Engineering of Biological

Processes

Lecture 3: Yields and stoichiometry
Mark Riley, Associate Professor
Department of Ag and Biosystems
Engineering
The University of Arizona, Tucson, AZ
2007
Objectives: Lecture 3
Biosynthetic processes (anabolic)
Case studies - cholesterol
Stoichiometry and modeling cellular
requirements
"You are what you eat"

Stoichiometry
Provides information on fundamental
constraints
Substrate conversion to product
Cell mass from substrate

Yields and yield coefficients
Mass based = kg of this from kg of that
Y
(output / input)
Y
x/s
Y
p/s
Y
ATP/O2

Y
m
x/s
maximal yield of cell mass from
substrate
Yield
Overall
Instantaneous

Ratio of rates
Ratio of yields

Theoretical = Y
Observed = Y
YIELD
Cell metabolism
Y
lactate / glucose
= ranges from 2 to 0 based on environment

The basic reaction is:
Glucose + 2 P
i
+ 2 ADP 2 Lactate + 2 ATP + 2 H
2
O
Glucose [g/L]
Bacterial
dry cell weight
[mg/L]
Slope = dX/dS
7 (mg/L) / (g/L)
Yield of cell mass from substrate
Y
x/s
Glucose [mM]
Bacterial
dry cell weight
[g/L]
Aerobic
Y
x/s
=58 mg/mol
Anaerobic
Y
x/s
=22 mg/mol
Cell composition
Dry weight vs. wet weight
70% of the composition is water

Dry weight consists of:
Element E. coli Yeast
C
O
N
H
P
S
K
Na
Others
50%
20%
14%
8%
3%
1%
1%
1%
<1%
50%
34%
8%
6%
1%
<1%
<1%
<1%
<1%

CH
x
O
y
N
z
In a very simplistic interpretation of
metabolism, the following applies:

Cells + medium + O
2
(sometimes) more cells +
product + CO
2
+ H
2
O

Medium contains sugars, amino acids,
cofactors and the elements in the previous
table.


Stoichiometric calculations
Based on 1 mole of C in the input

CH
m
O
n
+ a O
2
+ b NH
3


c CH
a
O
b
N
d
+ dH
2
O + eCO
2


This is normalized to 1 mole of C.
Could also be normalized to 1 mole of the C source compound

Perform elemental balances to determine the
unknown values of the cofactors

Example
C
6
H
12
O
6
+ a O
2
+ b NH
3


c C
4.4
H
7.3
O
1.2
N
0.86
+ dH
2
O + eCO
2

2/3 of the glucose C goes to biomass
What are the stoichiometric coefficients,
and Y
x/s
, Y
x/O2
? MW
glucose
= 180
MW
cell
= 89.62
MW
oxygen
= 32
MW
ammonia
= 17
Generalized growth reaction
C
6
H
12
O
6
+ a NH
3
+ b O
2
a CH
1.8
O
0.5
N
0.2
+
b CH
x
O
y
N
z
+ gCO
2
+ dH
2
O

Normalized to 1 mole of carbon source compound
Where a, b, a, b, g, d, x, y, z depend on the type of cell
involved.
a, b, a, b, g, d, are stoichiometric coefficients
When little info is available about cell composition, use
an approximated cell composition of
CH
1.8
O
0.5
N
0.2

This yields a MW of a cell ~ 24.6

Generalized growth reaction
180
6 . 24
Y
s x
a

C
6
H
12
O
6
+ a NH
3
+ b O
2
a CH
1.8
O
0.5
N
0.2
+
b CH
x
O
y
N
z
+ gCO
2
+ dH
2
O
g of cells from g of glucose
Lack of information
Unfortunately, the elemental balances
often do not provide enough information
to completely solve for the stoichiometric
coefficients.
Respiratory quotient
RQ = Y
CO2/O2

Molar basis
Moles of CO
2
produced from moles of O
2

Provides information on the metabolic state of the cell

A high RQ means that much CO
2
is produced and
hence the metabolism is operating at high efficiency
Aerobic metabolism
CH
m
O
n
+ a O
2
+ b NH
3


c CH
a
O
b
N
d
+ d CH
x
O
y
N
z
+ eH
2
O + fCO
2

RQ = ?
Degree of reduction
Electron balance
g = # of available electrons / g of atomic C
g Or, this can be described as:
g = # of available electrons / # of Cs

Provides another independent equation
Degree of reduction
C = 4
H = 1
N = -3
O = -2
P = 5
S = 6
CO
2
= +4 (C) + -2 (O) = 0

C
6
H
12
O
6
= 6(4) + 12(1) + 6(-2) = 24
g = 24 / 6 (# carbon atoms) = 4

C
2
H
5
OH = 2(4) + 6(1) + (-2) = 12
g = 12 / 2 (# carbon atoms) = 6



Example yeast grown on glucose
C
6
H
12
O
6
+ 0.48 NH
3
+ 3 O
2

0.48 CH
1.8
O
0.5
N
0.2
+ 3.12CO
2
+ 4.32H
2
O
To grow yeast to 50 g/L in a 100,000 L reactor, determine:
a) mass of glucose and ammonia required
b) O
2
required
c) Y
x/s
and Y
X/O2

MW
glucose
= 180
MW
cell
= 24.6
MW
oxygen
= 32
MW
ammonia
= 17
HW #1 questions
1) What kind of cell would you use to produce
androstenedione? Your answer should
describe the attributes of such a cell (don't
just state, "a cell that produces andro"). An
answer longer than 4 sentences is too much.
2) Producing cholesterol is an energy intensive
process. How much energy (in terms of # of
ATP molecules) is consumed in producing
one cholesterol molecule from a source of
glucose?