Lecture 1 introduction

Industrial MicrobiologyMBIO 4510

Lecture 1 – Introduction to Industrial Microbiology

Industrial microbiology is the commercial exploitation of microorganisms to produce valuable economic, environmental and socially important products, or to carry out important chemical transformations.

What is Industrial Microbiology?


Madigan, M.T. 2003. Brock Biology of Microorganisms 10th ed. New Jersey: Prentice Hall. P 967

Lecture 1 – Introduction to Industrial Microbiology Fermentation Products

Food, beverage, food additives and supplements

Dairy products (yogurt, cheese)

Alcoholic beverages (beer, wine)

Amino acids, vitamins


Health Care Products

Antibiotics – over 4000 isolated, only 50 used regularly

• β-lactams, penicillins, and cephalosporins

• Aminoglycosides (streptomysin)• Tetracyclins

Important to develop new antibiotics due to abuse/misuse of current antibiotics


Health Care Products

Alkaloids, steroids, vaccines

Therapeutic recombinant human proteins (insulin, interferons, blood-clotting factors, human growth hormone)

More recombinant therapeutic products to be developed


Production of microbial enzymes Proteases, carbohydrases, Taq

polymerase

Industrial chemicals and fuel Methane, ethanol, H2, propane, etc.

Environmental roles of microorganisms

Waste water treatment, desulphurization of fuels, leaching of metals, use of microbes to reduce usage of synthetic pesticides

Lecture 1 – Introduction to Industrial MicrobiologyOverview of a Fermentation Process

Waites et al. 2001. Industrial Microbiology: An Introduction. Oxford: Blackwell Science. P 2

Lecture 1 – Introduction to Industrial MicrobiologyFermentation process – Upstream

Processing1. Fermentation Organism

need suitable cells to produce desired products (bacteria, fungi, yeast, animal cells)

improve strain to enhance productivity and yield

maintain purity of cultures

Lecture 1 – Introduction to Industrial MicrobiologyFermentation process – Upstream

Processing



produce usable products or effects be available in pure culture be genetically stable, or genetically

mutated produce spores or other reproductive

structures to allow easy inoculation grow rapidly and produce product

quickly in large scale culture* be easily separated from products not be harmful to humans, plants,

animals, etc

To be useful for commercial processes, cells must:

Lecture 1 – Introduction to Industrial Microbiology Fermentation process – Upstream

Processing

2. Fermentation Medium

need cost-effective carbon and energy sources, essential nutrients

media often wastes from other processes, such as sugar processing wastes, lignocellulosic wastes, cheese whey and corn steep liquor


Processing

3. Fermentation

industrial microorganisms cultivated under controlled conditions to optimize growth of organism and production of microbial products

must avoid environmental conditions that trigger regulatory mechanisms (repression, feedback inhibition)


Processing


Fig. 9.2

Fig. 9.3

Fig. 9.4

Genentech 12,000 L animal cell bioreactor

Lecture 1 – Introduction to Industrial Microbiology Fermentation process – Downstream

Processing includes all processes after fermentation

involve cell harvesting, cell disruption, product purification from cell extracts or the growth medium

must be rapid and efficient to purify product and to maintain stability of product

safe and inexpensive to dispose of wastes


Primary Metabolites: produced during active growth

(trophophase) amino acids, organic acids, alcohol

fermentation products, vitamins

Secondary Metabolites: produced during stationary phase after

microbial biomass production has peaked (idiophase)

generally not essential for growth or reproduction

antibiotics, citric acid

Lecture 1 – Introduction to Industrial Microbiology Fermentation process





Economics of fermentation determined by cost of raw materials, utilities, labour and maintenance, fixed charges, working capital charges, etc.

P roduc ts

H igh vo lume, low value produc ts

L ow volume, high va lue produc ts

Scale up? $$$$

Mammalian

Prokaryotic

Yeast

Undisclosed

60 million of patients

Clinical Trials 500

0

5

10

15

20

25

30

2001 2004

US

Bil

lion

50

39

21

12

Therapeutic Monoclonal Antibodies

Datamonitor report “Mabs are hottest segment of biotech industry” articles in “Fierce Biotech” and “Bioprocess International”

Mabs generate revenue of $20 billion

14% annual growth expected 2006-2012 and outstrips other sectors of pharmaceutical industry

(Avastin, Herceptin, Remicade, Rituxan, Humira, and Erbitux) are 6 blockbusters.

Butler, M. (2005) Applied Microbiology and Biotechnology 68: 283-291.

The demand for mammalian cell culture products

1986 1988 1990 1992 1994 1996 1998 2000 2002 2004 2006 2008

Cu

mu

lati

ve p

rod

uct

ap

pro

vals

0

10

20

30

40

50

60

Dem

and

(kg

)

0

500

1000

1500

2000

2500

3000

number of productsKg capacity demand

enbrel

infergen

basiliximab

epo

herceptin

rituximab

abciximab

humulin

Annual demand (kg)

1e-3 1e-2 1e-1 1e+0 1e+1 1e+2 1e+3 1e+4 1e+5 1e+6 1e+7 1e+8 1e+9

Pric

e ($

/g)

1e-4

1e-3

1e-2

1e-1

1e+0

1e+1

1e+2

1e+3

1e+4

1e+5

1e+6

1e+7

1e+8

1e+9

Plasma HSA

penicillin

lysineethanol

Pharmaceutical Prices

Lecture 9 Animal Cell BiotechnologyScaling up the production process

Butler, M. 2004. Animal cell culture and technology 2nd ed. London and New York:Garland Science/BIOS Scientific Publishers. P203.

Pre-purification vs selling price of biological products

Selling price ($ per kg)

1e-2 1e-1 1e+0 1e+1 1e+2 1e+3 1e+4 1e+5 1e+6 1e+7 1e+8 1e+9 1e+10Con

cent

ratio

n in

sta

rting

med

ium

(g/l)

1e-7

1e-6

1e-5

1e-4

1e-3

1e-2

1e-1

1e+0

1e+1

1e+2

1e+3ethanol

citric acidamino acids

penicillin

bulk enzymes

insulin

m.antibodies

factor VIII

therapeutic enzymes

Wurm,F (2004) Nature Biotech 22: 1393

Milestones in the development of animal cell technology

1880Roux maintained embryonic chick cells in saline solution

18901900

Harrison grew frog nerve cells by the 'hanging drop' technique.

1910Carrel used aseptic techniques for long term cell cultures.Rous and Jones used trypsin for sub-culture of adherent

cells.1920

The 'Carrel' flask was designed for cell culture.19301940

Antibiotics were added to culture medium.Earle isolated mouse L fibroblasts.Enders grew polio virus on cultured human cells.

1950Gey cultured HeLa cells.Eagle developed a chemically defined culture medium.

1960Hayflick and Moorhead showed that human cells have a

finite lifespan.Ham grew cells in a serum-free medium.Harris and Watkins fused human and mice cells.

1970Kohler and Milstein produced an antibody-secreting

hybridoma.Sato developed serum-free media from hormones and

growth factors.1980

Human insulin was produced from bacteria.Monoclonal antibody (OKT3) used for human therapy.Recombinant tPA licensed for human therapy.

1990Humanized chimeric antibodies used for human therapyStem cells isolated

Technology

Lecture 1 introduction