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Q IWA Publishing 2009 Water Science & Technology—WST | 60.1 | 2009
9
Influence of temperature on the hydrolysis, acidogenesis
and methanogenesis in mesophilic anaerobic digestion:
parameter identification and modeling application
A. Donoso-Bravo, C. Retamal, M. Carballa, G. Ruiz-Filippi and R. Chamy
ABSTRACT
The effect of temperature on the kinetic parameters involved in the main reactions of the
A. Donoso-Bravo (corresponding author)
C. Retamal
M. Carballa
G. Ruiz-Filippi
R. Chamy
School of Biochemical Engineering,
Pontificia Universidad Católica de Valparaı́so,
General Cruz 34,
Valparaı́so,
Chile
E-mail: andres.donoso.b@mail.ucv.cl
anaerobic digestion process was studied. Batch tests with starch, glucose and acetic acid as
substrates for hydrolysis, acidogenesis and methanogenesis, respectively, were performed in a
temperature range between 15 and 458C. First order kinetics was assumed to determine the
hydrolysis rate constant, while Monod and Haldane kinetics were considered for acidogenesis
and methanogenesis, respectively. The results obtained showed that the anaerobic process is
strongly influenced by temperature, with acidogenesis exerting the highest effect. The Cardinal
Temperature Model 1 with an inflection point (CTM1) fitted properly the experimental data in the
whole temperature range, except for the maximum degradation rate of acidogenesis. A simple
case-study assessing the effect of temperature on an anaerobic CSTR performance indicated that
with relatively simple substrates, like starch, the limiting reaction would change depending on
M. Carballa
Department of Chemical Engineering,
School of Engineering,
University of Santiago de Compostela,
Rúa Lope Gómez de Marzoa s/n,
15782 Santiago de Compostela
Spain
temperature. However, when more complex substrates are used (e.g. sewage sludge), the
hydrolysis might become more quickly into the limiting step.
Key words
| acidogenesis, anaerobic digestion, hydrolysis, modeling, temperature
INTRODUCTION
Nowadays, anaerobic digestion might be considered as a
the methanization process. On the contrary, when treating
consolidated technology with more than 2200 high-rate
complex organic matter or anaerobic treatment is carried
reactors implemented worldwide, treating different types of
out at low temperature, hydrolysis is often considered as the
wastes and wastewaters coming from different sectors, such
limiting step (Vavilin et al. 2008).
as agro-food industry, beverage, alcohol distillery and pulp
and paper industries (van Lier 2008).
The temperature at which the anaerobic digestion
occurs can significantly affect the conversion, kinetics,
However, there are a number of factors which affect
stability, effluent quality, and consequently, the methane
the anaerobic digestion process, including the substrate
yield of the process (Sanchez et al. 2001). Microorganisms
characteristics, the reactor configuration, the operational
are generally divided into three thermal groups, psycro-
parameters, such as hydraulic retention time (HRT), solids
philes, mesophiles and thermophiles, with optimum
retention time (SRT) and organic loading rate (OLR),
temperatures below 208C, 25 –408C and higher than 458C,
and the environmental factors like temperature and pH
respectively (van Lier et al. 1997), and it has been
(Banerjee et al. 1998).
demonstrated that the anaerobic degradation rate of
As a complex multi-step process, the overall kinetics of
organic matter increases with temperature when psychro-
waste utilization during anaerobic treatment is governed by
philic,
the kinetics of the slowest step, which often corresponds to
compared (Sanchez et al. 2001). However, anaerobic
doi: 10.2166/wst.2009.316
mesophilic
and
thermophilic
conditions
are
10
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
Water Science & Technology—WST | 60.1 | 2009
digestion has been traditionally performed in mesophilic
extract were added according to Field et al. (1988). Starch
range (35 – 378C) despite the temperature of certain waste-
(1.0 –3.0 g/L),
waters might be either warmer or cooler. Treating these
(0.1 –25.0 g/L) were used for hydrolysis, acidogenesis and
glucose
(0.1 –1.0 g/L)
and
acetic
acid
wastewaters at their natural temperatures would often be
methanogenesis assays, respectively. Several temperatures
beneficial because of the reduced resources and costs
were tested for the different stages, i.e. hydrolysis (12, 22,
(Kettunen & Rintala 1997).
