11 μήνες)

173 εμφανίσεις

2007
-
2008

B
IOTECHNOLOGY


B. BIOLOGICAL FUELS


1

The need for biological fuels

2

Raw materials

These include wastes and crops;


wastes Dry Wastes


Wet wastes

Crops

In

the

future
,

crops

may

be

grown

specially

for

energy

production,

perhaps

on

land

unsuitable

for

growing

foodstuffs
.

Sugar

ca
ne

is

already

being

grown

in

Brazil

for

this

purpose
.



A. ETHANOL PRODUCTION

1
-

Substrates

include

sugar

cane
,

cassava

roots
,

cellulose

waste

and

corn
.


Cassava

roots

contain

starch

which

must

be

hydrolysed

to

sugars
,

and

cellulose

waste
,

such

as

timber

and

straw,

needs

quite

complex

pre
-
treatment

with

ligno
-
cellulase

enzymes

or

chemicals
.


2
-

At

present
,

alcohol

production

is

similar

to

the

traditional

process

but

much

research

is

taking

place
.


It

is

hoped

that

more

efficient,

genetically

engineered

M
.
O
.
s

will

be

developed

and

that

newer

fermentor

designs

and

immobilized

enzyme

technology

will

improve

efficiency
.


3
-

Distillation

costs

can

be

reduced

by

using

a

cheap

fuel
,

and

bagasse

(the

waste

from

sugar

cane)

has

proved

to

be

an

economical

fuel

for

raising

steam

for

the

process

by

combustion
.

4
-

A

range

of

M
.
O
.
s

have

been

used

in

the

production

of

ethanol
,

using

many

different

carbohydrates

as

substrate
.



Traditionally,

ethanol

production

has

relied

upon

the

use

of

yeasts
,

mostly

Saccharomyces

species
.


5
-

Zygomonas

mobilis

has

been

used

in

South

America

for

many

years

in

the

production

of

tequila
,

and

in

Indonesia

and

Africa

to

make

palm

wine
.



However,

its

use

in

the

western

world

is

quite

new
.

Recent

research

into

Zygomonas

has

shown

that

it

is

more

efficient

than

yeasts

in

converting

sugar

to

ethanol
.


6
-

A

technique

has

been

developed

to

produce

ethanol

using

Zygomonas

in

a

continuous

culture

process,

rather

than

the

more

traditional

batch

culture

methods
.



6.4

The production of methane


(
1
)

Sewage



(
2
)

Urban

waste
,

landfill

gas



(
3
)

Biogas

fermentors


However,

while

this

is

a

useful

small
-
scale

process
,

it

is

unlikely

to

be

commercially

avaible

on

a

large

scale

because
:

methane

can

be

produced

far

more

cheaply

from

coal

at

present
;


natural

gas

is

cheaper

than

microbially

produced

methane
.



There

are

many

natural

sources

of

methane



Gas

is

expensive

to

store,

transport

and

distribute

at

present
.



It

is

expensive

and

difficult

to

liquefy
.


(
4
)

Agricultural

wastes

Some

farms

now

place

animal

manure

and

other

crop

residues

into

anaerobic

digestion

tanks
.



Here,

the

waste

is

fermented

by

M
.
O
.
s

and

the

methane

produced

is

collected,

liquefied

and

used

to

power

farm

machinery
.



In

some

cases

it

may

be

used

to

fire

boilers
,

which

heat

glasshouses

and

produce

early

crops

of

tomatoes
,

peppers

and

other

vegetables
.

C. P
HARMACEUTICALS

PRODUCED

BY

M.O.
S
:


1. Dextrans

Dextrans

are

polysaccharides

produced

by

lactic

acid

bacteria
,

in

particular

members

of

the

genus

Leuconostoc

(e
.
g
.

L
.

dextranicus

and

mesenteroides
)

following

growth

on

sucrose
.



2
-

Vitamins, amino acids and organic acids


1. Vitamins


Vitamin

B
2

(
riboflavin
)

is

a

constituent

of

yeast

extract

and

incorporated

into

many

vitamin

preparations
.



Vitamin

B
2

deficiency

is

characterized

by

symptoms

which

include

an

inflamed

tongue
,

dermatitis

and

a

sensation

of

burning

in

the

feet
.



2. Amino acids

Amino

acids

find

applications

as

ingredients

of

infusion

solutions

for

parenteral

nutrition

and

individually

for

treatment

of

specific

conditions
.



They

are

obtained

either

by

fermentation

processes

similar

to

those

used

for

antibiotics

or

in

cell
-
free

extracts

employing

enzymes

isolated from bacteria.
3. Organic acids

Examples

of

organic

acids

(
citric
,

lactic
,

gluconic
)

produced

by

M
.
O
.
s
.

