Lecture 7 - industrial microbiology Flashcards
what is industrial microbiology?
◼ Industrial Microbiology uses microorganisms,
typically grown on a large scale, to produce
commercial products or to carry out important
chemical transformations.
◼ Enhancements of metabolic reactions that the
microorganisms are already capable carrying
out - overproduction
◼ Not microbial biotechnology where bugs are
genetically engineered to produce substances
they can’t normally produce.
◼ Often yeasts and molds and some bacteria,
often Streptomyces
what are pharmaceutical products?
❑ some of the most important industrial products
❑ include antibiotics, steroids
❑ all are secondary metabolites
what are Specialty chemicals and food additives
❑ vitamins e.g. B12 and riboflavin
❑ amino acids
◼ e.g. glutamic acid (monosodium glutamate)
◼ aspartic acid and phenylalanine (aspartame, for diet soft
drinks)
what are commodity chemicals?
❑ inexpensive chemicals produced in bulk
❑ include ethanol, acetic acid, lactic acid, glycerol, citric acid
❑ increasing use of ethanol as additive for petrol (gasohol)
what are examples of alcoholic beverages?
❑ beers, wines, spirits
how is vinegar produced?
◼ vinegar = vin aigre (sour wine)
oxidation oxidation
◼ ethanol ———> acetaldehyde ————> acetic acid
CH3CH2OH CH3CHO CH3COOH
◼ Acetic acid bacteria (Acetobacter,Gluconobacter)
◼ very acid tolerant
◼ strictly aerobic but do not fully oxidise acetic acid to
CO2
◼ process has high oxygen demand
what are the methods to produce vinegar?
◼ Open-vat (Orleans method)
traditional, surface culture, inefficient
◼ Trickle method continuous culture
◼ Bubble method stirred tank reactor
highly efficient, 90 - 98 % conversion of ethanol to acetic acid
what is the trickle method?
Vinegar generator
Alcohol trickled through
wood shavings,
aerated from below
Acetic acid bacteria form
biofilm on wood,
convert alcohol to acid
Operated continuously,
liquid recirculated until
desired acid
concentration is
achieved, 4-5 days
what is fermentation?
◼ Large scale process
◼ 5 to 500,000 litres
◼ Anaerobic – not complex
◼ Aerobic – very complex
batch vs continuous culture…
◼ Batch cultures are closed systems, nutrients are added at the start, growth will eventually decline due to diminishing nutrients or accumulation of toxic waste products
◼ Continuous cultures are open systems, nutrients are added and culture is removed
continuously, organisms grow under steady state conditions
Batch cultures are more common because
◼ The product may only be required in small amounts intermittently
◼ A high product concentration may be required for DSP
◼ Certain products are only made during the stationary phase of growth e.g. some antibiotics
◼ Strain instability may necessitate regular renewal of culture
◼ Continuous processes are technically more difficult
the advantages of continuous culture are
◼ Productivity for a given size of culture vessel is increased
◼ Growth rate can be maintained at optimum for product
formation but has limited uses e.g. ethanol production,effluent treatment.
what is the growth and product formation?
◼ Primary metabolites formed during the growth phase e.g. ethanol production by yeast
◼ Secondary metabolites formed during stationary phase e.g. penicillin production by Penicillium chrysogenum.
Growth medium greatly influences product formation
Often over-produced
Often produced by spore-formers
primary vs secondary metabolites…
◼ Primary metabolite forms during exponential growth
❑ e.g. Alcohol, a by-product of microbial growth so formed as the microbes grow
◼ Secondary metabolite is produced near the end of the growth phase
❑ More complex and important metabolites
❑ Not essential for growth
❑ Highly dependent on growth conditions
❑ Often closely related but highly distinct compounds produced by closely related microbes
❑ Highly overproduced unlike primary metabolites
❑ Often produced during spore formation
❑ Can get interactions between primary and secondary metabolite pathways
◼ Tetracycline – 72 enzymatic reactions
◼ Erythromycin – 25 enzymatic reactions
what is the problem of scaling uo of large scale cultivation
◼ Vessels used: fermenters or bioreactors
◼ Most industrial processes are aerobic
◼ Provision of oxygen and mixing are important factors
◼ Transition from lab scale to large scale may be difficult
what are the stages in scale up?
Stages in scale-up
◼ Lab flask - process shown to be of commercial interest
◼ Lab fermenter - 5-10 L. Check effects of alterations of
growth conditions to optimise process
◼ Pilot plant stage - 300-3000 L. Conditions for commercial
scale can be tried out. Introduce instrumentation and
computer control of system.
◼ Commercial fermenter 10,000-500,000 L
what is downstream processing?
processing the biomass or the product on a large scale e.g.
