Energy from Biomass

Anaerobic biological processes

(Biogas plant Midlum near Bremerhaven)





Material for the lecture

Hochschule Bremerhaven, SS 2010

Prof. Dr.-Ing. Dieter Lompe





Seite 1
Content page
1 Structure of micro organisms 2
1.1 Pro- and eukaryotic cells 2
1.2 Prokaryotic cells 2
1.3 Classification of bacteria regarding their metabolism 5
2 The metabolism of bacteria 5
2.1 Basic metabolic processes 5
2.2 Respiration, fermentation, methanogenesis 8
2.2.1 The respiration 8
2.2.2 The fermentation 10
2.2.3 Acetogenesis and methanogenesis 11
3 Kinetics of growth and metabolic processes 13
3.1 Cell division and growth 13
3.2 Influence of Temperature 16
3.3 Influence of pH-value 17
3.4 Growth kinetics 19
3.4.1 Michaelis-Menten kinetic 19
3.4.2 Inhibitions 21
3.4.3 Monod-kinetic 24
4 Bioreactor design 26
5 Applications 28
5.1 General process design 28
5.2 Gas production 31
5.2.1 Gas quality 31
5.2.2 Gas yield 33
5.2.3 Reaction rates 34
5.2.4 Example: Digestion of slaughterhouse waste 39
Literature 40






Seite 2
1 Structure of micro organisms
1.1 Pro- and eukaryotic cells
Living cells can be subdivided in two types, the smaller and more simple prokaryotic cells
(e.g. bacteria) and the bigger and more complex eukaryotic cells (e.g., protozoas, fungi incl.
yeasts, plants, animals) (Fig. 1).














Fig. 1: Sketch of a pro- and eukaryotic cell (in red: Membranes) (Fritsche 1990)

1.2 Prokaryotic cells
Fig. 2 shows the inner and outer structure of a bacterial cell. In spite of very different types of
bacteria the general structure of bacteria is very similar.





Seite 3











Fig. 2: Structure of a bacterial cell (Fritsche 1990)












Fig. 3: a) basic shapes of bacterial cells, b) basic forms of flagella (Fritsche 1990)





Seite 4
The nucleus contains the genetic information coded in a conglomerate of DNA. This
macromolecule has a length of 1200 m at Escherichia coli, which means 500 times longer
than the cell.
Plasmids are small DNA molecules with additional information for individual genes.
Proteins are produced within ribosomes (app. 5000 to 50.000 per cell). Ribosomes receive
the information from DNA via mRNA, material is taken from cell plasma and energy is used
as ATP (see below).
Sometimes cells contain granula, that are stored substances like PHB (poly-hydroxo butyric
acid) or poly-phosphates.

The bacterial material contains in average the following main elements:
C 50%
O 20%
N 14%
H 8%
P 3%
S 1%
K 1%
Ca 0,5%
Fe 0,2%
These figures may vary very much depending on individual types of bacteria and on growth
conditions.




Seite 5

1.3 Classification of bacteria regarding their metabolism
Bacteria metabolism is classified into groups regarding their carbon source, their energy
source and their electron acceptor (s. fig. 4)













Fig. 4: Classification of metabolisms

If bacteria use oxygen as final electron acceptor of their metabolism they are called aerobic
or anoxic bacteria. Anaerobic bacteria use other electron acceptors.

2 The metabolism of bacteria
2.1 Basic metabolic processes
Metabolism is used to get energy from substrates and to produce new cell material for
growth. For a better understanding the metabolism is subdivided in degradation metabolism
or Catabolism, e.g. oxidation of glucose or other organic substances. From this oxidation the
CO
2
autotroph
organic Subst.
heterotroph
Radiation
phototroph
Energy-source
Organ. Subst.
chemo-
organotroph
Inorgan. Subst.
chemo-
lithotroph
Chemoorgano-
heterotroph
animals, fungi,
many bakteria
Chemoorgano-
autotroph
some methane
bakteria
Photoheterotroph
some Purpur-
bakteria a. Algae
Photoautotroph
Plants, Algae
Cyanobacteria
Chemolitho-
autotroph
nitrifying bacteria
iron oxidizing bact.
Chemolitho-
heterotroph
some methane
bakteria
Carbon-source




Seite 6
reaction enthalpy is used to produce ATP (Adenosine-tri-phosphate) from ADP (Adenosine-
di-phosphate) as an energy storage. This energy is used on the other hand to synthesize
new material for cell growth in the Anabolism.
There are many links between catabolism and anabolism, e.g. by using substances from a
partially degradation already for the anabolism, sometimes by converting these intermediate
products within the intermediate metabolism.
In order to use energy from reactions in amounts that enable the storage of energy in
molecules like ATP, e.g. from the oxidation of glucose, micro organisms use a lot of small
reaction steps which led normally to an transport of electrons. Therefore those reactions are
also named redox-reactions, because of they are a combination of reduction and oxidation.
Often these reactions are called also hydrogen-transfer-reactions, because of hydrogen is a
combination of one proton and one electron.
As an example, the very exothermic oxidation of hydrogen with oxygen can be written in
general as in (eq.2.1),
O
2
+ H
2
H
2
O (2.1)

but in detail as a transfer of electrons and hydrogen to oxygen(eq. 2.2).

