Extended Monod Kinetics for Substrate,

Product, and Cell Inhibition

Keehyun Han* and Octave Levenspiel

Department of Chemical Engineering, Oregon State University,
Corvallis, Oregon 9733 1
Accepted for publication August 6, 1987

A generalized form of Monod kinetics is proposed to account for all kinds of product, cell, and substrate inhibition. This model assumes that there exists a critical
inhibitor concentration above which cells cannot grow,
and that the constants of the Monod equation are functions of this limiting inhibitor concentration. Methods for
evaluating the constants of this rate form are presented.
Finally the proposed kinetic form is compared with the
available data in the literature, which unfortunately is
very sparse. In all cases, this equation form fitted the
data very well.

For substrate inhibition:

for product inhibition:

for cell inhibition:

INTRODUCTION
Although microbial growth is a very complex phenomenon, the overall growth can often be regarded as a single
chemical reaction with a simple rate expression. Many
equations are used for this purpose. Among these, the simplest and most popular is the one proposed by Monod
who assumed that a single essential substrate is the growth
limiting factor. I Monod kinetics can be expressed as
Substrate (A)
r, = k

Cells (C)

more Cells (C)

+ Product (R)

cell mass produced

where r, is the growth rate of cells; k is the reaction rate

constant; CA is the substrate concentration; C, is the cell
concentration; and C, is the Monod constant.
Unfortunately, there are often inhibitory effects in the
real world. In this article, we propose a generalization of
the Monod expression which takes into account inhibition
effects caused by high concentration of either substrate, or
cell, or product, or other inhibitory substance. This proposed expression is

In the extreme where C, -G C,: the above eqs. (2)-(5) all

reduce to the simple Monod expression of eq. ( 1 ) .

HOW TO EVALUATE THE CONSTANTS OF THE

GENERALIZED MONOD EQUATION FOR
PRODUCT OR CELL INHIBITION
As with Monod kinetics, the constants in eq. (2) with
C, = C, or C, = C, can be evaluated from a Lineweaver-

Burk plot of C , / r , vs l / C A .Thus inverting eq. (2) gives

cc

- -

r,

C,(1 - c,/c:)m1
k ( l - C,/Cf)" CA

-+

1
k ( l - C,/CF)"

This expression can account for and represent the six common patterns of inhibition illustrated by the LineweaverBurk plot of Figure 1:

* Present address: Biochemical Engineering Program, School of Engineering, University of California, Irvine, CA 92717.

(a) represents noncompetitive inhibition, where n > 0

and m = 0;
(b) represents competitive inhibition, where n = 0 and
m < 0;
(c) represents generalized uncompetitive inhibition, where
n>m>O;
(d) represents generalized uncompetitive inhibition, where
m>n>0;

Biotechnology and Bioengineering, Vol. 32, Pp. 430-437 (1988)

0 1988 John Wiley & Sons, Inc.

CCC 0006-3592/88/040430-08$04.00

where C, is the inhibitor concentration, C: is the critical

inhibitor concentration above which reaction stops, and n
and m are constants.

I ///

( a ) noncompetitive
m=O, n>O

(d) uncompetitive
m>n>O

(b) competitive
m<O, n = O

(c) uncompetit ive

n>m>O

(e 1 uncompet i t i ve
m=n>O

( f ) general case
m<O, n>O

Figure 1. Lineweaver-Burk plot for various forms of product or cell inhibition, based on the
expression of eq. (2).

(e) represents uncompetitive inhibition, where n = m >

0; and
(f) represents the general case, for any n > 0 and m < 0.

and on taking logarithms

kobs and CMobs are then given by the intercepts and abscissas on Figure 1, as shown in Figure 2. Note that

y;:

kobS = k 1 - -

with constants k and n

+ h(k)

(9)

ln(CMds)= m ln(1 - C,/C,*) + ln(C,)

(10)

ln(kobs)= n ln(1 - C,/C:)

and

(7)

Consider the evaluation of k and n. For this, make a series of runs at different inhibitor concentration. If the value
of C: is known from experiment, and it usually is, then a
plot of ln(kobs)vs ln(1 - C,/Cf), as shown in Figure 3,
will give k and n from eq. (9). If C,* is not known, guess
C: until a straight line is obtained, then proceed as above.
CMand m are found in a similar way with eq. (10).

and
with constants CMand m

(8)

In ( kobJ

Figure 2. Evaluation of kms and CMh requires making series of runs at

various concentrations of inhibitor.

find k from this point,

Figure 3. Procedure for evaluation of k, n, and C,? for either cell or

product inhibition, adapted from ref. 4.

