Stoichiometry and Energetics

Material balance on biochemical

reactions The universal principle that materials cannot be
created or destroyed (unless there is a nuclear
reaction) holds in biochemical systems. In any
closed system the total mass of every element,
C, N, O, H, P, etc, is constant over time. As a first
step in analyzing a biochemical reaction system,
the system and its boundary and the surroundings
must be defined. Often a physical entity, such
as a cell or a bioreactor, is defined as a system,
and material and energy balance is performed
on the system. In other cases the balance is
done on a set of reactions that Is a subset of the
reaction network in the cell. Material balance can
be performed on either a compound (a chemical
species) or an element. For a chemical species ί,
the material balance is the rate of accumulation of
i = rate of inputs of i – rate of outputs of i + net rate
of production consumption of i from all reaction. For
a given element, such as C, N, O, it can shift from
one compound to another due to chemical reaction,
but the total amount is conserved. Thus elemental
balance can be written as a rate of accumulation
of elements = rate of input of i-rate of output of a.

Stochiometry 1
Stoichiometry and Energetics
Typically, chemical reactions are written as a
single or a system of stoichiometric equations.

∑a C ai H bi N ci Odi = 0 In the equation the

i
stoichiometric coefficient (α) has opposite sign
for reactants and products. By convention
the products are given positive stoichiometric
coefficients, while reactants negative.
The heat of reaction (ΔH ro ); is positive
for endothermic reaction and negative for
exothermic ones. When multiple reactions occur
in a system, an overall stoichiometric equation
can be written that is a linear combination of the
stochiometric equations of component reactions.

When dealing with biochemical systems

stoichiometric equations are used for describing
reactions in biochmecial pathways, as well
as for depicting complex systems such as the
conversion of nutrients into cells or organisms.
The formulation of traditional chemical reactions
and biochemical reactions are virtually identical.
When using stochiometric equations for biomass
transaction we will only consider C, H, O, N, and
P in most cases, as other elements participate
only in a small fraction of all biochemical reactions
and contribute only a very small quantity to the
biomass. Stochiometric equations can also be
written for reactions that occur inside a physical
system, such as a cell or all reaction (whether they
are enzyme catalysed reactions or cells grown on
nutrients and producing product) inside a reactor.
In the case of dealing with a physical system,
the matierals can flow into te system (inputs0
and out of the system(outputs). The material
balance will involve the flow of materials (inputs
and outputs) and the reactions in the system.

Most biological reactions (almost all) occur in

aqueous solution. A large number of biological
moelucles, including amino acids, proteins,
organic acids, phosphorylated sugars have
functional groups whose charge is dependent
upon the chcemical environment. In presenting
treactions involving those molecules, the change
balance is not explicityly written. Because

2 Stochiometry
 the reactions occur in an environment that
water molecules are the predominant species,
sometimes even water molecules is not explicityly
written. In performing measurement for matieral
blance, balancing on water is virtually impossible
unless isotope labeling is used. Many require
cosubstrates or cofactors, which are largely
“recycled” or “regenerated” by another reaction
or reactions. Those reactions are thus “coupled”.

Diversity in Stochiometry
Since the biological reactions are catalyzed by
enzymes in the organism, the starting point in
setting up the stoichiometric equation is to consult
the biochemical pathways. In addition to various
textbook, a number of websites also provide
valuable data; some are more comprehensive and
covers important pathways of general interest,
including the KEGG (http://www.genome.ad.jp/
kegg), Brenda, and ExPASy (http://www.expasy.ch .

In the course of evolution, most key metabolic

pathways involved in energy metabolism,
biosynthesis of building blocks, (amino acids, fatty
acids, nucleic acids) are very well conserved,
meaning that the basic reaction mechanisms
and the enzymes are very similar. The pathways
are used from the most primitive organisms to
mammals. For example, for organisms utilizing
glucose to derive energy aerobically, they all
convert each mole of glucose to six moles of CO2
and six moles of H2O, using basically the same
set of reactions. However, amongst this seemingly
very similar reaction network, there are also many
important differences between different species.

Consider glucose metabolism, both yeast

Sacchryomyces cerevisiae and bacterium
Xymomonas mobilis can convert glucose to ethanol,
but they do have some subtle differences in their
conversion pathway. In Sacchanomyces the typical
glycolysis pathway (Embden-Meyerhoff-Parnas
(EMP) pathway) is used, whereas in Xymomonas
mobilis, the Entner Doudoroff pathway is used.

