Simplified Modeling of Fed-Batch

Alcoholic Fermentation of Sugarcane

Blackstrap Molasses
Attilio Converti,1 Saleh Arni,1 Sunao Sato,2
Joao Carlos Monteiro de Carvalho,2 Eugenio Aquarone2
1

Department of Chemical and Process Engineering G.B. Bonino,

University of Genoa, via Opera Pia 15, 16145 Genoa, Italy; telephone: +39
010-3532593; fax: +39 010-3532586; e-mail: converti@unige.it
2
Faculdade de Ciencias Farmaceuticas, Universidade de Sao Paulo,
Avenida Prof. Lineu Prestes 580, 05508-900 Sao Paulo, SP, Brazil
Received 20 December 2002; accepted 29 April 2003
Published online 18 June 2003 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/bit.10750

Abstract: Simplified modeling based on material balances for biomass, ethanol and substrate was used to
describe the kinetics of fed-batch alcohol fermentation of
sugarcane blackstrap molasses. Maintenance requirements were previously shown to be of particular significance in this system, owing to the use of massive inoculum to minimize inhibitions; therefore, they were taken
into consideration for kinetic modeling. Average values
of biomass and ethanol yields, productivities, and substrate consumption rates, calculated at the end of runs
performed either at constant or exponentially varying
flow rates, demonstrated that all of these parameters
were influenced by the initial sugar-feeding rate, FoSo.
Under conditions of substrate shortage (FoSo 300 gS
h1), the amount of carbon dioxide produced was higher
than that corresponding to the stoichiometry of sucrose
fermentation to ethanol, indicating that an appreciable
fraction of the carbon source was likely consumed by
respiration. Besides, the biomass yields either on substrate, YX/S, or ethanol, YX/E, as well as the product yield
on substrate, YE/S, notably decreased. These results are
in agreement with the relatively high specific rate of anaerobic substrate consumption for maintenance estimated for this system (mas = 0.789 gS gX1 h1), which
was responsible for the consumption of more than 70%
of the fed carbon source. The proposed equations derived from the Monod model proved to be a useful tool to
easily predict the performance of this process. 2003
Wiley Periodicals, Inc. Biotechnol Bioeng 84:8895, 2003

Keywords: maintenance requirements; fed-batch process; alcoholic fermentation; Saccharomyces cerevisiae;

sugarcane blackstrap molasses

INTRODUCTION
Ethanol fermentation of various raw materials and agroindustrial residues was extensively studied either in batch or
continuous cultures. Among the various renewable waste
materials used for fuel ethanol production, sugarcane blackstrap molasses is particularly suitable for that purpose be-

Correspondence to: A. Converti

2003 Wiley Periodicals, Inc.

cause it is cheap and plentiful in the sugar industry (Laluce,

1991), but the presence of some inhibitors in molasses
(Converti et al., 1988) requires process optimization. Although a fed-batch process was shown to be of crucial importance in industrial practice several decades ago, relatively few empiric efforts were made in the past to model it,
mainly because of the difficulties related to the continuous
variation of the reaction volume. Most of the experimental
work was focused on ethanol production, either under constant (Borzani, 1987; Koshimizu et al., 1984) or variable
feeding conditions (Aquarone et al., 1986; Carvalho et al.,
1990, 1993; Echegaray et al., 2000; Krauter et al., 1987).
Fed-batch fermentation with cell recycling is by far the
most employed process in fuel ethanol producing countries
like Brazil, because the high biomass levels ensured by this
mode of operation allow, at the same time, reduction of the
deleterious effects of substrate and/or product inhibitions
and minimization of the costs associated to the inoculum
preparation step. In particular, linearly or exponentially decreasing flow rates allowed ethanol productivity to be increased by no less than 10% (Carvalho et al., 1990; Krauter
et al., 1987). The use of high inoculum levels was also
proposed for xylose-to-xylitol bioconversion to face the
presence of inhibitors released during acid hydrolysis of
lignocellulosics (Felipe et al., 1997; Parajo et al., 1996).
Preliminary results of fed-batch fermentations of blackstrap molasses, performed using exponentially decreasing
flow rate, were previously used to empirically relate growth
yield and cell productivity to the starting carbon sourcefeeding rate (FoSo) (Carvalho et al., 1990, 1993). Maintenance requirements were likely to be significant in those
works because large fractions of the carbon source had to be
consumed to maintain biomass, which was used at high
concentration either to accelerate the process or to overcome the above inhibition problems. However, no systematic kinetic study was performed up to now on this topic.
Additional tests were then performed in this work, at rela-