Citric

and

lactic

acids

also

have

widespread

uses

in

the

food

and

drink

and

plastics

industries,

respectively
.

Gluconic

acid

is

also

used

as

a

metal
-
chelating

agent

in,

for

example,

detergent

products
.


3 Iron
-
chelating agents

Growth

of

many

M
.
O
.
s

in

iron
-
deficient

growth

media

results

in

the

secretion

of

low

molecular

weight

iron
-
chelating

agents

called

siderophores
,

which

are

usually

phenolate

or

hydroxamate

compounds
.


-
The

therapeutic

potential

of

these

compounds

has

generated

considerable

interest

in

recent

years
.



4 Enzymes


1
-

Streptokinase and
streptodornase


Mammalian

blood

will

clot

spontaneously

if

allowed

to

stand
:

however,

on

further

standing,

this

clot

may

dissolve

as

a

result

of

the

action

of

a

proteolytic

enzyme

called

plasmin
.



Plasmin

is

normally

present

as

its

inactive

precursor
,

plasminogen
.


Streptokinase

is

administered

by

intravenous

or

intra
-
arterial

infusion

in

the

treatment

of

thrombo
-
embolic

disorders
.


2
-

L
-
Asparaginase

-
L
-

Asparaginase
,

an

enzyme

derived

from

E
.

coli

or

Erwinia

carotovora
,

has

been

employed

in

cancer

chemotherapy

where

its

selectivity

depends

upon

the

essential

requirement

of

some

tumors

for

the

amino

acid

L
-
asparagine

.



-

Normal

tissues

do

to

require

this

amino

acid

and

thus

the

enzyme

is

administered

with

the

intention

of

depleting

tumor

of

asparagine

by

converting

it

to

aspartic

acid

and

ammonia
.



3
-

Neuraminidase

-
Neuraminidase

derived

from

Vibrio

cholerae

has

been

used

experimentally

to

increase

the

immunogenicity

of

tumour

cells
.


-
It

is

capable

of

removing

N
-
acetylneuraminic

(
sialic
)

acid

residues

from

the

outer

surface

of

certain

tumor

cells,

thereby

exposing

new

antigens

which

may

be

tumor

specific

together

with

a

concomitant

increase

in

their

immunogenicity
.


-
-
In

lab

animals

administration

of

neuraminidase
-
treated

tumour

cells

was

found

to

be

effective

against

a

variety

of

mouse

leukaemias
.



4 β
-
Lactamases

-

β
-
Latamase

enzymes,

whilst

being

a

considerable

nuisance

because

of

their

ability

to

confer

bact
.

resistance

by

inactivating

penicillins

and

cephalosporins

are

useful

in

the

sterility

testing

of

certain

antibiotics

and,

prior

to

culture,

in

inactivating

various

β
-
lactams

in

blood

or

urine

samples

in

patients

undergo

therapy

with

these

drugs
.

-

One

other

important

therapeutic

application

is

the

rescue

of

patients

presenting

symptoms

of

a

sev
e
re

allergic

reaction

following

administration

of

a

β
-
lactamase

-

sensitive

penicillin
.



3
-

A
PPLICATIONS

OF

M.O.
S

IN

THE

PARTIAL

SYNTHESIS

OF

PHARMACEUTICALS
:


3.1 Production of antibiotics

Alexander

Fleming's

accidental

discovery

of

penicillin

in

1929

is

well

known
.



He

found

the

mould

Penicillium

notatum

contaminating

a

Petri

dish

of

pathogenic

bacteria

and

inhibiting

their

growth
.



He

isolated

penicillin

but

it

was

not

until

the

Second

World

War

that

it

was

successfully

produced

on

a

large

scale
.


At

first,

it

was

grown

in

static

liquid

culture

in

flasks,

shallow

pans

and

bottles,

but

this

process

was

inefficient

and

it

was

not

possible

to

produce

enough

penicillin

to

meet

demand
.



Two theories have been proposed to
explain antibiotic production.

1
-

Antibiotics

are

secondary

metabolites
,

so

they

may

be

produced

to

keep

enzyme

systems

operative

when

the

microbe

has

run

out

of

nutrients

and

cell

division

is

no

longer

possible
.


Normally,

when

the

substrate

has

been

used

up
,

the

enzymes

of

that

particular

pathway

would

be

broken

down
.


-
Then,

if

a

new

nutrient

supply

was

found
,

there

would

be

a

delay

while

the

necessary

enzymes

were

produced
.