Stochiometry 3
 C6H12O6 → 2C2H5OH +
2CO2 ∆Go = -56.5 kcal/mole

ADP + Pi → ATP +
H2O ∆Go = 7.3 kcal/mole

In Sacharomyces cerevisiae the net reaction is:

C6H12 O6 + 2 ADP + 2Pi →
2 C2H5OH + 2CO2 + 2ATP + 2H2O

∆Go = 41.9
-
kcal/mole

Macroscopically one mole of glucose is converted

to 2 moles each of ethanol and CO2. ATP is used in
intracellular chemical work and recycled back toADP.
Whereas in Xymomonas mobilis, the net reaction is:
C6H12O6 +ADP + Pi → 2 C2H5OH + 2CO2 +ATP + 2H2O

∆Go = 49.2 kcal/mole

It is clear that the pathways that the two

microorganisms use have different energetic
efficiencies even though they both produce
two moles each of ethanol and carbon dioxide
from one mole of glucose. Different pathways
with different efficiencies are not only seen in
different microorganism, but may also be used
under different environmental or physiological
conditions in the same organism. For example,
there are two pathways for ammonia assimilation
into organic nitrogen in E. coli. When the ammonia
concentration in the environment is high, E. coli
assimilates ammonia by reductive amination of
α – ketoglutarate. The reaction is catalyzed by
NADP-dependent L – glutamate dehydrogenase

COOH COOH
C =0 H 2 N ⋅ CH
NH 3 + CH 2 + NADPH 2 → CH 2 + NADP + + H 2O
+
CH 2 CH 2
COOH COOH
The product, L-glutamate, can then be the
amino group donor to transfer the amino group
to other α-keto acid to form other amino acids.
When the ammonia concentration is low, E.
coli employs an energetically more expensive
pathway with a higher affinity for ammonia (a

4 Stochiometry
 lower km) to assimilate ammonia. The enzyme
catalyzing the reaction is glutamine synthetase.
COOH COOH
H 2 NCH H 2 NCH
CH 2 + ATP + NH 3 → CH 2 + ADP + Pi
CH 2 CH 2
COOH CONH 2
The amino group in glutamine can be transferred
to α position of α-ketoglutarate to form
glutamate as catalyzed by glutamate synthase.
Glutamine + α-ketaglutarate
+ NADPH2 → 2
+
glutamate + NADP+
The second molecule of glutamate is recycled
to be used in the slutamine synthetase
catalyzed reaction. The net reaction is
a − ketoglutarate + ATP + NH 3 + NADPH 2 + → glutamate + NADP
In the above two examples the same chemical
conversions, from glucose to ethanol and from
α-ketoglutarate and ammonia to glutamate, are
accomplished. But different biochemical pathways
are used with different energetic efficiency.

Stoichiometry for cell growth

From the stoichiometric and energetic point of
view, cells degrade and convert the nutrients (i.e.
carbon and energy sources for chemotrophs) to
derive chemical energy and produce biomass and
product. The chemical energy is used to assimilate
NH3 (in microbes and plants), to make other building
blocks (fatty acids, amino acids, nucleic acids, etc.)
and to make polymeric components of biomass
(proteins, lipids, nucleic acids, etc.) and to grow.

Stochiometry 5
 The cost of making polymers (protein, nucleic
acids) from building blocks is rather high. The
incorporation of each mole of nucleotide into DNA
or RNA requires 2 moles of ATP. The synthesis of
each peptide bond or the addition of each amino
acid, through translation costs at leat 4 ATP. The
energetic cost of synthesizing a unit amount of
biopolymers which constitute the bulk of biomass
thus can be estimated, at least in principle.

In addition to assembling the materials to make

cell mass, cells also maintain a membrane
potential (pH and small electric gradient) and an
osmotic pressure across membrane, they also
take up or excrete some compounds against
gradient, and then all require energy. Thus, a large
portion of nutrients taken up by cells is used for
generation of energy, to sustain their functions.

In many cases the system on which the material

balance is to be performed is “abstract”, without
a real physical entity. In other cases the material
balance is performed on a physical system, such
as a bisector, a cell or a population of cells. For
example, one can balance all glycolysis reactions in
the cell. In this case the system is a set of reactions
catalysed by the glycolytic enzymes, not a physical
entity. The inputs are glucose and other compounds
needed to carry out the reactions, including NAD+,
ADP, Pi, and the outputs are pyruvate, NADH2+, ATP.