tively low FoSo values and using exponentially increasing

flow rates, in order to check the general validity of the
previous approach and to extend it to ethanol formation and
substrate consumption, under a wider range of experimental
conditions. Simplified kinetics based on experimental mass
balances between the initial and final states of fermentations
rather than on concentrations was used to avoid considering
any influence of volume variation.
A similar model was adopted with success for an automated fed-batch culture of Kluyveromyces fragilis (Nor et
al., 2001). However, most of the attempts made to rigorously model fed-batch processes (Lee et al., 1999) were
complex and often required numerical solutions, including
efforts based on differential algebraic system models (Enfors, 2001), deterministic or probabilistic models (Saucedo
and Karim, 1998), Kalman filters (Zoretto and Wilson,
1996), neural networks (Costa et al., 1998), and fuzzy logic
(Besli et al., 1995). The present simplified modeling, aimed
at predicting and controlling the performance of fed-batch
industrial fermentation of sugarcane blackstrap molasses,
was used to study the effects of maintenance requirements
on the yields and kinetic parameters. Although already evidenced in a few studies using this carbon source (Carvalho
et al., 1990, 1993), these effects were never discussed in
detail before.

MATERIALS AND METHODS

Homogeneous yeast suspensions with concentration of 52
2 g L1 (dry weight) were prepared by suspending 550 g of
wet-pressed commercial yeast (Saccharomyces cerevisiae)
in 3.0 L of distilled water by means of a 6-flat-blade turbine
with diameter 7.5 cm. The mash, prepared by diluting blackstrap molasses in distilled water, was firstly clarified to
remove suspended solids (Vitolo et al., 1985) and then
supplemented with penicillin V (0.3 mg L1) to prevent
contamination. After addition of 0.5 g L1 urea to ensure the
desired nitrogen content, the pH was adjusted to 4.55.0 by
means of concentrated H2SO4. The concentration of total
reducing sugars after these operations was 221 12 g L1
(expressed as glucose). No adjustment of pH was necessary
because it varied little during every test, likely due to the
presence of buffering substances in molasses (Converti et
al., 1988).
Fed-batch runs were carried out in a 14-L New Brunswick fermentor, without any air supply. Three liters of the
above cell suspension was added into the empty fermentor,
and the impeller speed and temperature were adjusted to
200 min1 and 32C. The mash was fed at selected feeding
rates up to a final volume of 10 L. Experiments were carried
out in 5 replicates at each selected value of FoSo, either at
constant or exponentially varying flow rates. It was previously demonstrated that ethanol productivity was not affected by periodic feeding rather than continuous feeding
(Borzani et al., 1988); therefore, the selected volume of
mash was added at time intervals of 15 min. Each fed-batch

run was followed by a batch step during which sugar was

nearly completely consumed.
Sugars and ethanol concentrations were measured as previously described (Carvalho et al., 1990). Cell concentration
was measured by the dry weight method, filtering 5-mL
samples through Millipore membranes with 1.2-m pore
diameter, washing with 50-mL of distilled water, and drying
at 105C for 4 h. During tests at FoSo < 300 gS h1, the exit
gas from the reactor was dried (Jansen et al., 1984) and
absorbed in Ba(OH)2 solutions of suitable concentration.
Carbon dioxide evolution was determined titrimetrically
(ISO, 1997), taking into account both the headspace volume
and the average CO2 solubility in water at 32C and pH
4.55.0 (Perry et al., 1998).
THEORETICAL ASPECTS
Definitions of Flux Parameters
Because of the progressive change in the volume (V), the
flow rate (F) of a fed-batch process can be defined as:
dV
= FoeKt,
dt