HAN AND LEVENSPIEL: EXTENDED MONOD KINETICS

43 1

Example of Product Inhibition

Aiba and co-workers5 also proposed an equation form to

represent product inhibition. Using the best values of the
constants of their equation form, we find

The best example of product inhibition is in alcohol fermentation, in which alcohol, the product of reaction, inhibits cell growth.' Bazua and Wilke3 made 108 runs in a
well mixed flow reactor, or CSTR, where each run gave a
value of rate at a specific C , , C R , and C c . Plotting this
data as in Figure 2 gives values of kobS and CM,,bs shown in
Table I.
Since CM0bs does not seem to change in any systematic
manner with inhibitor concentration, CR, we take it to be
constant. This means that the product of reaction, alcohol,
acts as a noncompetitive inhibitor, and the following equation form4 applies
rc/Cc = k(1 - CR/Cf>"CA/(CA + C,)

kobs

The generalized Monod equation for substrate inhibition

is written as eq. (3). Unfortunately, the method described
for cell or product inhibition cannot be used for substrate
inhibition because there is no difference between substrate
and inhibitor. Since the Lineweaver-Burk plot method
cannot be used, we proceed as follows: first evaluate k and
n at high C, ; then evaluate C , and rn from the data in the
low range of C, values.
When C, % CMobs eq. (3) simplifies to

(12)

Let us compare this result with other proposed equations.

First of all Bazua and Wilke3 represented their data by

rC/cC
kDbs

- CR/93.6)'.'

= 0.448(1

(13)

c,

kCbs

CMobr

(g/L)

(h-')

(g/L)

4.37
29.19
61.29
81.30

0.415

0.230
0.230
0.259
0.223

0.368
0.289
0.141

= k(1

- cA/C,*)"

( 15)

and on taking logarithms

Table I. Values of the observed rate constants as extracted from the

experimental results of ref. 3.

c,

(14)

HOW TO EVALUATE THE CONSTANTS OF THE

GENRALIZED MONOD KINETICS FOR
SUBSTRATE INHIBITION

(11)

0.424(1 - CR/83.57)0.3w

.oi3cR)

The fit of the experimental data to these proposed expressions, eqs. (12)-(14), is shown in Figure 5. As can be
seen, the model proposed here fits the data well.

Since C,* was not reported in the original article the procedure of Figure 3 is then used to find C,*, k , and n. The
final result, shown in Figure 4, is then
kobs

= 0.494 exp( -0

As with product or cell inhibition, when the C,* value is

not known, the trial error method of Figure 3 using
eq. (16) gives the values of C,*, k , and n. Next, for the
low range of C, values we rearrange eq. (3) to give

In (kobr)
-0.86

Y intercept = In k

I \

q--.

:. k = 0.424 h i '

0.4

C; = 83.57 g A
gives a straight

&

0.3

0.2

4-2.0
c

- 4.0

::,

-2.0

-3.0

-1.0

In I--

Figure 4. Evaluation of k, n, and C t of eq. (11) for alcohol fermentation; data from ref. 3, see Table I.

432

0.1

from' Aiba et a15

tt

eq.'13
from Bazua and Wilke3

20

40

60

, I
80

alcohol concentration (g/.t)

Figure 5. Increasing the concentration of inhibitor (alcohol) depresses
the observed rate constant.

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 32, AUGUST 1988

Calculate C,,, from eq. (17), and plot as in Figure 6. The

slope and intercept on this graph gives C, and m.
0 : data

from Uemura et aI6

Substrate Inhibition: Example 1

Uemura and co-workers6studied the microbial attack on
n -pentane by a pentane-consuming bacterium (JTB-271).
The results were read from Figure 5 of Edwards' paper,7
and are shown in Table 11. A plot of these data points in
Figure 7 clearly shows substrate inhibition since the rate of
reaction slows at high substrate concentration. When C, 2
65 mmHg, assume that C, S CMobs, in which case eq. (3)
is well approximated by eq. (15). It will be shown later
that this is a reasonable assumption.
Following the procedure outlined above, we first make
the plot of Figure 8, from which we find C,* = 700 mmHg,
k = 0.117 h-', and n = 1.82. CMobs
values are then calculated from eq. (17) and listed in Table 111. Finally, the plot
of Figure 9 gives C, = 30 mmHg and m = 66.8. At C, =
65 mmHg, we find CMobs
= 0.045 mmHg, hence the assumption that C, S CMobs
and m > 0 is well satisfied. The
final result is then

eq. 18, best fit of the

general Monod equation

0 '

200

400

600
partial pressure of n-pentane (mm Hg)

Figure 7. Data of ref. 6 clearly shows that high concentration of

n-pentane, the substrate, inhibits the reaction; see Table 11.

(18)

This expression fits the reported data well, as shown in

Figure 7.

-2.8

-3.0

-0.4

In I--

::)

-0.2

Figure 8. This graph gives k, n, and C,* for n-pentane substrate inhibition; data of ref. 6.