Here one can also see a difference between

balancing on a reaction system and on a cell with
respect to the same pathways. Take fermentation
of glucose by yeast, Saccharomyces cerevisiae,
as an example. Balance on the cell using
glucose as feed (input) and ethanol and CO2 as
the products (output). If the yeast is resting (not
growing) and all the glucose consumed goes to
ehanol, then the balance on the cell as a system
is glucose → 2 ethanol + 2CO2. On the other
hand, if one balances on the reaction system or
implicitly balancing on intracellular glycolysis and
fermentation reactions, then the balance equation is:
glucose + 2ADP + 2Pi → 2

6 Stochiometry
 ethanol + 2CO2 + 2ATP+2H2O

ADP, Pi and ATP do not cross cell cytoplasmic

membrane. Macroscopically glucose is the only
input into the cell, and 2CO2 and ethanol are the only
outputs. One can view the cell as an overall system,
the intracellular ethanol production as a subsystem.
There must be another subsystem intracellularly
that convert ATP to ADP and Pi, so that the sum
of the two subsystems equals the overall system.
Biomass Formula That subsystem is the many intracellular reactions
or “work” which consumes ATP. For example the
mechanical work and osmotic work both require
ATP. Thus, two subsystems, one generating
ATP the other consumes ATP, are coupled.

A chemical formula can be written for any

compound or a chemical species with a defined
composition, such as glucose, glutamic acids, a
protein like human insulin, a molecule of DNA. For
a mixture of molecules with known composition,
such as a mixture of proteins, one can derive a
weighted average molecular formula. For example
for a mixture of proteins consisted of n species
Cαi Hβi Nγi Oδi, each with a mole fraction ai., the
average molecular weight can be expressed as

MWAve =
∑a M
W i i

The average molecular formula C∑ a H

α’i
i
β’i

Nγ’i Oδ’i , will have their coefficient as

∑ ai a i
ai ’
=
∑ ai
However, when the molar composition of the
protein mixture is uncertain, as in the case of the
cellular proteins, this approach is not applicable.
One has to resort to various methods of estimation.
In the same vein, if the molar composition of the
cell, including all the biopolymers and building
blocks, is all known, by summing up the equations
for all those components, one will be able to write
a “formula” of the cell. In practice the precise molar
formula cannot be easily written by summing
up all components in the cell and developing
a weighted average formula. Rather, a cell

Stochiometry 7
 cormula can be written, but largely by doing an
elemental analysis (usually considering only key
component elements, C, N, O, H, maybe also P, S).

With a formula for biomass, one can develop

a stoichiometric equation for converting
nutrients to cells. Considering the case cells
take up organic nutrients, and use oxygen and
ammonium additionally. The stoichiometric
equation for “synthesizing” the cell, from nutrients,
and those for transforming building blocks
into polymer and other components of cells.
Ca i H b i Ngi Od i + aO2 + bNH 3 → cCa ' H b ' Ng 'Od ' + dCO2 + eH 2 O

Where subscript i indicates different nutrients,

if there are multiple organic substrates.
The formula of biomass is basically the
molar elemental composition of cells.
The energy for biosynthesis of biomass is
derived from catabolism of nutrients. In most
living organism, carbohydrate metabolism
through glycolysis and tricarboxylic acid (TCA)
cycle contributes most to energy generation.
Upon complete oxidation of glucose to CO2
and H2O via TCA cycle, the energy efficiency
is calculated from the stoichiometric equation

C6H12O6 + 6O2 + (30~33)ADP +36 Pi

→ 6CO2 + 6H2O + (30~33)ATP H2O
stoichiometric coefficient of ATP
varies with organisms. It depends
on the efficiency of ATP sythase.
If one considers biomass as an assembly of all
the other constituents, one can write the equation”
a ' Ca " H b " N g "Od " (average protein formula) +
a "Ca " H b " N g "Od "
(average DNA formula) + …
(RNA) + … → biomass

The energetics of assembling the constituents

into cells cannot be easily estimated. With the
uncertainty about the energetic efficiencies as
described above, it is very difficult to obtain a
biomass equation with a high degree of confidence.