30, 37 and 458C), acidogenesis (12, 22, 30, 37, 42 and 458C)
The influence of temperature has been extensively
and methanogenesis (12, 25, 37 and 458C). Sodium
studied on the rate-limiting methanogenic phase at
bicarbonate was used as buffer at a concentration of
both mesophilic and thermophilic conditions (Hegde &
1 g/gCODadded (Soto et al. 1993) in the hydrolysis assays,
Pullammanappallil 2007). However, little attention has been
while
paid on the effect of temperature on hydrolysis and
phosphate was used at concentrations ranging from
acidogenesis. Hydrolysis rate constants are highly depen-
0.007 – 0.070 M and 0.009 M, respectively (Retamal 2008).
for
acidogenesis
and
methanogenesis
assays,
dent on temperature since hydrolysis is a biochemical
The temperature of the experiments was maintained by
reaction catalyzed by enzymes, which are very thermally
using a thermostatic water bath. The bottles were inocu-
sensitive (Sanders et al. 2000). Moreover, a better under-
lated with anaerobic biomass coming from a mesophilic
standing of temperature effects on acidogenesis can result in
sewage sludge digester at a final concentration of
the improvement of digester stability due to physical
1.03 ^ 0.14 gVSS/L. Different substrate concentrations
separation of phases, the increase in the concentration of
were tested in duplicate or triplicate at each temperature
soluble organics and the optimization of biological nutrient
and the substrate consumption over time, pH and sus-
removal processes (Banerjee et al. 1998). In addition, few
pended solids concentration at the end of the experiments
studies have been carried out on the effect of temperature
were the parameters monitored.
on the overall process of anaerobic digestion. Banik et al.
For acidogenesis and methanogenesis, a biomass lineal
(1998) studied the effect of temperature on the overall
range study was performed at 378C with glucose (0.09 g/L)
kinetic parameters of the anaerobic biomass but just up to
and acetic acid (0.12 g/L), respectively. For acidogenesis,
258C. Since temperature variations may not have the same
three biomass concentrations, 0.20, 0.54 and 1.07 g VSS/L,
effect on the different stages of anaerobic digestion
were used and the volumetric consumption rates of glucose
(hydrolysis,
were
acidogenesis
and
methanogenesis),
more
calculated.
The
glucose
consumption
rate
at
knowledge is required to identify and select corrective
0.54 g VSS/L was double than the value obtained at
actions for temperature disturbances on anaerobic reactors.
0.20 g VSS/L, while no differences were observed between
The objective of this study was to determine the kinetic
0.54 and 1.07 g VSS/L. Hence, a biomass concentration of
parameters that characterize hydrolysis, acidogenesis and
0.54 g/L was selected. For methanogenesis, two biomass
methanogenic stages at different temperatures in order to
concentrations, 0.32 and 0.65 g VSS/L, were used and the
develop a mathematical model describing the influence of
volumetric consumption rates of acetic acid were 9.6·1023
temperature on each single stage.
and 1.9·1022 g/L·d, respectively. Thus, a biomass concentration of 0.4 g VSS/L was used.
MATERIALS AND METHODS
Determination of kinetic parameters
Experimental set-up
Kinetic equations
Batch experiments were run in glass serum bottles with
First order kinetics was considered for the hydrolysis
a total liquid volume of 200, 250 and 100 mL for
of particulate organic matter (Equation 1), while Monod-
hydrolysis,
assays,
type (Equation 2) and Haldane-type (Equation 3) kinetics
respectively. Macronutrients, micronutrients and yeast
were assumed for acidogenesis and methanogenesis
acidogenesis
and
methanogenesis
11
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
(Bernard et al. 2001).
