THE MEDIA IN

FERMENTATION
Nutritional Requirements

Introduction

Microorganisms obtain energy for support of biosynthesis

and growth from their environment in a variety of ways
Need detailed investigation to establish the most suitable
medium for an industrial fermentation process, but certain
basic requirements must be met by any such medium.
nutritional requirements are complex and varied
Not only are the types of microorganisms diverse
(bacteria, molds, yeast), but the species and strains
become very specific; the products
All microorganism requires water, source of energy, C, N,
Mineral, vitamins, oxygen (aerobic).
On a small scale its easy to maintain but on large scale
very difficult to maintain the satisfactory growth.

Medium criteria

Produce max yield of prod. or biomass per g

substrate used
Produce max concentration of prod.
permit max rate of prod formation
Permit min yield of undesired prod
Consistent quality & available throughout year
Cause min problems during preparation
Cause min problems during production & down
stream process

Consideration

availability of nutrients required by

microorganisms: energy, grow, production
fermentation conditions and condition change
during fermentation (e.g. substrate pH, MC,
incubation temperature)
stage of growth of micro-organisms product
presence of other competing micro-organisms
Type of reactor
Preparation & downstream process (e.g.
separation, recovery, effluent treatment)
Problems in upscale (e.g. agitation, viscosity, gas
transfer)

Natural or formulated-defined media?

Natural Media:

Cheaper
High variation/batch
Unpredictable
product/biomass
More complex in
down stream process
Difficult to detect
small yield
improvement

Formulated-MEDIA

Easily reproduce
Specific to the strain
More expensive
More predictable
result
More simple process
cheaper
Easier to detect
process improvement

Natural media

MEDIA FORMULATION

Essential stage in design of successful laboratory experiments, pilot-scale

development & manufacturing processes

The constituents of a medium must satisfy:

the elemental requirements for cell biomass and metabolite production

must be an adequate supply of energy for biosynthesis and cell maintenance

Some components need for product formation but not for biomass
formation
Stoichiometry for growth & product formation:
C(energy sources)+N
source+othersbiomass+Product+CO2+H2O+heat

This equation should be expressed in quantitative terms

It should be possible to calculate:
~ the minimal quantities of nutrients which will be needed to produce biomass
~ the substrate concentration in order to produce required product yield

Commercially, trade secret

Microbial Nutrient

Nutrient: combination of elements require for

energy, biosynthesis of cellular matter, products
in cell operation, maintenance and reproduction
predominant atomic constituents of organisms, C,
H, N, O, P, and S, 40-50% C, 30-50% O2, 6-8%
H2, 3-12 N
All living cells contain water as their predominant
constituent
Cell: lipids, polysaccharides, nucleic acids,
proteins and few common salts
Nutrients divided into 3 group (often
overlapping): energy supplier, C supplier and any
& everything else supplier

Water

Major comp. require in fermentation use

for nutrient solvent and solvent within the
cell
it has some unusual properties ionizes into
acid and base, and has a propensity for
hydrogen bonding
May need treatments
Consider: pH, salts dissolved, mineral,
effluent contaminant
Base on water availability: Solid state
fermentation (SSF) & Submerged liquid
fermentation (SmF)

SSF

Substrate requires
pretreatment (e.g SR,
Demerit sieving)

Merit

Low water availability

reduce contamination by
bacteria & yeast

Higher level of aeration

porous

Similar with natural mold

environment

Higher yiled/product
low nutrient dilution
Lower in production cost
low energy for agitation
& aeration

Poor grow of high

moisture microbial

Difficult in biomass
determination (mixed
up)

Difficult in control
process (e.g pH, O2,
MC)

Static condition, no
agitaion

Difficult in scale up
(e.g aeration)

SSF-microbial

Filamentous fungi are

the best
Good tolerance in low
water activity
the hyphal features
give the power to
penetrate into solid
substrate
Penetration increase
accessibility of all
nutrient within
substrate