Another difficulty in developing a stoichiometric

8 Stochiometry
 equation for biomass arises from “turn-over” of
cellular materials. Both protein and mRNA “turns
over” in the cells. In other words, after being
synthesized they serve their function for a period of
time, and then get degraded. Some components
become degradation products and are excreted
or catabolized, others become monomers
like amino acids and recycled to build cellular
biopolymers. The balance in the intracellular
concentration of protein P is described as

dP
=a - β P - μ P
dt
2 O
where α is the average synthetic rate, β is the rate
constant for degradation (turn over), and μ is the rate
of cell expansion which causes P, the intracellular
concentration of proteins, to decrease due to
the expansion of cell volume. The degradation
rate of protein and mRNA and varies widely. The
formula for the synthesis of biopolymers can
reasonably accurately incorporate energetic cost
(i.e. specify the number of moles of ATP used to
synthesize a mole of biopolymer), or can specify
the energetic cost associated with α. However,
knowing steady state cellular composition P is not
sufficient to determine α unless β and μ are also
determined. In practice, even though μ can be
determined accurately, β is largely undetermined.
In other words, cells with the same P
and the same μ may have very different
α and β even both have the same dP/dt.

Other factors that contribute to the wide range of

diversity in the biomass formation equation, even
among microorganisms of similar composition, is
the energetic components which are not directly
related to “cell composition”, or to the synthesis
of biomass. Such factors include the energetic
cost of transport and maintenance of membrane
potential and osmotic pressure. Nutrient uptake
and product excretion often require energy. Some
nutrients are transported across cell membrane by
passive or facilitated diffusion for which there is not
a net energy cost. For many others, the transport
is coupled to the concentration gradient of Na+/
K+ or H+ across the membrane. In other cases
the transport is directed coupled to the hydrolysis
of ATP. The maintenance of the concentration

Stochiometry 9
 gradient of a chemical species requires energy
Stoichiometric Equation for (otherwise, no matter how small the permeability
Bi
Biomass F
Formation
i the cellular membrane is, eventually the two sides
of the membrane will reach equilibrium). Transport
C6H12O6+ of molecules utilizing H+ or Na+/K+ gradient is
¦ i CD i H E i NJ i OG i  aO
O2  bNH
N 3 o cC
CD ' H E ' NJ 'OG '  dCO
CO2  eH 2 O thus energy dependent indirectly. In many cases,
the cost of those transport process are not
Inputs Outputs clearly known. In Eukaryote, organelles provides
Subsrates consumed ĺ biomass produced+ (CO2 + water) produced
compartmentalization in all the cell. Electric or
Can write four balance equations on elements carbon, nitrogen,
pH potential exists across those membranes
hydrogen, and oxygen
There are four unknown
separating the cytosol and lumen of organells.
Reaction intermediates are transported across
these organelles. Maintaining these gradients and
transport across orgenell is also energy dependent.

The stoichiometric equation for biomass can be

written to include many atomic species, especially
P, S. However, usually it includes C, H, N, and
O. For each elemental species, an elemental
balance equation can be written. In the case that
a biomass equation considers only C, H, N, and
O, four conservation equations can be derived
for C, H, N, O. In the stoichiometric equations for
biomass using NH2, O2 and a carbon source five to
six chemical species are involved in the reaction.
The four elemental balance equations are certainly
not sufficient to determine the five stoichiometric
coefficients. The equation is thus undetermined.
One has to rely on experimentally measured
values to obtain the stoichiometric coefficients.
Unlike simple stoichiometric chemical reaction
equations like CH3 CO COOH → CH3 CHOH
COOH, for which a unique set of stoichiometric
coefficient exists, a wide range of the combination
of stiochiometric coefficients exist for biomass
equation. There is thus no single equation for all
biomass forms. There is a wide diversity on how
organisms use the same raw materials to build cells.

The biomass equation presented in this chapter

describes only the conversion of substrate into
biomass. Implicitly we considered only the organic
matters in the cells only. In addition to their organic
constituent, cells contain a large amount of water.
In fact, the water content of cells is typically from
75-85%. In plant cells the fraction can be even
higher under some conditions. The stoichiometric
equation described above considers only the
dry biomass, not wet biomass. In general, the

10 Stochiometry
 symbol, x, will be used to denote dry biomass.

Like writing an average molecular formula

for a mixture of compounds, there are many
different ways to present the biomass formula.
Conventionally one can set one atomic coefficient
(usually carbon) to be 1.0 or set the formula weight
to be equal to a convenient number such as
100. Biomass is made up of more than the four
elements of C,H,N,O. Other major constituents
include P, S, Mg, Fe, and other minerals. Those
are often referred to as ashes, since their mass
is determined after treatment of cells in a high
temperatures (~450oC) furnace to complete oxidize
the combustible portion of the cells. Therefore
with a formula Cα, Hβ, Nr, Oб, the formula weight
accounts for only the portion excluding the ashes.