O
2
+ 2H
+
+ 2e
-
H
2
O (2.2)

In this reaction, oxygen was reduced and hydrogen was oxidized.
The free energy from this reaction G
0
can be calculated by using the difference of the redox
potential R between the electron-donator (Hydrogen, -0,42V) and the electron-acceptor
(Oxygen, +0,81 V) and the Faraday-Constant F (96480 As/mol). This calculation leads to a
free energy from this reaction of G
0
= -237,4 kJ/mol (eq. 2.3).
(2.3)

For the storage and transport of energy and hydrogen mainly two substances are used by
cells: ATP and NADH respectively (s. fig.5).
F R G =
0




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Fig. 5: Structure of ATP and NADH (Fritsche 1990)

The free energy for the ATP ADP-reaction has an amount of 34 KJ/mol. If organism are
able to portion the free energy from the degradation of substrates by using many small
redox-reactions with this energy amount of 34 KJ/mol each of these reactions let cells
produce one ATP from ADP.
NADH is often used in cells to store and transport hydrogen (s. fig. 5), but there are also
other substances for this purpose (e.g. FAD).





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2.2 Respiration, fermentation, methanogenesis
Fig. 6 shows the main types of bacterial metabolism where as an example of organic
substrates, but also for cell-material a unit of [CH
2
O] is used.


Respiration



Fermentation


Metabolism of phototrophic organisms



Metab. of chemolithotrophic organisms



Methanogenesis

Fig. 6: Main types of bacterial metabolism (Fritsche 1990)

2.2.1 The respiration
In the respiration-metabolism substrates (e.g. organic substrates, e.g. glucose) are oxidized
by using oxygen. Normally molecular oxygen O
2
is used, if there is dissolved oxygen in the




Seite 9
aqueous system available which is named aerobic metabolism or metabolism under aerobic
condition. If there is no molecular oxygen available, but oxygen within substances like nitrate
or sulphate a respiration-metabolism under anoxic conditions takes place, but with a small
difference in the available free energy:
aerobic conditions:
C
6
H
12
O
6
+ 6 O
2
6 CO
2
+ 6 H
2
O, G
0
= -2870 KJ (2.4)

anoxic conditions with nitrate:
C
6
H
12
O
6
+4,8 HNO
3
6 CO
2
+ 4,8 H
2
O, G
0
= -2669 KJ (2.5)

The respirative degradation of glucose consists of mainly four steps:
1. Conversion of 1 mol glucose into 2 mol pyruvate C
3
H
4
O
3
(Yield: 2 mol ATP, 2 mol
NADH)
2. Decarboxylation (split off of carbon dioxide) of pyruvate (Yield: 1 mol NADH)
3. Further degradation within the citric-acid-cycle or Krebs-cycle (Yield: 2 mol ATP, 7
mol NADH, 2 mol FADH)
4. The respiratory chain, where all hydrogen from NADH or FADH is transferred to
oxygen (Yield: 34 mol ATP by using 6 mol oxygen)
In total, 38 mol ATP are yielded from the aerobic oxidation of 1 mol glucose. If other ways of
transferring energy within a bacteria are neglected an efficiency h of the aerobic biological
oxidation of glucose can be calculated:
(2.6)

with: n
ATP
: moles ATP per mol glucose

Using eq. 2.4 the efficiency of the aerobic oxidation of glucose results in (38*34/2870):
h= 45%,
which is more than normal power plants achieve.
0
0
Gl
ATP ATP
G
G n

=




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But the result shows also, that 55% of the input-energy is converted to heat. That leads at
aerobic processes with high reaction rates to a high temperature in the reactor.

2.2.2 The fermentation
Fermentation processes consist often of the same first three steps like the respiration
process, but due to a lack of oxygen hydrogen cannot be transferred to oxygen. Therefore
other products with a high hydrogen content were produced and released by those micro
organisms.
Depending on the intermediate products to which hydrogen is transferred, different
fermentation products are yielded. Often organic acids like acetic acid, propionic acid, butyric
acid, or lactic acid or alcohols like methanol or ethanol are produced (s. fig. 7).
As an example ethanol is produced be the yeast Saccaromyces cerevisae from glucose by
transferring hydrogen from NADH to acetaldehyde.
C
6
H
12
O
6
2 CO
2
+ 2 C
2
H
5
OH, G
0
= -197 KJ (2.7)

Some organisms are able to release molecular hydrogen. This process can be used for
biological production of hydrogen from organic waste.
Organisms which produce lactic acid are widely used for the conservation of food like
sauerkraut, sour milk, yoghurt, cucumbers, some cheese or animal food like silage.
Because of the respiratory chain cannot be used in fermentation processes the energy yield
per amount of substrate and therefore also the growth rate are much lower than in aerobic
processes.
Many fermentative bacteria and yeasts are fastidious anaerobic, but there are also
organisms which are able to adapt to anaerobic and aerobic conditions or which are tolerant
to some oxygen.