-
It

has

been

suggested

that

making

a

secondary

metabolite

keeps

the

enzymes

active
,

so

that

the

microbe

can

quickly

take

advantage

of

any

new

food

supply
.

-
2
-

Some

scientists

think

antibiotic

production

is

for

ridding

of

the

cell

toxic

metabolic

waste
.

-
-

Although

not

toxic

to

the

organism

producing

them
,

these

substances

could

still

be

highly

toxic

to

other

M
.
O
.
s
.


-
If

the

toxin

phenylacetic

acid

is

added

to

a

culture

of

Penicillium
,

penicillin

production

is

increased
.

This

observation

supports

this

theory
.


-

It

is

of

course,

possible

that

both

theories

are

correct

since

they

are

not

contradictory
.


T
HE

INDUSTRIAL

PRODUCTION

OF

ANTIBIOTICS
;


PENICILLIN PRODUCTION

1
-

M
.
O
.

the

organism

used

for

production

of

penicillin

was

Penicillium

notatum
,

but

the

mostly

common

used

is

P
.

chrysogenus

.


2
-

Inoculum

Preparation
;

a

pure

inoculum

in

sufficient

volume

and

in

the

fast

growing

(
logarithmic
)

phase

so

that

a

high

population

density

is

soon

obt
a
ined
.

3
-

The

fermenter
;

A

typical

fermenter

is

closed
,

vertical
,

cylinderical
,

stainless

steel

vessel

with

convexly

dished

ends

and

25

-

250

m
3

capacity
.



The

height

is

usually

two

to

three

times

its

diameter
.



4
-

Oxygen

supply
;

Penicillin

fermentation

need

oxygen
,

which

is

supplied

as

filtered

sterilised

air

from

a

compressor
.


5
-

Temperature

control
;

The

production

of

penicillin

G

is

very

sensitive

to

temperature
,

the

tolerance

being

less

than

1

C
.



Heat

is

generated

both

by

the

metabolism

of

nutrients

and

by

the

power

dissipated

in

stirring
,

and

has

to

by

removed

by

controlled

cooling
.


6
-

Defoaming

agents
;

The

fermenter

system

stirred

vigorously

and

aerated

usually

foam
,

so

provision

has

to

made

for

adding

defoaming

agents
.

7
-

Instrumentation
;

The

vessel

is

fitted

with

several

probes

to

detect

foaming
,

temperature
,

pH
,

O
2
-
tension

and

exhaust

gas
.


8
-

Media
;

Not

all

the

nutrients

required

during

fermentation

are

initially

provided

in

the

culture

medium
.



Provision

is

therefore

made

to

add

these

while

the

fermentation

is

in

progress
.

The

media

used

is

corn

steep

liquor

(
CSL
)
.


9
-

Transfer

and

sampling

systems
;

Appropriate

pipework

is

provided

to

transfer

the

inoculum

to

the

vessel
,

to

allow

taken

routine

sample

and

to

transfer

the

final

content

to

the

extraction

plant
.

10
-

The

optimum

temperature

and

pH

for

growth

are

not

those

for

penicillin

production

they

must

be

changed

du
r
ing

the

process
.


11
-

The

production

phase

begin

with

the

addition

of

phenylacetic

acid

(
PAA
)
.

12
-

PAA

supplies

the

side

chain

of

penicillin

G
.

13
-

PAA

is

toxic

for

the

M
.
O

so

it

must

be

supplied

in

small

quantities

without

approaching

the

toxic

level
.

14
-

Termination
;

The

harvest

is

carried

out

shortly

after

the

first

signs

of

faltering

in

the

efficiency

of

conversion

of

the

most

costly

raw

material

to

penicillin
.


15
-

Extraction:

A
-

Removal

of

the

cell
;

penicillin

G

is

extracellular

the

first

step

is

to

remove

the

cells

by

filtration
.


B
-

Isolation

of

penicillin

G
;

Penicillin

G

is

very

unstable
,

so

it

must

be

quickly

extracted

by

organic

solvent

(
amyl

acetate
)

from

the

acidified

aqueous

solution
.

C
-

Treatment

of

cr
u
de

extract
;

first

formation

of

an

appropriate

salt
,

charcoal

treatment

to

remove

pyrogens

and

sterilization

by

using

dry

heat
.


Interferons

are

antiviral

chemicals
,

which

also

have

some

tumour

inhibiting

properties
.


These

used

to

be

extracted

from

human

fibroblast

cells,

but

yields

were

minute
.


Recombinant

DNA

methods

have

now

been

used

to

synthesize

interferons

using

a

suitable

bacterium
,

such

as

Escherichia

coli
.

Some

other

anti
-
tumour

pharmaceuticals

are

also

made

microbiologically
.