0
Table III. C,,

value extracted from ref. 6.

Figure 6. For substrate inhibition m and C , are evaluated at low C,

Table 11. Experimental result of ref. 6 taken from Figure 5 of ref. 7.

5.000
6.875
10.938
15.313
19.688
25.625
30.625
65.000
1oo.OOo
135.625
199.375
268.7 50

0.0227
0.0378
0.0564
0.0792
0.0911
0.0992
0.0992
0.0991
0.0817
0.0786
0.0633
0.0489

5.000
6.875
10.938
15.313
19.688
25.625

0.0227
0.0378
0.0564
0.0792
0.091 1
0.0992

20.44
14.03
11.11
6.42
4.32
2.61

Substrate Inhibition: Example 2

Kortan* reported substrate inhibition by n -butanol to
Arthrobacter AK 19. The experimental results of Table IV
are from Figure 2 from the article by Wayman and Tseng.'
As in the previous example C,*, k, and n values are obtained from Figure 10. In this example, we assume that
eq. (3) is well approximated by eq. (15) when C , 3
HAN AND LEVENSPIEL: EXTENDED MONOD KINETICS

433

In (CkbJ
L

0.2 vol %. Then C , and m values come from Table V and

Figure 11. The final expression from our model is:
n

rc/Cc = 0.29(1 - CA/0.98)

CA

LA

+ 0.014( 1 - CA /0.98)23.2
(19)

- 2.0

Wayman and Tsengs model, with values of constants

taken from their article, gives
r

rc/Cc

- 1.0
,I

-0.03

-0.04

In

-0.02

0.309

when CA< 0.03

+ 0.011

CA

0.309[cA/(CA

(1-2)

L A

and

rc/Cc

-0.01

+ O.Oll)]

0.328(CA - 0.03)

when CA> 0.03

The fit of our eq. (19) and Wayman and Tsengs eq. (20)
to Kortans data is shown in Figure 12.

Figure 9. Evaluation of m and C , for n-pentane substrate inhibition;

data of ref. 6, see Table 111.

DISCUSSION

Table IV. Experimental result of Kortan taken from Figure 2 of ref. 9.

Many inhibition models have been proposed in the literature; some only for substrate inhibition, others only for

C*
(vol. S)

CKC

0.0125
0.02
0.03
0.05
0.06
0.09
0.2
0.3
0.4
0.5
0.6
0.8

0-l)

Table V.

0.155
0.215
0.230
0.255
0.260
0.258
0.235
0.205
0.175
0.132
0.110
0.060

c*=
(vol. %)

CMobvalue extracted from the data of ref. 8.