F=

(1)

where Fo is the starting flow rate, t the fermentation time,

and K a time parameter on which the flow rate pattern
during fermentation depends. For K 0, <0, and >0, as
those investigated in this study, corresponded to constant
flow rate and exponentially decreasing and increasing flow
rates, respectively.
Integration of Eq. (1) allows for calculation of Fo for each
K value:
Fo =

KVf Vo
,
eKT 1

(2)

where Vo is the starting volume and Vf the final volume after

the fermentor filling-up time (T).
The volume of fermentation broth after a given time can
be obtained by integrating Eq. (1) between t 0 and t and
substituting into Eq. (2):
V = Vo +

Vf Vo
eKT 1

eKt 1.

(3)

Definition and Calculation of Kinetic Parameters,

Yields, and Consumed Carbon Fractions
Modeling of fed-batch process should consider the continuous variation of all kinetic parameters during fermentation
due to volume variation, mass balance equations for biomass, substrate, and dissolved oxygen (Enfors, 2001), and
mass transfer (Di Serio et al., 2000). However, as maintenance requirements are usually more significant in fedbatch cultures than in batch or continuous ones (Heijnen,
2001), the resulting system of differential equations does
not allow analytical solutions, especially when the flow rate

CONVERTI ET AL.: MODELING FED-BATCH FERMENTATION OF BLACKSTRAP MOLASSES

89

continuously varies. For this reason and to provide a set of

equations easily and rapidly applicable to predict the behavior of industrial fed-batch fermentations, we used average
values of the main kinetic parameters and yields between
the starting and final states of each run. Such a simplified
approach was utilized with success in studies dealing with
the influence of operating conditions on kinetics and yields
of different fermentations, during which they varied along
the time as well (Carvalho et al., 1990, 1993; Converti and
Domnguez, 2001; Mussatto and Roberto, 2001; Parajo et
al., 1996). Although these values differ from the instantaneous ones, the resulting average kinetic analysis can provide a clear, even if rough, picture of the effects of operating
conditions on fermentation performances.
Previous work (Carvalho et al., 1990, 1993) revealed that
the average productivities of biomass, rX, and ethanol, rE,
were Monod-type functions of the starting substrate-feeding
rate (FoSo):
rX =

XfVf XoVo rXmaxFoSo

=
,
Vftf
kX + FoSo

Ef rEmaxFoSo
rE = =
,
tf kE + FoSo

(4)

(5)

where Xo and So are the starting concentration of biomass

and the substrate concentration in the feeding mash, Xf and
Ef the final concentrations of biomass and ethanol, rXmax
and rEmax the maximum values of rX and rE, and kX and kE
the corresponding saturation constants, respectively.
Since the fed-batch runs lasted a total time longer than the
filling-up time (tf > T) to complete substrate consumption, it
was Sf 0. When both growth and product formation linearly depend on substrate consumption in the absence of
any external electron acceptor, as is the case of alcoholic
fermentation, two contributions can be distinguished in the
substrate consumption rate, the one associated to growth
and product formation and the other to biomass endogenous
turnover (Roels, 1983):
rS =

SoVf Vo
rX
rXmaxFoSo
= a + mas X = a
+ mas X,
Vf tf
Y XS
Y XS kX + FoSo
(6)

where YaX/S is the true anaerobic growth yield, mas the

specific rate of anaerobic substrate consumption for maintenance processes, and X the cell concentration at the instant t.
Correspondingly, the yields of biomass on substrate (YX/S),
ethanol on substrate (YE/S) and biomass on produced ethanol
(YX/E) were calculated as the ratios of their respective rates:

90

YXE =

rX XfVf XoVo
=
.
rE
EfVf

(9)

Dividing rS (Eq. 6) by rX (Eq. 4) and substituting the definition of the macroscopic yield factor, YX/S (Eq. 7), we
obtained the equation proposed by Heijnen (2001) to estimate mas, which is valid for any system, independently of
the operation mode:
1
YXS

1
YaXS

maS
,

(10)

where is the average specific growth rate. The values of

mas and YaX/S were estimated from the slope and intercept of
the straight line obtained plotting the experimental data of
1/YX/S versus 1/.
The average specific growth rate was calculated integrating the material balance equation for biomass applied to a
fed-batch culture with no exit flow:
=

1 XfVf maxFoSo
ln
=
,
tf XoVo kX + FoSo

(11)

where max is the maximum theoretical value of .

The average specific ethanol productivity () was calculated as the ratio of to YX/E (Roels, 1983):
v=

vmaxFoSo
=
,
YXE kE + FoSo

(12)

where max is the maximum theoretical value of .

The experimental fraction of the carbon source consumed
by the fermentative metabolism (including growth) was
given by the ratio of the experimental data of YX/S collected
at variable FoSo to YaX/S. Conversely, the experimental fraction addressed to maintenance processes was obtained by
multiplying the ratio of mas to the experimental data of by
YX/S collected at variable FoSo. The fraction used for respiration (for FoSo < 300 gS h1) was experimentally determined at the end of each run from the excess CO2 production with respect to the fermentation stoichiometry. The
addition of these experimental fractions was finally used to
check the quality of the underlying data. On the other hand,
the theoretical values, calculated using the values of max
and kX estimated by nonlinear regression (see later), were
utilized to predict the theoretical behavior of YX/S by means
of Eq. (10). The theoretical fraction of the carbon source
used for the growth was then calculated as the ratio of the
theoretical values of YX/S to YaX/S. Finally, the theoretical
fraction used for cell maintenance was calculated as the
difference necessary to close the theoretical material balance.
RESULTS AND DISCUSSION

rX XfVf XoVo
,
YXS = =
rS SoVf Vo

(7)

rE
EfVf
YES = =
,
rS SoVf Vo

(8)

Flow Rate Patterns

Fed-batch fermentation of blackstrap molasses was studied
either at constant (K 0) or exponentially varying feeding

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 84, NO. 1, OCTOBER 5, 2003

rates (K 0). To select the ranges of flux variables significant for the process (T, t, and K), a theoretical study was
performed on their influence on the flow rate patterns.
These variables were parametrically varied keeping constant one of them, alternately.
Figure 1a shows the simultaneous dependence of the flow
rate on t and K, for a given fermentor filling-up time (T
5 h). The starting flow rate (Fo) is a decreasing function of
K (Eq. 2), while the flow rate (F) decreases with time for K

Figure 1.

< 0 or increases for K > 0 (Eq. 1). Besides, it is a decreasing

function of K for 0 < t < 1 h, whereas it follows an opposite
behavior for t 3 h. The most interesting effects on the
flow rate can be observed in the range 0.8 < K < 0.4 h1,
which was consequently selected to perform the subsequent
experimental study.
On the other hand, for K < 0, the flow rate decreases with
increasing either the fermentation time (t) or the fermentor
filling-up time (T), whereas it follows an opposite trend for

Dependence of the flow rate of a fed-batch process on the fermentation time, t, the fermentor filling-up time, T, and the time parameter, K.