An

example

is

bleomycin
,

a

glycopeptide
,

made

by

Streptomyces

verticillus
.

This

drug

has

the

ability

to

disrupt

the

DNA

and

RNA

of

tumour

cells
.



Steroid biotransformation

Since

steroid

hormones

can

only

be

obtained

in

small

quantities

dir
e
ctly

from

mammals
,

attempts

were

made

to

synthesize

them

from

plant

sterols

which

can

be

obtained

cheaply

and

economically

in

large

quantities
.



However,

all

adrenocortical

steroids

are

characterized

by

the

presence

of

an

oxygen

at

position

11

in

the

steroid

nucleus
.


-
More

recent

advances

involving

the

employment

of

M
.
O
.
s

in

biotransformation

reactions

utilize

immobilized

cells

(both

living

and

dead
)
.

-

Immobilization

of

microbial

cells
,

usually

by

entrapment

in

a

polymer

gel

matrix
,

has

several

important

advantages
.



Chiral

inversion

Several

clinically

used

drugs
,

e
.
g
.

salbutamol

(a

β
-
adrenoceptor

agonist
),

propranolol

(
a

β
-
adrenoceptor

antiagonist
)

and

the

2
-
arylpropionic

acids

(
NSAIDs
)

are

employed

in

the

racemic

form
.



-

It

has

thus

been

suggested

that

the

enantiomerically

pure

S(+)

form

could

be

administered

clinically

to

give

a

reduced

dosage

and

possible

less

toxicity
.


4
-

USE

OF

M
.
O
.
S

AND

THEIR

PRODUCTS

IN

ASSAYS



Microbiological assays

In

microbiological

assays

the

response

of

a

growing

population

of

M
.
O
.
s

to

the

antimicrobial

agent

is

measured
.



The

usual

methods

involve

agar

diffusion

assays,

in

which

the

drug

diffuses

into

agar

seeded

with

a

susceptible

microbial

population

and

produces

a

zone

of

growth

inhibition
.


In

the

commonest

form

of

microbiological

assay

used

today,

samples

to

be

assayed

are

applied

in

some

form

of

reservoir

(porcelain

cup,

paper

disc

or

well)

to

a

thin

lay

of

agar

seeded

with

indicator

organism
.



The

drug

diffuses

into

the

medium

and

after

incubation

a

zone

of

growth

inhibition

forms,

in

this

case

as

a

circle

around

the

reservoir
.



Vitamin and amino acid bioassays

-
The

principle

of

microb
.

bioassays

for

growth

factors

such

as

vitamins

and

amino

acids

is

quite

simple
.



-
Unlike

antibiotic

assays

which

are

based

on

studies

of

growth

inhibition,

these

assays

are

based

on

growth

exhibition
.



-

All

that

is

required

is

a

culture

medium

which

is

nutritionally

adequate

for

the

test

M
.
O
.

in

all

essential

growth

factors

except

the

one

being

assayed
.

-
If

a

range

of

limiting

concentrations

of

the

test

substance

is

added,

the

growth

of

the

test

M
.
O
.

will

be

proportional

to

the

amount

added
.



Carcinogen and mutagen testing

-
A

carcinogen

is

a

substance

which

causes

living

tissues

to

become

carcinomatous

(to

produce

a

malignant

epithelial

tumor)
.




-
A

mutagen

is

a

chemical

(or

physical)

agent

which

induces

mutation

in

a

human

(or

other)

cell
.

The Ames test



The

Ames

test

is

used

to

screen

a

wide

variety

of

chemicals

for

potential

carcinogenicity

or

as

potential

cancer

chemotherapeutic

agents
.



-
The

test

enables

a

large

No
.

of

compounds

to

be

screened

rapidly

by

examining

their

ability

to

induce

mutagenesis

in

specially

constructed

bacterial

mutants

derived

from

Salmonella

typhimurium
.



Use of microbial enzymes in sterility testing

-

Sterile

pharmaceutical

preparations

must

be

tested

for

the

presence

of

fungal

and

bacterial

contamination

before

use
.


-
If

the

preparation

contains

an

antibiotic,

it

must

be

removed

or

inactivated

where

membrane

filtration

is

the

usual

recommended

method
.


-

However,

this

technique

has

certain

disadvantages
.

Accidental

contamination

is

a

problem,

as

is

the

retention

of

the

antibiotic

on

the

filter

and

its

subsequent

liberation

into

the

nutrient

medium
.



6 Insecticides

-

Like

animals,

insects

are

susceptible

to

infections

which

may

be

caused

by

viruses,

fungi

bacteria

or

protozoa
.