It should be obvious that since the biomass

composition is dependent on the cultivation
condition and on the growth stages, both
the biomass formula and the values of the
stoichiometric coefficients for the biomass equation
change accordingly. The biomass composition
varies with other physical, chemical variables in
the environment. When a complex medium is
used for cultivation or when additional products
(in addition to CO2 and H2O )are produced, other
terms are added to the equation accordingly.

Biomass formation equation is basically a summary

of the material balance (i.e. stoichiometric
relation) of nutrient uptake and the outputs of
cells and excreted products. Experimentally,
the equation is merely an approximation, since
a precise measurement of biomass formula is
very difficult to obtain. Furthermore, the equation
always entails production of H2O, CO2 and
consumption of O2. For CO2 and O2, experimental
measurements are at least available, although
accurate determination is not easy. For H2O, the
measurement is extremely difficult since cells grow
in aqueous environment. Therefore, the utility of
the biomass formula is largely in the analysis of
material’s flow using material balances (metabolic
flux analysis) for which the formation of biomass
must be accounted for in the overall balance.
Conversion and Yield Coefficient

Stochiometry 11
 The stochiometric equation of biomass can
be seen as a conversion of new materials (or
input) into cells and other products (outputs).
A yield coefficient is often used to describe
the conversion efficiency. This term yield is
loosely defined and used in different units.
Virtually any pair of output and input can be
combined to give a yield coefficient. The most
commonly used ones are shown in Table III.

The basic form of yield coefficient is

ΔX
Yx / s =
In the Δcase
S that multiple nutrients and products
are invovlved, for example, in mammalian cell
Yield on biomass = mass of biomass produced culture, a stoichiometric ratio (βίј) for a pair of
mass of substrate consumed nutrients/products is often used. For example, in
mammalian cell culture, a large fraction of glucose
consumed by cells is converted to lactate. Under the
condition that glucose concentration is controlled
at a very low level, much less lactate is produced.
It is not dimensionless.
dimensionless The units can be g cells/g The stoichiometric ratio of lactate to glucose is
glucose. The value is dependent on the units often used to characterize the metabolism of
chosen for the expression.
p
glucose. Yield coefficient and stoichiometric ratio
can be applied to a wide variety of substrates,
with a variety of units. Referring to the biomass
formation equation, on a mass basis it is basically
Yield coefficient of biomass the product of the stoichiometric coefficient of
the biomass and the formula weight of biomass
• Determined experimentally divided by the product of the stoichiometric
• The efficiency differs from organisms, coefficient and the formula weight of the substrate.
growth conditions
conditions, and substrates

The biomass composition and the biochemical

pathways used for cell growth may change
under different conditions. As a result, the
Yield Coefficient on Biomass biomass stoichiometric equation, and the
• Is affected by the type of substrate
yield, may also change. Thus, yield can be an
Dry Biomass
E coli growing on different carbon sources instantaneous value obtained at a particular
time point; alternatively, it can be an average
glucose

succinate
succ ate

slope = yield value over a long period of cultivation.

S b t t consumed
Substrate d

• Is affected by the medium, growth condition (yield is higher at optimal

conditions, i.e., optimal pH, temperature, osmolality, nutrient composition dx
Aerobic
Yx / s =
Yeast grown aerobically and Anaerobically
ds
Dry Biomass