Water Science & Technology—WST | 60.1 | 2009
Finally, a nonlinear optimization by least squares
vh ¼ 2kh ·Sh
ð1Þ
S1
v1 ¼ vmax 1
KS1 þ S
S2
v2 ¼ vmax 2
S2
KS2 þ S2 þ K21
ð2Þ
ð3Þ
procedure is applied to calculate the unknown parameters
by minimizing a cost function (Equation 6), which measures
the difference between the experimental measurements and
the corresponding simulated value (the values obtained
with the linearization method are used as initial values in
the simulation process).
where vh is the hydrolysis reaction rate (g/L·d), Sh is the
hydrolysis substrate concentration (g/L), kh is the hydroly21
sis rate constant (d
), v1 is the acidogenesis reaction rate
JðcÞ ¼ min
N
X
t¼1
2
vm ðtÞ 2 vðt; cÞ jc0 ¼cL
ð6Þ
(g/gVSS·d), S1 is the acidogenesis substrate concentration
(g/L), vmax1 is the maximum degradation rate for acidogen-
where vm is the velocity consumption obtained for
esis (g/gVSS·d), KS1 is the half saturation constant for
measurements, v is the corresponding simulated velocity
acidogenesis (g/L), v2 is the methanogenesis reaction rate
and N is the number of measurements.
(g/gVSS·d), S2 is the methanogenesis substrate concentration (g/L), vmax2 is the maximum degradation rate for
methanogenesis (g/gVSS·d), KS2 is the half saturation
Models of temperature effect
constant for methanogenesis (g/L) and KI is the inhibition
The influence of temperature on the hydrolysis constant rate
constant associated with substrate S2.
has been often described by the Arrhenius model. However,
this model is limited by a maximum value of temperature
above which the temperature effect can not be further
Parameters estimation
evaluated. In contrast, the Cardinal Temperature Model 1
The hydrolysis rate constant is obtained from Equation (1),
with an inflection point (CTM1) proposed by Rosso et al.
by representing ln S0 h/Sh versus time (slope). The determi-
(1993) is able to model all temperatures tested (Equation 7).
b ¼ bopt
ðT 2 T max ÞðT 2 T min Þ2
ðT opt 2 T min Þ ðT opt 2 T min Þ·ðT 2 T opt Þ 2 ðT opt 2 T max Þ·ðT opt þ T min 2 2TÞ
nation of the kinetic parameters for acidogenesis and
ð7Þ
where Tmin, Topt and Tmax were the minimum, optimum
methanogenesis was carried out from the initial consumption
and maximum temperatures, respectively (8C), and bopt is
rate (Retamal 2008) by using the linearization Lineweaver-
the optimum value of the kinetic parameter under study.
Burke method (Equations (4) and (5)). Since the initial
All these parameters were obtained from a nonlinear
concentration of biomass may exert an important effect
optimization by least squares procedure as described
on the biodegradation kinetics (Urra et al. 2008), these
previously. Figure 1 summarizes all the calculations
assays were performed within the biomass lineal range.
procedure.
1
KS1 1
1
¼
· þ
v1 Vm1 S1 Vm1
ð4Þ
Analytical methods
Starch concentrations were determined as the difference
1
KS2 1
1
S2
¼
þ
· þ
v2 Vm2 S2 Vm2 Vm2 ·KI
between the total sugar concentrations (Dubois et al. 1956)
ð5Þ
and the reducing sugar concentrations, i.e. glucose concentrations (Miller 1959). pH, Volatile Suspended Solids (VSS)
where cL ¼ ðKS1 ; KS2 ; KI ; Vm1 ; Vm2 Þ are the unknown
and Volatile Fatty Acids (VFA) were determined according
parameters.
to standard methods (APHA 1995).
12
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
Figure 1
|
Water Science & Technology—WST | 60.1 | 2009
Calculation procedure for the determination of the kinetic parameters and the evaluation of temperature influence. Case-study: acidogenesis. (A) Initial rates calculation;
(B) kinetic-profile construction; (C) model fit; (D) temperature influence; and, (E) temperature-model fit.
RESULTS AND DISCUSSION
to describe the effect of temperature over the whole
temperature range tested. The parameters of CTM1 are
Hydrolysis rate constants as a function
shown in Table 1. It can be observed that the optimum
of temperature
modeled temperature for hydrolysis (40.38C) is lower than
Figure 2a shows the hydrolysis rate constants of starch at
the different temperatures tested. It can be observed that the
hydrolysis rate constants increased with temperature up to
the value obtained in the experiments (378C), while the
optimum modeled hydrolysis rate constant (22.8 d21) is
higher than the experimental value (21.1 d21).