◼ Separation- filtration, centrifugation, disruption of cells
◼ Concentration- extraction, ultrafiltration, precipitation
◼ Purification- chromatography, crystallisation
◼ Drying
what are the properties of a useful microorganism?
◼ Large scale growth
◼ Spore or reproductive cell form
◼ Grow rapidly and produce the product rapidly
◼ Bulk quantities available for cheap
◼ Non-pathogenic
◼ Advantage – genetically amenable to
manipulation
what are products of industrial microbiology? and examples
◼ Natural products or reactions
◼ Gene manipulation to obtain new products, not naturally
produced e.g. insulin, interferons, factor VIII
◼ Microbial cells
❑ baker’s yeast, sold as compressed or active-dry yeast
❑ single-cell protein e.g. Quorn (mycoprotein)
❑ inoculants, for fermented dairy products e.g. yoghurt
◼ Enzymes
❑ extracellular enzymes produced in large amounts
❑ digest nutrients to which cell is impermeable
❑ proteases, amylases, lipases for “biological” washing
powder
❑ glucose isomerase for high fructose syrup (sweetener)
❑ heat-stable Taq polymerase, for PCR
◼ Pharmaceutical products
❑ some of the most important industrial products
❑ include antibiotics, steroids
❑ all are secondary metabolites
◼ Specialty chemicals and food additives
❑ vitamins e.g. B12 and riboflavin
❑ amino acids
◼ e.g. glutamic acid (monosodium glutamate)
◼ aspartic acid and phenylalanine (aspartame, for diet soft
drinks)
how do you extract and purify an antibiotic?
◼ Produce the antibiotic successfully in large-scale fermenters
◼ Downstream processing. Elaborate methods for extraction and purification are often necessary
what is brewing, distilling and commodity alcohol?
◼ Brewing is the term used to describe the
manufacture of alcoholic beverages from malted
grains
◼ Yeast is used to produce beer
◼ Two main types of brewery yeast strains
❑ Top fermenting — ales
❑ Bottom fermenting — lagers
◼ Distilled alcoholic beverages are made by heating
previously fermented liquid to a temperature that
volatilizes most of the alcohol
❑ Whiskey, rum, brandy, vodka, gin
◼ > 50,000,000,000 liters of ethanol are produced
yearly for industrial purposes
❑ Used as an industrial solvent and gasoline supplement
what are yeasts used for?
beer yeasts
ability to produce vigorous fermentation
flocculate
produce desired by-products (aroma and flavour)
starter culture – from previous batches
or pure strains
what is malting?
Barley - traditional raw material
high C (starch), low N
–> germination –>
Malt - contains various enzymes
amylases (starch ->maltose)
and proteases
essential for the brewing process
Dried malt - enzymes stable
what is mashing?
what is fermentation of ale and lager?
◼ Ale
❑ traditionally produced by batch fermentation in large, open vats
❑ using top-fermenting yeast
❑ strains of Saccharomyces cerevisiae
❑ wort is “pitched” with large inoculum
❑ lag phase, then rapid growth, may be aerated
❑ then anaerobic conditions allowed to develop to favour alcohol
production
❑ floating yeast skimmed off to inoculate further brews
❑ fermentation 5 - 6 days, 14 > 23oC
❑ pH falls from 5.2 > 4.1
◼ Lager
❑ produced by closed, batch fermentation
❑ using bottom-fermenting yeasts
❑ strains of Saccharomyces carlsbergensis (S. uvarum)
❑ fermentation 8 -14 days, 6 >12oC
what is maturation?
fresh beer –> filtration or centrifugation and surplus yeasts removed. then the beer is added to storage tanks at 4-8oC for a few days under natural conditioning
Lager – prolonged storage, several weeks at -1oC
Finishing
pasteurisation distribution to casks, bottles, kegs etc.
what is spoilage?
◼ By contaminants, after boiling of wort, from air, water,
pitching yeast
◼ Only certain bacteria and yeasts cause spoilage of beer
because
❑ low pH
❑ alcohol content
❑ hop extracts
❑ low storage temp
❑ anaerobic conditions
what are examples of spoilage organisms and methods of the prevention of spoilage
◼ Examples of spoilage organisms
❑ Lactobacillus acidity and cloudiness
❑ Acetobacter souring
❑ Wild yeast e.g. S. pasteurianus bitterness, cloudiness
❑ Pediococcus cerevisiae diacetyl, “ropy” beer
◼ Prevention of spoilage by
❑ aseptic and hygienic precautions
❑ regular cleaning and sterilisation of equipment
❑ use of pure strains
❑ pasteurisation
look at the screen shots to see the slides on mashing and the chemistry of industrial microbiology