Seite 11















Fig. 7: fermentative production of alcohol (left) and lactic acid (right) (Fritsche 1990)

2.2.3 Acetogenesis and methanogenesis
The production of methane containing biogas is a complex process consisting of four main
steps where many different micro organisms are involved.
1. If solid substrates are used from organic waste or agricultural products as a first step
hydrolysis is necessary in order to transfer the material into a dissolved phase to
make them accessible to other organisms. Some micro organisms often
fermentative organisms are able to produce special enzymes and to secrete these
enzymes (eponyms). These extra cellular enzymes brake up large molecules like
polymers (e.g. starch) to smaller components. These smaller components are soluble
and can be transferred via the cell-membrane into the cell.




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2. These smaller hydrolysis products are used by fermentative bacteria for their
metabolism. As described above those bacteria release alcohols, organic acids,
hydrogen and carbon dioxide. Due to the production of organic acids this phase is
also called acidogenic phase.
3. In a third phase other fermentative bacteria use alcohols and higher organic acids like
propionic acid as substrates and produce the smallest organic acid, acetic acid, as
well as hydrogen. Due to the main product of this phase it is named acetic phase.
The bacteria Syntrophobacter wolinii degrades e.g. propionic acid to acetic acid and
hydrogen:
CH
3
CH
2
COOH + 2 H
2
O CH
3
COOH + CO
2
+ H
2
, G
0
= -76,1 kJ/mol (2.8)

The degradation of butyric acid by Synthrophomonas wolfei is an example for the
degradation of a long-chain fatty acid:
CH
3
CH
2
CH
2
COOH + 2 H
2
O 2 CH
3
COOH + 2 H
2
, G
0
= -48,1 kJ/mol (2.9)

4. Finally, in the fourth phase special methanogenic bacteria (archae bacteria) produce
methane via two different ways. Firstly methane is produced from carbon dioxide and
hydrogen, e.g. by Methanosarcina:

H
2
+ CO
2
CH
4
+ 2 H
2
O , G
0
= -135,6 kJ/mol (2.10)

This hydrogen-consuming reaction is important for the whole process because of
hydrogen is an inhibiting substance of reactions of the acetic phase.

Secondly, methane is produced from acetic acid, e.g. by Methanotrix:

CH
3
COOH CH
4
+ CO
2
, G
0
= -31 kJ/mol (2.11)
Normally, in biogas-reactors methane is mainly produced via acetic acid, but
hydrogen plays an important role in the regulation of these processes.

The steps of biogas production are illustrated in fig. 8. Fig. 9 shows the interaction between
organisms of the acetic phase and the methanogenesis.




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Fig. 8 and 9: the four steps of methane production and the role of hydrogen in
methanogenesis (Fritsche 1990)
The very low values of free energy obtained from reactions (eq. 2.10 and 2.11) are the
reason for the very low growth rate of methane producing bacteria.

It is possible to use the steps one to three including fermentation to produce hydrogen from
organic substances like organic waste biologically (eq. 2.9) . In spite of only a small portion of
the substrate is converted to hydrogen some research is done in this respect (Fischer 2009).

3 Kinetics of growth and metabolic processes
3.1 Cell division and growth
If there is enough substrate available bacteria grow by means of cell division. Under
conditions where substrates are limited very much, some bacteria produce spores instead of
the cell division process. Spores have no metabolism and are very resistant against extreme
conditions like temperatures, pressures, radiation, drought and chemical disinfection and
they are able to outlive decades. under good environmental conditions spores are able to
switch back to normal metabolism and cell division.




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If there is no limitation of substrates under good environmental conditions bacteria double
their quantity and their mass by producing a new cell generation by cell division within a
characteristic period, the generation time. Fast growing bacteria have a generation time of
down to 15 or 20 minutes. Due to this duplication within a certain period cells grow
exponentially under this conditions.
Therefore the quantity of cells N after n generations starting with a quantity of N
0
can be
calculated to
N = N
0
2
n
(3.1)
or
(3.2)
respectively
(3.3)

If a balance of the cell quantity for a batch reactor under exponential growth is done the
maximum growth rate
max
is normally used to describe the process.
Based on the general form of a balance
Ac = Tr + Re (3.4)
with Ac: Accumulation
Tr: Transport across the system boundaries
Re: Reaction of the balanced material
the balance of the quantity of cells for the system described is:
dN/dt =
max
N (3.5)
After integration and use of the initial condition N (t=0)= N
0
follows:
ln(N/N
0
) =
max
t (3.6)
With (eq 3.3) follows:
t = n ln2 /
max
t (3.7)
The generation time or duplication time t
D
follows for one generation or n=1:
t
D
= ln2 /
max
t (3.8)
n
N
N
2
0
=
) 2 ln( ln
0
n
N
N
=




Seite 15
From eq. (3.6) can be derived the exponential character of unlimited bacterial growth:
N/N
0
= exp(
max
t) (3.9)

Example:
Many aerobic heterotrophic bacteria have a generation time of approximately 30 minutes.
From eq. (3.8) the maximum growth rate can be derived as:

max
= ln2 / 0,5 h = 1,4 h
-1
(3.10)
How fast a lack of substrate is a limitation for growth shows the following calculation: If a
bacteria has the size of a cube with the dimension of 1 m its volume is 10
-18
m
3
. If a density
like water is assumed (10
6
g/m
3
) the bacteria has a mass of 10
-12
g.
After 67 hours of exponential growth the bacterial mass can be calculated from eq. (3.9), set
up for the mass m instead of the quantity N to:
m = 10
-12
exp(1,4 *67) = 2,18 * 10
28
g (3.11)
Compared with the earth-mass of 6 * 10
27
g more than three times of the earth mass would
have been grown.