CONVERTI ET AL.: MODELING FED-BATCH FERMENTATION OF BLACKSTRAP MOLASSES

91

K > 0 (Fig. 1b). An intermediate value of K (0.4 h1),

among those selected for tests, was used for comparison.
These parametric variations pointed out that T 5 h would
be the most interesting flux conditions to investigate experimentally the influence of the fermentor filling-up time on
fed-batch fermentation performance. However, the value T
10 h cannot be considered to be of industrial interest,
because it would be a fermentation time that is too long in
terms of ethanol productivity.
Finally, Fig. 1c shows that, after a given fermentation
time (t 2 h), the flow rate increases nearly linearly with
K at very short fermentor filling-up times (T 3 h),
whereas, for T 4 h, it grows with K up to a maximum
value depending on T and subsequently quickly decreases.
But the most important finding is that, at every T value, F
practically converges to the same value when K 1.0 h1,
which means that such a parameter should hardly influence
the performance of a fed-batch process with quick exponential flow rate decrease. These results suggested that the most
interesting flow rate variations should take place in the
ranges: 0.8 < K < 0.4 h1 and 5 T 7 h.
Kinetic Parameters and
Maintenance Requirements
Fed-batch fermentations of sugarcane blackstrap molasses
were performed at low starting substrate-feeding rates (25 <
FoSo < 150 gS h1) to integrate previous results as well as to
collect a wide range of data necessary to check earlier proposed models (Carvalho et al., 1990, 1993). The experimental data of all these fermentations are summarized in
Table I.
Empiric Monod-type relations were shown to describe
well the dependence of biomass yield and productivity on
FoSo (Carvalho et al., 1990, 1993). A large number of ad-

ditional equations, with or without physical meaning, were

tested (results not shown), but only those of the Monodtype, among the biotechnological models available in the
literature, appeared to ensure satisfactory determination coefficients. Well-accepted models, considering or ignoring
substrate and product inhibitions (Roels, 1983), were shown
to fit the experimental data poorly.
Figure 2a shows that volumetric biomass and ethanol
productivities (rX and rE) depended on FoSo according to
Eqs. (4) and (5). Similar behaviors were obtained for the
corresponding specific rates ( and ), described by Eqs.
(11) and (12), respectively. A standard nonlinear regression
program (Table Curve for Windows), able to minimize the
deviations between model predictions and experimental
data, was used to estimate the kinetic parameters appearing
in those relations (rXmax, rEmax, max, and max) as well as
the corresponding saturation constants (kX and kE). As expected by the direct relation between specific and volumetric rates, the values of kX estimated by Eqs. (4) and (11) as
well as those of kE estimated by Eqs. (5) and (12) were close
(Table II).
Substrate appeared in the same figure to be effectively
consumed even at very low FoSo values, which suggests that
maintenance processes could have become predominant under these conditions. This behavior is consistent with Eq.
(6), which was proposed to describe the dependence of rS on
FoSo. However, because of the continuous variation of X
with time, it was not possible to estimate the kinetic parameters appearing in that equation from the experimental values of rS; therefore, the approach reported by Heijnen
(2001) was followed for maS estimation. Using literature
data from different bioprocesses, this author demonstrated
the general validity of Eq. (10) and proposed that the dependence of YX/S on could be due to the expenditure of
energy required to maintain cellular functions: while in

Table I. Experimental results of fed-batch fermentations of sugarcane blackstrap molasses (Xo

52 2 g L1)
Test
no.

T
(h)

1
2
3
4
5
6
7
8
9

5.0
5.0
5.0
5.0
5.0
7.0
7.0
7.0
7.0

10
11
12
13

8.0
8.0
7.0
7.0

tf
(h)

K
(h1)

Soa
(gS L1)

FoSoa
(gS h1)

Results from previous work (Carvalho et al., 1990,

7.0
0
221 14
310 19
6.57.0
0.2
217 10
480 24
6.5
0.4
222 11
716 38
6.06.5
0.6
218 12
962 50
6.07.0
0.8
211 2
1202 11
7.58.0
0
227 10
228 11
7.58.0
0.2
224 15
416 29
7.0
0.4
224 15
668 45
7.0
0.6
227 12
964 54
9.0
8.5
8.0
7.5

Results
+0.4
+0.3
+0.2
+0.1

obtained in this work

212 4
25 2
224 8
50 2
221 7
99 6
223 11
150 10

Xf a
(gX L1)