-

The

use

of

M
.
O
.
s

to

spread

diseases

to

particular

insect

pests

offers

an

attractive

method

of

bio
-
control,

particularly

in

view

of

the

ever
-
increasing

incidence

of

resistance

to

chemical

insecticides
.


-

However,

any

M
.
O
.

used

in

this

way

must

be

highly

virulent,

specific

for

the

target

pest

but

non
-
pathogenic

to

animals,

man

or

plants
.


-

It

must

be

economical

to

produce,

stable

on

storage

and

preferably

rapidly

acting
.

Bacterial

and

viral

pathogens

have

so

far

shown

the

most

promise
.



MICROBIAL DEGRADATION

-

Biodegradation

and

biodeterioration

The

use

of

M
.
O
.
s

to

break

down

substances

is

usually

called

biodegradation
.



However,

M
.
O
.
s

often

break

down

substances

in

a

way

that

is

not

beneficial

to

humans,

for

example

in

causing

food

spoilage
.


This

activity

is

generally

called

biodeterioration
.

Sewage

Sewage

is

composed

of

the

following
:
-

a
-

Human

waste

made

up

of

human

excreta

mixed

with

waste

household

water
.


This

contains

many

M
.
O
.
s

including

potential

pathogens
.


A

major

pollutant

from

waste

household

water

is

detergent,

which

causes

persistent

foam

and

has

high

levels

of

phosphates
.


b
-

Industrial

wastes

which

are

variable

in

nature,

depending

on

the

industry
.


Some

can

be

very

toxic

to

M
.
O
.
s

and

must

undergo

pretreatment

so

that

they

do

not

kill

or

inhibit

the

M
.
O
.
s

which

degrade

the

sewage
.


Many

industries

are

required

to

treat

their

own

sewage,

either

wholly

or

partially
.

c
-

Road

drainage

consists

of

rain

water

together

with

grit

and

other

debris

which

enters

the

sewers

from

roadside

gutters
.


Sewage treatment

Sewage

is

treated

in

two

or

three

stages

as

follows
.


Primary treatment.


Materials

which

will

settle

out

are

removed
.

The

sedimented

solids

pass

on

to

a

digester

for

further

treatment,

while

the

liquid

(effluent)

continues

into

the

secondary

treatment

stage

.

Secondary treatment.

Aerobic

M
.
O
.
s

are

used

to

break

down

most

of

the

organic

matter

in

the

effluent
.

Any

sludge

produce

in

this

process

is

passed

on

to

anaerobic

digesters
.


Tertiary treatment

This

involves

chemical

and

biological

treatment

which

renders

the

sewage

effluent

fit

for

drinking
.

However,

this

is

a

very

expensive

treatment,

so

it

is

only

carried

out

when

absolutely

necessary
.

There

are

two

main

reasons

for

treating

sewage
.



Firstly,

sewage

can

contain

pathogens

which

cause

diseases,

such

as

Salmonella

typhi

(typhoid),

pathogenic

Escherichia

coli

(gastroenteritis)

and

Ascaris

lumbricoides

(roundworm)
.



Secondly,

by

treating

sewage,

pollution

of

the

environment

can

be

avoided
.


Microbial Mining

-

Some

bacteria

are

useful

in

extracting

metals

from

low
-
grade

ores
.


-

This

is

because

they

are

chemoautotrophic

which

means

they

derive

their

energy

from

inorganic

chemicals
.


-

Bacteria

of

the

genus

Thiobacillus

are

used

commercially

to

extract

copper

and

uranium

from

otherwise

uneconomic

reserves
.


-
Cobalt
,

lead

and

nickel

may

also

be

extracted

in

this

way

in

the

near

future
.



The

extraction

process

may

require

extremes

of

environmental

conditions,

such

as

heat

and

pH
.



Genetic

engineering

techniques

are

being

used

to

confer

acid
-

and

heat

resistance

on

these

M
.
O
.
s
.


Problems of biologically active
biotechnology products:

Vaccines

and

antibiotics

are

obvious

examples

of

biologically

active

products,

and

care

must

be

taken

to

prevent

their

indiscriminate

dispersal
.



Contaminants

in

otherwise

safe

processes

may

produce

toxic

molecules

that

could

become

incorporated

into

final

products,

leading

to

food

poisoning
.

Allergenic

reactions

to

produce

formulations

must

also

be

guarded

against
.



Overuse

of

antibiotics

in

agriculture

could

lead

to

carry
-
over

into

human

foods,

resulting

in

possible

development

of

antibiotic

resistance

in

human

disease

organisms
.


Many

countries

now

restrict

the

use

of

antibiotics

in

agriculture
.