Due to the limited amount of substrates and nutrients the growth of bacteria in a closed
system as a batch-reactor consists of four phases. During the lag-phase the cell adapts its
enzymes to available substrates without growth (s. fig. 3.1).








Fig. 3.1: Phases of bacterial growth (Fritsche 1990)




Seite 16
Then exponential growth takes place as long as substrate is not limiting. Under substrate
limitation bacteria growth slowlier use more internal substances for their metabolism and
more bacteria die. That results during this stationary phase in a approximately constant
biomass concentration. After all substrate is consumed the quantity and mass of bacteria is
reduced during the decay phase due to the death of bacteria and the metabolism of all
remaining internal substrates.

3.2 Influence of Temperature
If for a chemical reaction of the substrates A and B result in a product C
A + B -> C
a pseudo-first-order reaction rate can be defined as
r = k
A
c
A
(3.12)
with
k
A
: rate-coefficient
c
A
: concentration of substrate A
the rate-coefficient depends on temperature often according to the Arrhenius-equation:
(3.13)
with
k
0
rate coefficient at reference temperature
e
A
activation energy
R universal gas constant
T Kelvin temperature

Biological reaction are enzyme-catalysed reactions. Enzymes are proteins which catalyses
reactions in a certain range of temperatures. Regarding theses ranges bacteria can be
subdivided into mesophilic bacteria with a temperature range between 5 and 45C and a
range for optimal growth between 35 and 40C. THermophilic bacteria grow between 40 and
85C with an optimum between 0 and 60C. There also other species as e.g. psychrophilic
) exp(
0
T
e
k k
A

=




Seite 17
bacteria (optimum at app. 5 to 10C) and extreme thermophilic bacteria (optimum at app. 90
to 100C).

Example:
The influence of temperature can be calculated by using eq. (3.13). The relation of reaction
rates for two temperatures k
1
and k
2
can be derived as follows:

(3.14)

with
e
A
= 50 KJ/mol (Block 1992) and
R = 8,314 KJ/(kmol K) follows for usual temperatures 10, 20, and 30 C:
k
30/20
= 1,97 and
k
10/20
= 0,48.
It can be seen that a temperature increase or decrease of 10 C within the temperature
range of mesophilic bacteria results in twice respectively half of the reaction rate.

3.3 Influence of pH-value
All metabolic processes in cells are controlled by enzymes that consist of amino acids.
Amino acids contain different pH-sensitive side chains and therefore the activity of enzymes
depends on the pH-Value. The pH-Value inside cells is often regulated, but membrane
proteins and exoenzymes depend directly on the conditions of the surrounding medium. All
organisms produce enzymes for a certain range of pH-Value and are not able to grow below
or above a specific pH-limit.
Bacteria that produce e.g. lactic acid are able to grow at low pH-values down to pH4. Many
other bacteria cannot grow under this conditions that makes this process useful for
conservation of food products like some cheese, sour cucumbers, or sauerkraut.
In Table 3.1 some examples for bacteria are mentioned which grow at different optimal pH-
values.

=
2 1
1 2
1
2
) (
exp
T T
T T e
k
k
A




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pH-value Medium Organism
1 volcanic wells Thermoplasma acidophilum
2 citric acid, mine water Aspergillus, Thiobacillus
3 vinegar Acetobacter
4 sauerkraut lactic acid producing bacteria
5 cheese, sourdough lactic acid producing bacteria, yeasts
6 to 8 fresh water, sea water many micro organisms
9 alkaline soil, dung Nitrosomonas, Nitrobacter, urea degrading
bacteria, e.g. Bacillus pasteurii
10 fermented indigo leaves Bacillus alcalophilus
Tab. 3.1: pH-values, media, and micro organisms able to grow under these conditions
(Fritsche 1990, translated)

Additionally, the pH-value influences the dissociation of dissolved substances which are
often substrates for bacteria. The species Nitrosomonas for example oxidises ammonia to
nitrite. It could be shown (Dombrowski 1991) that only neutral molecule ammonia (NH
3
) can
be used, but not the ion ammonium (NH
4
+
) which is the product of the dissociation of
ammonia in water.
Fig. 3.2 shows the strong influence of pH-value of the concentration of the substrate
ammonia.
In biogas-reactors there is a specific risk, if bacteria producing organic acids are faster than
those consuming acids with the consequence of decreasing pH-values. At low pH-values
methane producing and acid consuming bacteria are not able any more to grow which results
in even lower pH-values to a level where acid producing bacteria stop their metabolism also.
Then no biological reaction takes place any more in the reactor which is a main accident for
a biogas plant.