Anaerobic

Substrate consumed

12 Stochiometry

Partition of a substrate for
biomass and product formation
xt 2
Yx / s =
∫
xt 1
dx
st 2
∫
st 1
ds
Depending on the nature, a nutrient (or commonly
called substrate), may be either completely
conserved in biomass, or only partially converted
to biomass. In the latter case, the rest is excreted
as converted product. For example, carbon
source usually serves as energy source (except
in photosynthetic organism), thus, part of it is
converted to biomass, and the rest is excreted
as CO2 and other products. The fraction of
carbon atom conserved in biomass is always
less than 1.0. For Na, K, Fe, basically all that is
taken up by cells is retained in biomass for nearly
all chemoorganotrophs. The yield coefficient
based on atom basis is 1.0 mole cellular atom/
mole uptake atom. For nitrogen source, the
yield is variable depending on the organism and
the growth conditions. If there is no excreted
nitrogen containing compound, then the yield
is basically 1.0 mole cellular N/mole uptake N.
Knowing the theoretical maximum efficiency that
a raw material can be converted to a product
is important in process design and innovation.
The return on the investment of improving the
production yield is higher if the yield is substantially
below the theoretical maximum than if the yield is
already approaching the maximum. To estimate
such a maximum one needs to consider both
material and energetic aspects. Largely one
needs to consult the biochemical pathways of
the organism. The approach to determine the
theoretical product yield is dependent on whether
the pathway is energetically favorable or not.
Products that are converted directly from the raw
materials in an energetically favorable fashion.
In other words, the producing cell confers a
biochemical pathway to convert the raw materials
to the product with a net generation of chemical
energy. For example, for ethanol production by
yeast, one mole glucose can be converted to two
moles of ethanol, and generates two moles of
ATP. The reaction in thus energetically feasible,
Stochiometry 13
 and the theoretical maximum is (2x48/180=)
0.51 Kg ethanol/Kg glucose. The true yield in
process is lower because some glucose will be
diverted to make biomass and for the cells other
energetic needs. The basic way of calculating the
theoretical yield of such products is to tabulate the
pathway and examine its energetic conversion.
The second example, production of glutamic acid
from glucose and ammonia in corynebacterium
glutamican encompasses the following pathways.
a. Synthesis of a product is energetically feasible
if a net production of ATP and/or 1 NADH, results in
the process. As seen in the example, the theoretical
maximum conversion based on carbon is 5/6 (mole
carbon in glutamate/mole carbon in glucose), or
on a mass basis (143/180=) 0.79 for glutamine.
b. Products which require energetic contribution
of energy derived from the catabolism of raw
materials.Some products from biosynthetic
pathways require the catabolism of substrate
to make the reaction energetically favorable. In
this case, from the biochemical pathways the
biosynthetic reaction equation requires the supply
of energy in the form of NADPH/NADH or ATP.

aC a H b Og N d ( substrate1) + bC a' H b ' Og ' N d ' ( substrate 2) + cATP →

The energy must be supplied by catabolism
of the substrate, assuming catabolism
is primarily derived from substrate:

Ca H b Og N d + b ' O2 + d ' ADP + d ' Pi → c ' C

O 2 + d ' ATP

The maximum yield is the combination of the two

reactions that gives a net production ofATPequivalent
is zero. For first appropriation, one can assume one
mole NADH is equivalent to three moles of ATP.

C. Metabolic engineering of the pathways. In the

past two decades our ability to alter the genetic
makeup and their metabolic pathway has been
greatly enhanced. This has enabled one to con-
struct new pathways by introducing enzymes
which are not normally present in the producing
organism. Many of such efforts are, of course,

14 Stochiometry

cATP → Ca " H b "Og " N d " ( product ) + cADP + cP
aimed at producing new products. Many others
are aimed at alternating the carbon or energy
efficiency of the process. The estimation of the
maximum theoretical yield of a product from
a substrate is now not just based on the path-
ways in the producing organism, but on path-
ways, or the combination of segments of path-
ways, available in the living system. One may
even take one step further by combining with
protein engineering to create new enzymes to
facilitate improved carbon flow or energy effi-
ciency for product formation. Therefore, maxi-
mizing the thermodynamic efficiency for the
conversion of substrates to product is desired.
Now that we learned how to determine biomass
yield and the theoretical yield for products we
turn our attention to the case that biomass and
product are performed simultaneously. For aerobic
organisms growing without producing any product
(other than CO2, H2o)), the substrate consumed
can be largely divided into two categories. One part
is incorporated into biomass, the other combusted
to CO2 and H20 to generate energy for synthesis
i
of biomass. The yield is the ratio of biomass to
the mass of substrate consumed (combining both
parts of substrate). When a product is formed,
one can hypothesize that a third part of substrate
is consumed for product formation. The amount
of substrate goes to the third part is estimated
from the theoretical yield. Since biomass yield
can only be estimated experimentally, one can
subtract the substrate consumed for product
formation from the total substrate consumed to
estimate the substrate that is used for biomass
(including the part incorporated into the cell
and the part used for energy generation).
For anaerobic process, recall that product
formation and growth is coupled. The organic
acid or solvent formation is part of the process
generating the energy from substrate. Since each
mole of CO2 derived from sugar produces only small
amounts of energy, the vast majority of substrate
carbon is converted to product and only a very
small amount goes to biomass. The estimation of
Stochiometry 15
 biomass yield is not critical in most process design.

16 Stochiometry
Stochiometry 17
18 Stochiometry