CM,,
(vol. %)
~~~

0.155
0.215
0.230
0.255
0.260

0.0125
0.02
0.03
0.05
0.06
a

0.0107
0.0066
0.0069
0.0043
0.0033

low concentration range only.

In ( kobJ

Y intercept = In k = -1.23
k = 0.29 h i

.;

.1

slope: n = ~

/
4

-2.0

-1.5

-2.0

this <ne is straight

I

-1.0

- 0.5

4-2.8
I

1-3.2

- 0.06

434

- 0.04

- 0.02

1-6.0
0

( );

( );

In I - -

Figure 10. Evaluation of k , n , and C: for n-butanol substrate inhibition; data of ref. 8.

In I - -

Figure 11. Evaluation of m and C , for n-butanol substrate inhibition;

data of ref. 8.

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 32, AUGUST 1988

concentration predict that inhibitor concentration should be

infinite to stop the reaction.

product inhibition, as shown in Table VI. However the kinetic form proposed here should be useful for all forms of
inhibition. In addition if there are no inhibitory effects (n =
0 and m = 0), the generalized equation reduces to simple
Monod kinetics.
Finally, the key characteristic of the proposed model is
its assumption that there exits a critical inhibitor concentration above which reaction ceases. This is what actually
happens in many, if not most situations. Many of the product inhibition models incorporate a critical inhibitor concentration. On the contrary, for substrate inhibition only
Wayman and Tsengs model includes this feature. Consequently, those models which do not have a critical inhibitor

Substrate Inhibition
Edwards compared the goodness of fit of five kinetic
models for substrate inhibition to the reported data of
Uemura et aL6 Table VII lists these models, Table VIII
gives the best-fit value of the constants of these models,
and Figure 13 displays the results as dotted or dashed
lines. Also included in this figure is the curve from the
model developed here, or eq. (18). This figure shows that
one should be able to more clearly discriminate between
models at high substrate concentration.

Table VI. Kinetic models for inhibition; equations adapted from p. 256 and p. 259 of ref. 10.
Equation

Reference

For substrate inhibition:

1

-rC -- k

Andrews and Noack

+ c M / c A + CAIK/

CC

Webb
Yano and co-workers

Aiba and co-workers

c = k [exp(-CA/Kl) - exp(-CA/C~)]

Teissier-type

CC

(uis ionic strength)

Webb

Wayman and Tseng

c = k -

CA

CC

+ CA

cM

- K1(CA- C J ,

when CA> C i

For product inhibition:

c=kCC

CA
CM

rC -k

+ CA

(1 - KCR)

K(CR - KI)

Dagley and Hinshelwood

Holzberg and co-workers

CC

% = k
CC

c=kCC

I--

CA
exp( -KCR )
CM + cA

rc = k - - CA
Cc

c
cc

CM

= k(1

KI

+ CA Kl + CR

-3T5L

Ghose and Tyagi

Aiba and co-workers
Jerusalimsky and Neronova
Bazua and Wilke

CM + CA

Levenspiel

HAN AND LEVENSPIEL: EXTENDED MONOD KINETICS

435

,Wayman and Tseng's modelg

0'
0

0.2

\
\

I .o

0.8

0.4
0.6
n-butanol (vol %)

Figure.U. Final fit to Kortan's data*on n-butanol substrate inhibition.

Table VII. Kinetic models for growth with inhibitory substrates; from ref. 7.
Reference

Equation

Haldane (function 1)
Webb (function 2)
Yano and co-workers (function 3)
Aiba and co-workers (function 4)

rc = k[sxp(-?)

- erp(-z)]Cc

Teissier-type (function 5)

Table VIII. Values of the constants in Table VII for data of ref. 6;
taken from the Table V of ref. 7.

k
Function

0-l)

1
2
3
4
5

0.4113
1.550
0.2529
0.1905
0.1320

CM
(mmHg)

K/
(mmHg)

K
(mmHg)

47.36
267.4
34.88
20.86

47.36
4.274
98.91
194.4
268.2

205.0
307.4

15.51

Product Inhibition

Cell Inhibition
Cells also can be regarded as a reaction product, so the
equations and methods developed for product inhibition kinetics should be applicable to cell inhibition. Unfortunately,
we have not been able to test this supposition, because we
were unable to find any data in the literature suitable for
testing.
Finally, if a number of inhibitory factors act at the same
time, for example substrate inhibition and product inhibition, we may expect to represent this situation with the
generalized Monod equation as follows

Figure 5 compares the prediction of various models with

the data on classical alcohol inhibition as reported by Bazua
and Wilke.3 It can be seen that Levenspiel's e q ~ a t i o n ,a~
special case of the model proposed here, fits the data best.

436

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 32,AUGUST 1988

0.10
present model predicts
this limiting point

0.08
0.06

0.04
0.02
0

200

400

600

700

partial pressure of n-pentane (mm Hg)

Figure 13. Fit of various proposed substrate inhibition equations of
Table VII to the data of ref. 6 . Note that data at high pentane concentrations would sharply discriminate between models.

References

NOMENCLATURE
CA
CC

substrate concentration (substrate/volume)

cell concentration (cell/volume)
c,
inhibitor concentration (inhibitor/volume)
chf
Monod constant
Chf,
observed value of C,
Clf
product concentration (product/volume)
critical concentration above which reaction cannot proC*
ceed; C: for inhibitor, C : for cell, C i for substrate,
C,* for product
iandj
positive integers
k
reaction rate constant (time-')
k*s
observed value of k
K , K,, and Ks constants in Table VI and Table VII
nandm
constants
rC
rate of cell formation (cell mass/volume . time)
This work was carried out under NSF Grant no.: CBT-851.8737.

1. J. Monod, Ann. Rev. Microbiol., 3, 371 (1949).

2. C. N. Hinshelwood, The Chemical Kinetics of the Bacterial Cell
(Clarendon, Oxford, 1952).
3. C. D. Bazua and C. R. Wilke, Biotechnol. Bioeng. Symp., 7 , 105
(1977).
4. 0. Levenspiel, Biotechnol. Bioeng., 22, 1671 (1980).
5. S. Aiba, M. Shoda, and M. Nagatani, Biotechnol. Bioeng., 10, 845
(1968).
6. N. Uemura, J. Takahashi, and K. Ueda, J . Ferment. Technol., 47,
220 (1969).
7 . V. H. Edwards, Biotechnol. Bioeng., 12, 679 (1970).
8. K. Kortan, M. S . thesis, University of Toronto, Ontario, Canada,
1972.
9. M. Wayman and M. C. Tseng, Biotechnol. Bioeng., 18, 383 (1976).
10. A . Moser, in Biotechnology, Volume 2, H. Brauer, Ed. (Verlag
Chemie, Weinheim, 1985).

HAN AND LEVENSPIEL: EXTENDED MONOD KINETICS

437