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pH-value
Fig. 3.2: Influence of temperature and pH-value on the ammonia-ammonium-equilibrium
(Dombrowski 1991)

3.4 Growth kinetics
3.4.1 Michaelis-Menten kinetic
In 1913 in Berlin Leonor Michaelis and Maud Menten set up a theory of enzyme catalysed
reactions that allows to calculate quantitatively the influence of the substrate concentration
on reaction rates.
Michealis and Menten assumed an equilibrium reaction between the substrate S and an
enzyme E with an enzyme-substrate-complex ES and a complete reaction from ES to the
product P and enzyme E:
(3.15)
Further on it was assumed a first order reaction for k
2
and k
3
and a second order reaction for
k
1
. Then the reaction rate r can be defined as
(3.16)

with concentrations in [ ]-brackets and calculated to
P E ES S E
k k k
+ +
3 2 1
,
dt
P d
r
] [
=




Seite 20
(3.17)

Further on for the ES-complex can be written:
(3.18)

For steady-state-conditions for the ES-complex it follows:
(3.19)

The unknown concentration [ES] can be expressed by using the total enzyme concentration
[E
0
]:
(3.20)
Putting (3.20) and (3.19) into (3.18) results in
(3.21)

using the Michaelis-Menten-constant k
M
as
(3.22)

Putting (3.21) and (3.17) into (3.16) results in:
(3.23)

If all enzyme E
0
is active as ES-complex the maximum production rate (see 3.17, 3.16) can
be reached:
(3.24)
Putting (3.24) into (3.23) results in the Michaelis-Menton-kinetic:
(3.25)


] [
] [
3
ES k
dt
P d
=
] )[ ( ] ][ [
] [
3 2 1
ES k k S E k
dt
ES d
+ =
0
] [
=
dt
ES d
] [ ] [ ] [
0
E E ES =
] [
] ][ [
] [
0
S k
S E
ES
M
+
=
1
3 2
k
k k
k
M
+
=
] [
] [
] [
3
S k
S
E k r
M
o
+
=
] [
3 max o
E k r =
] [
] [
max
S k
S
r r
M
+
=




Seite 21









Fig. 3.3: Michaelis-Menten-kinetic with k
M
= 50

Fig. 3.3 shows the meaning of the Michaelis-Menton-constant as that concentration where
the reaction rate reaches 50% of the maximum reaction rate. The constant indicates the
range of very low substrate concentration and indicates a range of very low concentrations
where a first-order-kinetic fits approximately to the Michaelis-Menten-kinetic. There the
reaction rate is limited due to a lack of substrate (substrate limitation). At very high
concentrations a zero-order-kinetic describes the Michaelis-Menten-kinetic roughly.

3.4.2 Inhibitions
3.4.2.1 Substrate inhibition
At a high substrate concentration level sometimes the enzyme-substrate-complex ES (s. eq.
3.15) reacts additionally with another substrate molecule to another complex ES
2
.
(3.26)
Because of the portion of enzyme bound in ES
2
is not available for producing product the
production rate of the product decreases with increasing substrate concentration. A kinetic
modelling the whole reaction results in the substrate reaction rate r
S
:

(3.27)
0,000
0,250
0,500
0,750
1,000
0 1 2 3 4 5 6 7 8 9 10 11 12
[S] / kM
v

/

v
m
a
x
first-order reaction
zero-order reaction
2
ES S ES +
H
S
S M
S
S S
k
c
c k
c
r r
2
max ,
+ +
=




Seite 22

This kinetic formulation is also named Haldane-kinetic. It is to see that the reaction rate r
s
is
always lower than r
s,max
.
Under the condition
c
s
>> k
M

and c
s
= k
H

eq. (3.27) approximately results in
(3.28)









Fig. 3.4: Haldane kinetic (k
M
=50, k
H
=500)

An well known application is the conservation of marmalade by adding glucose up to high
concentrations in order to avoid glucose degrading bacteria and yeasts.

3.4.2.2 Other inhibitions
There are several other types of inhibitions. Two of them are the competitive inhibition and
the non-competitive inhibition. Inhibitors are often heavy metal ions, toxic organic
substances, or the product of a biochemical reaction like alcohol.
2
max , S
S
r
r =
0,000
0,250
0,500
0,750
1,000
0 4 8 12 16 20
[S] / kM
r

/

r
m
a
x






.
No Inhibition
Substrate Inhibition




Seite 23
A model of competitive inhibition consists of a further reaction to eq. (3.15) where the
inhibitor I reacts with free enzyme E to another complex EI. Hence there is a competition
between reaction E with S and E with I.












Fig. 3.5: competitive product inhibition with variation of product concentration c
P
(Kus 1993)

With increasing inhibitor concentration the reaction seems to increase their Michaelis-
Menten-coefficient, but the maximum reaction rate stays constant and can be reached if the
substrate concentration is high enough.

On the other hand the non-competitive inhibition is modelled by adding a reaction to eq.
(3.15) where the inhibitor reacts with the ES-complex to an ESI-complex. Therefore the
Haldane kinetic can be understood as a special form of non-competitive inhibition with the
substrate as inhibitor.
Fig 3.6 shows a non-competitive product inhibition with a variation of the product
concentration which seems to result in a decreasing maximum reaction rate at constant
Michaelis-Menten-coefficient.