Cell nutrients

Nutrients required by cell: macronutrient &

micronutrient
Macronutrients needed > 10-4 M
make up about 95% of dw

e.g C, N, O, P, S
Micronutrient needed < 10-4 M
involve in enzyme function, maintain in
protein structure
e.g Zn, Cu, Mn, Ca, Na

Carbon

Biomass 50% C (db); skeleton of all cellular molecules

for building cell mass and forming product, as well as

acting as an energy source

Energy sources; secreted as CO2 & organic compounds

Single comp for energy & C sources; add nutrients
Energy used for metabolism-biosynthetic reaction & cell
maintaining

The main product of fermentation process will often

determine the choice of C source, particularly if the
product results from the direct dissimilation

The impurity of C source should be considered

sterilization may affect suitability of C source (e.g.

sugar separated from amino acid black N comp.
which may inhibit microbial grown

C sources
1- Carbohydrates: C, O, H & energy 0.2-25% of media
3 classes: mono-, di-, & polysacharides
Breakdown of poly- & di- to simpler sugars with enzyme
help major source of energy-rich comp
Lab media glucose, sucrose & fructose
Industrial media starch (glucose, dextrin), mollases
(sucrose), whey (lactose)
2 - Oils & Fats
Vegetable oil contains fatty acid, oleic, linoleic & linolenic
acid
Typical oil contain~2.4x energy of glucose/weight
Less volume space smaller fermenter (10 kcal 1,24 l
of oil; 5 l of glucose
act as antifoam too
3 - Hydrocaarbon & their derivatives
N-alkanes for organic acid, amino acid, vitamin etc
Provide energy ~ 2x C, 3x sugar

Nitrogen

10-14% of cell dw
N serve to build structural proteins, functional enzymes
& nucleic acids
N presents in cell as amino groups
Supplied as organic or inorganic sources
Organic acid faster growth & fulfill microorganism
requirement (e.g. Protein, peptide, amino acids etc)
N inorganic supplied as ammonia salts (NH4Cl,
NH4NO3), amonia gas, nitrate
In industries: soya meal, yeast extract, corn steep
liquor

Oxygen

key nutrient for aerobic microbes

20% of cell dw

microorganisms that are dependent on respiration for

generating energy require molecular oxygen as the
final hydrogen or electron

commonly found as a constituent of cellular water and

organic compounds (e.g. carbohydrate)

low solubility of oxygen in water

Oxygen transfer achieved by shaking, sparging,

aeration and agitation.

Trace elements

In mM level essential comp, added as distinct

comp in many media : P, S, K and Mg
In M level sometimes supplied from
quantities occurring in water or impurities of
other comp: Fe, Na, Ca, Cu, Co, Zn, Mn etc.
Supplied during cell cultivation
Various functions: serve in coenzyme, catalyse
reactions, vitamin synthesis, cell transport
Primary metabolite production is usually not very
sensitive to trace element concentration, but not
for secondary metabolite production

metallic elements can be supplied as nutrients in

the form of the cations of inorganic salts
K, Mg, Ca and Fe are normally required in
relatively large amounts and should normally
always be included as salts in culture media
Not possible to generalize ionic requirements
Oxygen is always provided in water
Aerobic organisms require molecular oxygen as
terminal oxidizing agents to fulfill their energetic
needs through aerobic respiration.
For obligate anaerobes oxygen is a toxic
substance

Growth Factors

organic comp, require in very low concentration,

perform specific catalytic/structural roles
vitamin, sterol, amino acids, fatty acids etc
Specific for certain microorganism
Required to stimulate microbial growth
Many natural C & N contain some growth factor; not
vital
Vitamins are growth factors which fulfill specific
catalytic needs in biosynthesis
Vitamin function as (part of ) coenzymes to catalyze
many reactions
The vitamins most frequently required are thiamin and
biotin.
Required in the greatest amounts are usually niacin,
pantothenate, riboflavin, and some folic derivatives,
biotin, vitamin B, and lipoic acid are required in