an optimum value of 21.1 ^ 2.2 d21 at 378C. However, a
The values obtained for the hydrolysis constant in this
lower value was obtained at 458C (7.9 ^ 1.7 d21), which is
study were largely greater than other values reported in
probably due to the fact that this temperature is in between
literature (Siegrist et al. 2002; Vavilin et al. 2008). This fact is
the optimum values for mesophilic and thermophilic
probably explained by the use of starch as substrate, which
conditions. In the temperature range of 12 to 378C, the
is one of the most readily hydrolysable substrates in contrast
hydrolysis rate constants fitted well the Arrhenius equation
to the more complex substrates, such as sludge or lipids,
and a value of 72 kJ/mol for the activation energy was
used in other literature studies. With regard to the
calculated, which is a typical value for enzymatic kinetics
hydrolysis constant dependency on temperature, Siegrist
under anaerobic conditions (Veeken & Hamelers 1999).
et al. (2002) also found that this parameter is strongly
On the contrary, the CTM1 model is more appropriate
influenced by temperature.
13
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
Figure 2
|
Water Science & Technology—WST | 60.1 | 2009
(a) Hydrolysis rate constants in function of temperature using starch as substrate; (b) Maximum degradation rates of acidogenesis in function of temperature using
glucose as substrate; (c) Maximum degradation rates of methanogenesis using acetic as substrate; (d) Influence of temperature on half saturation constant in
acidogenesis stage; (e) Influence of temperature on half saturation constant in methanogenic stage; (f) Influence of temperature inhibition constant in methanogenesis
stage. B, experimental data; —, CTM1 fit.
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
14
Table 1
|
Water Science & Technology—WST | 60.1 | 2009
CTM1 parameters for optimum degradation rates during hydrolysis, acidogenesis and methanogenesis using starch, glucose and acetic acid as substrates, respectively
Tmin (8C)
Parameter
Topt (8C)
Tmax (8C)
kH opt (d21)
Vm1opt (g/gVSS·d)
Vm2 opt (g/gVSS·d)
KIopt (g/L)
Hydrolysis
kh
4.2
40.3
45.5
22.8
–
–
–
Acidogenesis
Vm1
28.9
37.0
45.2
–
33.6
–
–
Methanogenesis
Vm2
11.1
34.1
46.3
–
–
0.72
–
KI
11.9
34.3
46.2
–
–
–
6.45
Influence of temperature on acidogenesis and
temperature of an anaerobic reactor, a 58C decrease would
methanogenesis
result in 50 and 10% slower kinetics of acidogenesis and
hydrolysis,
Maximum degradation rates
respectively,
with
almost
no
effect
on
methanogenesis.
Figure 2b shows the influence of temperature on the
maximum degradation rate during acidogenesis. Whereas
there was almost no difference between 12 and 308C, a
Affinity and inhibition constant
significant increase was observed at 378C with again a sharp
Figure 2b and Figure 2d show the influence of temperature
decrease above 408C. In this case, only the temperature
on glucose (acidogenesis) and acetic acid (methanogenesis)
range between 30 and 458C was considered for CTM1
affinity constants. Similar values were obtained by Siegrist
modeling, which showed that the optimum temperature for
et al. (2002), but these authors also found an increase of
acidogenesis is lower than that for hydrolysis (Table 1).
these parameters at thermophilic conditions (558C), while
Figure 2c shows the influence of temperature on the
in the present study, no significant variations were observed
maximum degradation rate during methanogenesis and the
for affinity constant of glucose in the temperature range
CTM1 fit over the whole temperature range tested. It can be
tested. This finding was also obtained by Banik et al. (1998)
observed that the maximum degradation rate during
for the global affinity constant of anaerobic population.
methanogenesis was negligible below 158C and it increased
However, the half saturation constant of acetic acid
progressively with temperature up to 30– 358C, with lower
decreased from 4.25 g/L at 258C to 1.5 g/L at 458C.
values again at higher temperatures. CTM1 model indicates
The assay carried out at 128C did not produce methane
that the optimum temperature for methanogenesis (34.18C)
during the 3 months of experiment, thus not being possible
is lower than for hydrolysis and acidogenesis (Table 1).
the calculation of this parameter at this low temperature.
Overall, the three reactions diminished their activity as
Concerning the inhibition constant of acetic acid (Figure 2f),
the temperature decreased, as also observed by Banik et al.
the influence of temperature was similar to that on the
(1998). However, the impact of temperature decrease is
maximum degradation rate (Figure 2e) and it could be
different for each stage. Assuming 378C as the operational
modeled by CTM1 (Table 1).