Efa
(gE L1)

1993)
24.7 1.0
24.6 0.8
25.5 1.6
26.6 0.6
26.4 0.8
24.8 0.7
25.1 0.7
26.0 0.6
26.8 0.6

64.7 3.1
63.4 3.7
64.0 3.0
64.4 1.9
63.4 0.7
67.2 1.6
66.6 2.2
66.9 1.4
67.1 1.9

17.7 0.2
19.4 0.5
21.4 0.7
22.0 0.6

29.6 1.1
42.3 1.7
53.4 1.6
58.6 2.1

a
Reported values on the left represent the mean of 5 replicates, while those on the right, the related
standard deviations.

92

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 84, NO. 1, OCTOBER 5, 2003

Figure 2. Fed-batch fermentations of sugarcane blackstrap molasses. Influence of the starting substrate-feeding rate, FoSo, (a) on the volumetric rates of
() substrate consumption, rS; () ethanol formation, rE; and () biomass growth, rX; and (b) on the yields of () ethanol on substrate, YE/S; () biomass
on produced ethanol, YX/E; and () biomass on substrate, YX/S. Continuous curves refer to values estimated by Eqs. (49).

batch cultures this contribution is negligible due to high

values, maintenance may generally be more important in
fed-batch processes and only becomes really significant
when the apparent specific growth rate (ap) is much lower
than max.
The values of mas (0.789 gS gX1 h1) and YaX/S (0.219 gX
1
gS ) were estimated with reasonable accordance (r2
0.892) from the slope and intercept of the straight line obtained plotting the experimental data of 1/YX/S versus 1/.
However, a range of practical significance should be used
in Eq. (10) to estimate a realistic value of YaX/S. In this
connection, the experimental values at variable FoSo were
fitted by nonlinear regression using Eq. (11) and max
0.093 h1 was estimated. Substituting this value into Eq. (6),
we calculated YaX/S 0.077 gX gS1 (corresponding to
0.089 C-molX C-molS1), which compares with the values

of the true growth yield reported for glucose fermentation

by S. cerevisiae (0.120.171 C-molX C-molS1) and that
(0.10 C-molX C-molS1) estimated from data of standard
combustion enthalpies and reduction degrees of yeast biomass and sucrose (Roels, 1983). Besides, as expected, it was
about 7 times lower than that reported by the same author
for the aerobic growth of S. cerevisiae on glucose (0.59
0.65 C-molX C-molS1).
The above mas value, corresponding to 0.673 C-molS CmolX1 h1, was about 25% less and 45 times higher than
the highest and lowest values reported in the literature for
the anaerobic growth of S. cerevisiae on glucose (0.150.9
C-molS C-molX1 h1) and more than one order of magnitude higher than that reported for the aerobic growth (0.012
C-molS C-molX1 h1) (Roels, 1983). These results on the
whole are in agreement with the well-known higher main-

CONVERTI ET AL.: MODELING FED-BATCH FERMENTATION OF BLACKSTRAP MOLASSES

93

Table II. Main kinetic parameters and yields of fed-batch fermentations of sugarcane blackstrap
molasses estimated by Eqs. (412)

rXmax 1.83 gX L1 h1
rEmax 10.5 gE L1 h1
rSmax 24.0 gS L1 h1

Kinetic parameters
max 0.093 h1
kX 148 gS h1a kX 139 gS h1b
max 0.526 gE gX1 h1
kE 49.1 gS h1c kEd 45.3 gS h1d
mas 0.789 gS gX1 h1

Yields
YX/Smax 0.077 gX gS1 YE/Smax 0.437 gE gS1
YX/Emin 0.057 gX gE1
YaX/S 0.219 gX gS1

YX/Emax 0.174 gX gE1

Value estimated by Eq. (4).