Seite 24












Fig. 3.6: Non-competitive product inhibition with variation of product concentration c
P

(Kus 1993)

3.4.3 Monod-kinetic
In 1942, the French scientist Jaques Lucien Monod (1910 1976, Nobel price in 1965) did
investigations of the growth of Escherichia coli with glucose. He found out that the growth of
these bacteria can be described very well with the kinetic of Michaelis and Menten in spite of
the fact that bacterial growth is much more complex than the simple enzyme reaction of
Michaelis and Menten. But because of that mathematical formulation fits very well to the
experimental results of many investigations with different bacteria it is well known and widely
used for many bacteria (by far not for all) as Monod-kinetic for bacterial growth under normal
growth-conditions without inhibition and without bacterial decay.
The normal formulation is shown in eq. (3.29):
(3.29)
with
m: specific growth rate
S S
S
c k
c
+
=
max





Seite 25
k
S
: Monod-constant or saturation constant
The growth rate of bacteria r
B
then can be expressed as:
(3.30)
or with eq. (3.29):
(3.31)

During exponential growth often the relation between growth rate and the rate of substrate
degradation or product production is a constant value which are named yield-coefficients,
e.g.:
(3.32)
with
Y
B/S
: Yield-coefficient between bacterial growth and substrate degradation.

(3.33)
with
Y
B/P
: Yield-coefficient between bacterial growth and product production
(3.34)
with
Y
S/P
: Yield-coefficient between substrate degradation and product production.

Yield coefficients are constants under conditions of exponential growth. During this growth
bacteria do a metabolism specialised to maximum growth. Under other conditions, e.g.
substrate limitation or inhibition many bacteria are able to change and adapt their metabolism
which results in other values of yield coefficients. Therefore yield coefficients from literature
has to be checked seriously whether they were obtained under appropriate conditions for
own purposes.


B B
c r =
B
S S
S
B
c
c k
c
r
+
=
max

S
B
S B
r
r
Y =
/
P
B
P B
r
r
Y =
/
P
S
P S
r
r
Y =
/




Seite 26
4 Bioreactor design
Most of anaerobic bioreactors are designed as a so called chemostat reactor or CSTR
(continuously stirred tank reactor), which is a completely mixed reactor (fig. 4.1).






Fig. 4.1: Chemostat reactor

If steady state conditions are assumed and if there are no bacteria in the influent a mass
balance for bacteria results in the following formulation:
(4.1)
with
V
R
: reactor volume
With the definition of the dilution rate D and the hydraulic residence time t
V


(4.2)
follows from eq. (4.1):
(4.3)

Using the Monod formulation eq. (3.29) eq. (4.3) results in:

(4.4)


V
0
c
B,a
.
V
0
c
B,a
.
R a B a B
V c c V + =

, ,
0 0
R V
V
V
t
D

= =
0
1
D =
1
max
,

=
D
K
c
S
a S

Seite 27















Fig. 4.2: Results of eq. 4.4 as well as bacteria concentration
(with S
S
: effluent substrate concentration, X
S
: effluent biomass concentration,
S
0
: influent substrate concentration

max
= 1,0 h
-1
, k
S
= 0,005 g/L, Y
X/S
= 0,5)

It is seen that for =D no biomass any more is in the reactor and no substrate degradation
takes place. At this wash-out-point the transport of bacteria out of the reactor is equal to the
maximum growth rate of bacteria. A slight increase of the dilution rate increases the transport
term resulting of a loss of all bacteria after steady state conditions are met again.
Therefore for a stable operation under real variations of operating conditions it is necessary
to set a dilution rate sufficient lower than the critical wash-out-dilution-rate.






Seite 28
5 Applications
5.1 General process design
One stage CSTR-types of reactors are widely used for decades for anaerobic treatment of
sewage sludge. A typical egg-shaped reactor is shown in fig. 5.1. Egg-shaped reactors were
developed to reduce the risk of sand sedimentation on the bottom and foam formation on the
top. But other more cylindrical reactors are used as well.








Fig. 5.1: Mesophilic anaerobic sewage sludge treatment on the waste water treatment plant
Bochum-lbachtal: Volume 2 x 8720 m
3
, 3 x 770 KW cogeneration plant (Oswald-Schulze)

If the content of dry matter is chosen in the range of 15 to 50% processes are sometimes
called dry fermentation (s. fig. 5.2) which need other transport and mixing devices. Those
systems are also offered as container-based fermentation systems where in particular
attention should be given on the heat loss across the container surface.







Fig. 5.2: So-called dry fermentation system with plug-flow-behaviour (Linde, BRV-process)




Seite 29













Fig. 5.3: Digester with CSTR- or plug-flow-behaviour for substrates with low solid content,
e.g. waste water (Schwarting-Uhde)

By adjusting a recirculation pump the behaviour of the Schwarting-Uhde reactor can be
chosen as CSTR (high recirculation) or as plug-flow-reactor (no circulation) (s. fig. 5.3).
Two stage systems are sometimes used to reach high performance systems by adjusting the
conditions in stage 1 for hydrolysis and acidification (pH 5 to 6,5) and in stage 2 for
acetogenesis and methanogenesis (pH 7 to 7,5) (Kaltschmitt 2001). This can be done with all
the material in two separate e.g. CSTR-systems or solid and liquid phases are separated in
between, like in the Wherle-system (s. fig. 5.4).
Other systems are available with low investment costs for discontinuous operation (batch
systems) (s. fig. 5.5).
An overview of various designs of anaerobic reactors is shown in Fig. 5.6.