Figure 3
|
Profiles of the initial reaction rate at different substrates concentrations at 378C for. acidogenesis (a) and methanogesis (b).
15
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
Figure 4
|
Water Science & Technology—WST | 60.1 | 2009
Steady state COD consumption rate during (W) hydrolysis, (A) acidogenesis and (B) methanogenesis between 15 and 458C. (a) starch-wastewater (b) sludge.
Dynamic behavior
Figure 3 shows the dynamic behavior of the consumption
rate at different concentration of substrate for acidogenesis
(Figure 3a) and methanogesis (Figure 3b) at one temperature. It can be observed that the reaction behaves according
to a Monod and Haldane model.
retention time (HRT) of 20 d and six temperatures (15, 25,
30, 35, 40 and 458C) were assumed for the simulation. For
each temperature, the kinetic parameters were calculated
from the results obtained in the previous experiments
in the case of starch-wastewater. For the anaerobic
sludge treatment, a hydrolysis constant value of 0.016 d21
(which is three orders of magnitude lower than that of
starch) determined by Retamal (2008) at 378C was used,
Case-study: continuous stirred tank reactor (CSTR)
assuming a CTM1 behavior for the same range of
A two-population and three-reaction model (Donoso-Bravo
temperature. In both cases, a 90% of particulate organic
2008) adapted from the two-reaction model developed by
fraction was assumed.
Bernard et al. (2001) was used to evaluate the influence of
The results of the variables were obtained at steady state
temperature on the performance of an anaerobic CSTR.
and they were considered as the initial conditions of the
A starch-wastewater containing 5.0 g COD/L and sludge
system for the next run. The system was run in total for
from an activated sludge plant were considered. A hydraulic
500 days, and the results are shown in Figure 4.
Figure 5
|
Non-degraded COD accumulation during hydrolysis, acidogenesis and methanogenesis at different temperatures. (a) starch-wastewater (b) sludge.
16
A. Donoso-Bravo et al. | Mesophilic anaerobic digestion: parameter identification and modelling
In general terms, it can be noted that the methanogenic
reaction rate is the lowest for the temperature range studied
Water Science & Technology—WST | 60.1 | 2009
complex substrates are used (e.g. sewage sludge), the
hydrolysis was clearly the limiting step.
and for the kinetic parameters determined. However, with
sludge (Figure 4b), the hydrolysis and the methanogenesis
rates are closer than with starch, which shows that the
ACKNOWLEDGEMENTS
nature of the substrate has a significant influence on the
overall process.
In order to evaluate the limiting step at different
This work was funded by 1060220 and 1090482 projects
from Fondecyt and ALEGAS from PUCV, Chile.
temperatures, a model was used to determine the nondegraded COD in each phase (Figure 5). Figure 5a shows
that, for starch-wastewater, an important accumulation of
particulate COD would occur below 308C, whereas at
higher temperatures the methanogenic reaction would turn
into in the limiting step. For sludge (Figure 5b), hydrolysis
remains as the limiting step for all studied temperatures.
That is the reason why the increasing use of sludgepretreatment methods prior to anaerobic digestion to
diminish the particulate organic fraction of the sludge,
and thus increasing the overall rate of the anaerobic
digestion.
CONCLUSIONS
Knowledge about the effect of temperature on the kinetic
parameters of the main anaerobic reactions represents an
important progress in the anaerobic digestion field. This
work shows that the anaerobic process is strongly influenced by temperature, with acidogenesis showing the
highest effect. Therefore, it can be concluded that this
stage plays a key role in the stability of the overall anaerobic
process, since a small change in the operational temperature (around 58C), results in a decrease of 50% in the
acidogenesis reaction rate, affecting consequently the
methanogenesis stage.
Moreover, CTM1 modeled properly the influence of
temperature on the different kinetic parameters involved in
anaerobic process in the temperature range tested, except
for the maximum degradation rate of acidogenesis, which
could be only modeled between 30 and 458C. The simple
case-study to asses the effect of temperature on an
anaerobic CSTR performance indicated that with relatively
simple substrates, like starch, the limiting reaction would
change depending on temperature. However, when more
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