Value estimated by Eq. (11).
c
Value estimated by Eq. (5).
d
Value estimated by Eq. (12).
b

tenance requirements and lower growth yield of the anaerobic metabolism with respect to the aerobic one (Heijnen,
2001).
Comparing in Table II the values of the saturation constants estimated by non-linear regression using Eqs. (4) and
(5), kX was shown to be much higher than kE, which suggests that, at very low FoSo values, maintenance requirements could have become so important that the cells, owing
to energy limitation, gave up growing almost completely.
Conversely, the yeast went on fermenting and breathing to
produce energy. Such a hypothesis appears to be consistent
with the behaviors of the experimental fractions of substrate
consumed by the fermentative metabolism (including
growth) and the maintenance processes (Fig. 3). Although
the latter experimental fraction decreased with increasing
substrate availability, it kept always higher than the former.
This result suggests that the maintenance requirements predominate in a fed-batch culture with massive inoculum and
supports the assumption that the sensitivity of biomass yield
to maintenance aspects would be due to low ap values
(Heijnen, 2001).

If the hypothesis that only fermentative metabolism (including growth) and cell maintenance are relevant in the
selected system were correct, then the addition of the theoretical fractions of the carbon source addressed to these two
activities should always be 100% (Fig. 3). The horizontal
straight line corresponding to the theoretical carbon balance
fits reasonably well the addition of the experimental fractions only for FoSo 300 gS h1 (6.0% standard deviation).
On the contrary, some losses were evident from the theoretical carbon balance for FoSo < 300 gS h1 considering
only these two activities. To shed light on this phenomenon,
the total amount of CO2 produced was experimentally determined at the end of each test, as described in Materials
and Methods. This production was higher than that corresponding to the stoichiometry of sucrose fermentation to
ethanol and succeeded in closing well the carbon material
balance (6.0% overall standard deviation); therefore, it was
ascribed to possible respiration. In fact, since no air was
supplied during fermentations, the starting oxygen level
present in the reactor (provided by the headspace) was always very low. At high FoSo values, it was insufficient to

Figure 3. Influence of the starting substrate-feeding rate, FoSo, on the experimental fractions of substrate consumed for () fermentative metabolism
including growth; () maintenance processes; () respiration; and () their addition. Curves refer to the theoretical fractions of the carbon source utilized
by the yeast only for growth and maintenance, while the straight line represents their addition.

94

BIOTECHNOLOGY AND BIOENGINEERING, VOL. 84, NO. 1, OCTOBER 5, 2003

sustain appreciable respiration, and the fraction of carbon

source consumed by this activity was negligible. As this
fraction grew progressively with decreasing FoSo, a threshold value was reached (FoSo 300 gS h1) below which it
became significant.
The above effects of maintenance requirements on the
kinetics are confirmed by the yield dependence on FoSo
(Fig. 2b). According to Eqs. (49), the yield of ethanol
(YE/S) and biomass (YX/S) on consumed substrate followed
typical Monod-type equations, while that of biomass on
produced ethanol (YX/E) shifted from a minimum toward a
maximum value. If compared with the constant values observed in continuous cultures, these trends are the result of
maintenance processes significance in substrate consumption during fed-batch processes, especially under conditions
of substrate shortage and high biomass level. It is easy to
verify from Eqs. (49) that, for FoSo 0, both YX/S and YE/S
tended to zero, while YX/E approached YX/Emin 0.057 gX
gE1. The achievement of such a minimum value suggests
that biomass growth decelerated under these conditions
more than ethanol formation, which, together with some
respiration, kept the sole activity able to sustain the energy
requirements for maintenance processes. On the other hand,
increasing FoSo, all three yields tended to maximum values
(YX/Emax 0.174 gX gE1; YE/Smax 0.437 gE gS1;
YX/Smax 0.077 gX gS1).
The simplified modeling presented in this work proved to
be a useful and simple tool to predict and control the performance of fed-batch fermentation of sugarcane blackstrap
molasses, but further work is needed to find the optimum
starting biomass level able to minimize at the same time
inhibition and energy requirements of the system.
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