Seite 30













Fig. 5.4: Two-phase and two-stage fermentation (Wherle)










Fig. 5.5: Batch system for discontinuous use without mixing (Aschmann in Biogas 2007)






Seite 31













Fig. 5.6: Overview of various anaerobic continuously operated reactor systems (Kaltschmitt
2001)

5.2 Gas production
5.2.1 Gas quality
The gas quality depends mainly on the substrate composition. The theoretical portion of
methane and carbon-dioxide for pure known substrates can be calculated stoichiometrically
(Biogas 2007):

(5.1)

Results of this calculation related to real, but pure substrates are shown in fig. 5.7. The
fermentation of real mixed substrates results often in a methane content of 50 to 60%.

4 2 2
4 8 2 4 8 2 2 4
CH
b a u
CO
b a u
O H
b a
u O H C
b a u

+ +

+




Seite 32













Fig. 5.7: Theoretical gas composition of pure substrates (Kaltschmitt 2001)

Besides the main components methane and CO
2
there other components present in low
concentrations (s. tab. 5.1). In particular hydrogen sulphide is important as a hazardous
component and as a substance that reacts to sulphur dioxide in incineration processes and
which results therefore in air pollution.





Tab. 5.1: Components of biogas (Kaltschmitt 2001)






Seite 33
5.2.2 Gas yield
Also the amount of gas that can be produced from substrates can be calculated
stoichiometrically. Tab. 5.2 shows some figures related to the input mass of dry matter
(Trockenmasse, TM)



Tab. 5.2: Gas yield of various substances (Biogas 2007)

In literature there are many data of gas yields for real substrates available that are normally
measured at laboratory conditions and long duration of the fermentation. Therefore in real
plants the gas yield under less optimum conditions sometimes might be lower than expected.
The following tables 5.3 to 5.7 show some gas yields and other data of real substrates. In
these tables mean
TM: dry matter (Trockenmasse), FM: wet matter (Feuchtmasse)
oTM organic dry matter




Tab. 5.3: Data of some manure substrates (Biogas 2007)






Tab. 5.4: Data of some agricultural products (Biogas 2007)




Seite 34




Tab: 5.5: Data of same waste material (Biogas 2007)







Tab. 5.6: Data of some food waste material (Biogas 2007)

The gas yield of unknown substrates has to be investigated in laboratory tests.

5.2.3 Reaction rates
There are many results available regarding one of the specific steps of anaerobic
degradation that are necessary for models and simulation. Due to the limited duration of this
lecture only some examples and general results can be discussed here.
Because of hydrolysis often is the slowest step of the degradation chain, anaerobic digestion
of solid material. On the other hand in solid material often organic substances are much
more concentrated than in liquid material.
Fig. 5.8 shows the influence of hydraulic residence time t
V
on the COD-conversion a for a
fixed bed bioreactor in order to achieve a high bacteria concentration (using plastic foam
cubes of 0,7 and 1,0 cm ). A highly concentrated wastewater from the treatment of sewage
sludge was used as substrate containing approximately 30% of COD as not degradable
under this conditions.




Seite 35











Fig. 5.8: COD conversion rate depending on hydraulic residence time for highly concentrated
wastewater (Spie 1991)












Fig. 5.9: Methane production rate depending on hydraulic residence time (Spie 1991)





Seite 36
For the same investigations fig. 5.9 shows the methane production rate which could be
achieved. The results show the typical behaviour of a pseudo first-order reaction at constant
biomass concentration. At low residence times that means operating the reactor at high load
conditions the specific reaction rate per reactor volume reaches highest values. But only a
small portion of the input material is converted to biogas at low residence times. Therefore
often lower specific production rates are chosen in order to achieve a better use of the
substrate which means a higher conversion. In general an economically design results in an
optimum operation point of a certain conversion rate at as high as possible reaction rates
under this specific conditions.
If reactors are operated at production rates high concentrations are present in the reactor
which might result in inhibition effects. Fig. 5.10 shows the inhibition of the degradation of
propionic acid by high concentrations of propionic acid. Due to other investigations it is
known that the undissociated acid is used as substrate by bacteria. Therefore the substrate
concentration in fig. 5.10 is expressed as undissociated propionic acid.













Fig. 5.10: Conversion rate of dissolved organic matter depending the concentration of
undissociated propionic acid (Kus 1993)





Seite 37
Due to slow hydrolysis residence times for the conversion of solid material are considerably
longer. Results from the digestion of sewage sludge show a conversion of up to 50% of
organic matter at residence times of 20 days in CSTR-reactors (s. fig. 5.11).













Fig. 5.11: Conversion of organic matter from sewage sludge by anaerobic digestion at
mesophilic conditions ( Pfeiffer 1989).

Depending on substrate quality sometimes far higher residence times are necessary in order
to achieve a high conversion.
Fig. 5.12 shows lab-scale results of anaerobic digestion of organic waste after aerobic pre-
treatment. Black squares indicate results under controlled temperature with a high
conversion after 40 days. Even if treatment periods from batch tests cannot be interpreted as
residence times under continuous operation, results show that far longer residence times
than 20 days are necessary to convert a portion of input material to biogas. In technical
applications hydraulic residence times of approximately 60 days are already realized.






Seite 38











Fig. 5.12: Gas yield of organic waste (Linke 1999)

If inhibition play a role in an individual application growth rates of bacteria and therefore the
production rate of biogas is lower than without inhibitions. That might result in higher
residence times necessary to achieve a high conversion of the input material.
Kaltschmitt reported results (Kaltschmitt 2001) of the inhibition by sodium at concentrations
of some g/L in some cases. In other cases bacteria were able to adapt to concentrations up
to 60 g/L without inhibition effects.
It is known that hydrogen as an intermediate product inhibits the degradation of organic acids
at concentrations of some mol/L.
Heavy metals are known also as inhibitor if they are not precipitated by sulphide. Kaltschmitt
reports inhibition effects at some 10 mg/L depending on the type of metal.
Also ammonia (not ammonium!)is known as inhibitor. Due to the dissociation equilibrium
between ammonium and ammonia the pH-value influences inhibition effects very much.
Kaltschmitt reports an inhibition of ammonia at concentration of 150 mg/L.






Seite 39
5.2.4 Example: Digestion of slaughterhouse waste
Fig. 5.13 shows a simplified example of anaerobic digestion of slaughterhouse waste. Gas
yield, conversion, nutrient concentration, and other parameters depend on the mixture of
different slaughterhouse wastes like flotation waste (high fat content), blood waste (high
protein and nitrogen content), or stomach contents (other organic materials) in any individual
case.
In this example a high conversion of 85% is assumed due to the quality of raw material and a
residence time of approximately 40 days. Depending on the water content of the input
material and pH-value high ammonia concentration might occur. Therefore attention should
be paid to inhibition of the process by ammonia. If the output material should be used as
fertilizer also attention has to be paid to a limitation of nutrients applied on agricultural areas
in order to prevent ground water contamination by e.g. nitrate.











Fig. 5.12: Energy and nutrient balance of anaerobic digestion of slaughterhouse waste

Further aspects that were not mentioned yet are biogas desulphurisation, off-gas cleaning,
gas storage, output storage, excess-heat utilization, the conversion of biogas to bio-methane,
or a post-treatment of output material solid-liquid separation, composting of solids, further
processing of the liquid phase, or nutrient separation from output material.
Biogas
reactor
gas
engine
Input:
2800 Mg/a dry matter (slaughterhouse waste)
124 Mg/a N
17 Mg/a P
9 Mg/a K
Ouput:
400 Mg/a dry matter
124 Mg/a N
17 Mg/a P
9 Mg/a K
Biogas
5000 m
3
/d
60% Methane
6 KWh/m
3
:
Energy: 1250 KW
Electricity: 500 KW
(40 % efficiency):
Heat: 537 KW
(43% efficiency)
Off-gas




Seite 40

Literature

Biogas 2007 Biogashandbuch Bayern, Materialienband, Juli 2007
Block 1992 Block,J.: Untersuchungen zur Kinetik der Abwasserreinigung durch ae-
rob thermophile Bakterien. In: VDI (Hrsg.): VDI-Fortschrittsberichte Rei-
he 15, Nr. 98, VDI-Verlag Dsseldorf 1992
Dombrowski 1991 Dombrowski, T.: Kinetik der Nitrifikation und Reaktionstechnik der Stick-
stoffelimination aus hochbelasteten Abwssern. In: VDI (Hrsg.): VDI-
Fortschrittsberichte Reihe 15, Nr. 87, VDI-Verlag Dsseldorf 1991
Fischer 2009 Fischer, L.: Wasserstoff aus Biomll. Spektrum der Wissenschaft, Juni
2009, p.14.
Fritsche 1990 Fritsche, W.: Mikrobiologie, Gustav-Fischer-Verlag, Jena 1990
Kaltschmitt 2001 Kaltschmitt, M., Hartmann, H. (Hrsg.): Energie aus Biomasse. Springer
Verlag Berlin Heidelberg New York, 2001
Kus 1993 Kus, F.: Kinetik des anaeroben Abbaus von Essig- und Propionsure in
Bioreaktoren mit immobilisierten Bakterien. In: VDI (Hrsg.): VDI-
Fortschrittsberichte Reihe 15, Nr. 115, VDI-Verlag Dsseldorf 1993
Linke 1999 Linke, B, Schelle, H.: Verfahren zur Biomethanisierung von Feststoffen.
in: Frdergesellschaft erneuerbare Energien e.V. (HRSG.): Stoffliche
und energetische Nutzung von Biomasse im Land Brandenburg. Tagung
im Mai 1999 in Paaren, Berlin 1999
Pfeiffer 1989 Pfeiffer, W.: Verfahrensvarianten der Faulung und Entseuchung Leis-
tungsvergleich. Dissertation an der TU Mnchen 1989, in: Roediger, M.:
Bemessungsvorschlag fr Schlammfaulungsanlagen. Korrespondenz
Abwasser 1997 (44), S.1856
Schgerl 1985 Schgerl, K.: Bioreaktionstechnik Band 1. Otto-Salle-Verlag Frankfurt,
Berlin, Mnchen 1985, S.132




Seite 41
Spie 1991 Spie, A.: Anaerobe Abwasserreinigung in neuen Bioreaktore mit Polyu-
rethan-Schaumstoffpartikeln zur Immobilisierung der Bakterien. In: VDI
(Hrsg.): VDI-Fortschrittsberichte Reihe 15, Nr. 85, VDI-Verlag Dsseldorf
1991