THAI NGUYEN UNIVERSITY
UNIVERSITY OF AGRICULTURAL AND FORESTRY

DAM HA LUONG THANH

BIOMETHANE POTENTIAL TEST FOR RAPID EVALUATION OF
ANAEROBIC DIGESTION OF SEWAGE SLUDGE FROM MULTIPLE
MATERIALS FOR A PROPOSED LARGE-SCALE DIGESTER

BACHELOR THESIS

Study Mode: Full-time
Major:

Environmental Science and Management

Faculty:

International Training and Development Center

Batch:

2012 - 2016

Thai Nguyen, 2016


DOCUMENTATION PAGE WITH ABSTRACT
Thai Nguyen University of Agriculture and Forestry
Degree Program Bachelor of environmental Science and Management
Student name

Dam Ha Luong Thanh

Student ID

DTN1353110232
Biomethane potential test for rapid evaluation of anaerobic digestion

Thesis Tiltle

of sewage sludge from multiple materials for a proposed large-scale
digester

Supervisor

Assoc. Prof. Dr. Nguyen The Hung, Thai Nguyen University of
Agriculture and Forestry, Vietnam

Abstract
This paper examines the biomethane potential from organic wastes. The biomethane
potential test is used to assess the suitability of different substrates for biomethane
production. A methodology for accurately estimating the biomethane potential from
multiple heterogeneous organic waste substrates is sought. Three main substrates were
identified as possible substrates for biogas production, namely: pig manure, food waste,
and beer processing waste. The biomethane potential of these substrates ranged from as
low as 82 L CH4 kgVS-1 for beer processing waste to as high as 112 L CH4 kgVS-1 for
food waste treatment. The objective of the paper is to suggest an optimum substrate mix
in terms of biomethane yield per unit substrate for the proposed anaerobic digester. This
should maximise the yield of biomethane per capital investment. Pig manure displayed
the highest biomethane yield (13 Lkg-1) followed by food waste (3 Lkg-1) and beer

processing waste (2 Lkg-1).
Keywords

biomethane potential test, anaerobic digestion, sewage sludge,
biogas

Number of papers

40 pages

Date of submission:

21/09/2016

Supervisor’s signature:

ii


ACKNOWLEDGEMENTS
Completion of my Bachelor Thesis at Thai Nguyen University of Agriculture and
Forestry has not been achieved by my efforts alone, but memorably contributed by
many wonderful people to whom I must express my great thanks.
My sincere gratitude is offered to Assoc. Prof. Nguyen The Hung who gave me a
precious opportunity to carry out this study along with his enthusiastic support
throughout my thesis with his patience and knowledge whilst allowing me the room to
work in my own way. I attribute the level of my Bachelor degree to his encouragement
and effort.
A word of thanks must be also recorded to Ms. Anurag Deo and Ms. Mette Axelsson
Bjerg


from Linköping University for their commitment and companionship as

teammates throughout this study, especially during the time they constructed their
Master Thesis of Biogas plant in Thai Nguyen Agriculture and Forestry (TUAF). I
would also like to offer special thanks to Mr. Duong Manh Cuong (MS), lecturer in
Faculty of Biotechnology and Food Technology, TUAF for his suggestions and
assistance in setting up my experiment.
I am also grateful to all laboratory technicians and managers, Mrs. Hoan, Mrs, Phuong,
and Mr. Lam from Faculty of Biotechnology and Food Technology, TUAF for their
whole-hearted help during the time I carry out my experiment.
To my family, my heartfelt thanks are expressed for their unconditional love and
belief.
DAM HA LUONG THANH

iii


TABLE OF CONTENS
List of figures................................................................................................................. vi
List of table ................................................................................................................... vii
List of abbreviation ...................................................................................................... viii
PART I: INTRODUCTION .........................................................................................1
1.1. Research rationale .................................................................................................1
1.2 Research’s objectives .............................................................................................3
1.3 Research questions .................................................................................................3
1.4 Limitation ...............................................................................................................3
PART II: LITERATURE REVIEW ............................................................................5
2.1 Sewage sludge ........................................................................................................5
2.1.1 Type of sewage sludge ....................................................................................5

2.1.2 Component of sewage sludge ..........................................................................7
2.2 Anaerobic sludge digestion ....................................................................................9
2.2.1 Fundamentals of anaerobic digestion ..............................................................9
2.2.2 Current status of anaerobic digestion process ...............................................10
2.2.3 Factors influencing the anaerobic digestion process .....................................12
2.2.3.1 pH ............................................................................................................13
2.2.3.2 Alkalinity .................................................................................................14
2.2.3.3 Temperature.............................................................................................14
2.2.3.4 VFAs .......................................................................................................15
2.2.3.5 Ammonia .................................................................................................16
2.2.4. Nutrient requirements for anaerobic digestion .............................................17
2.2.5 Anaerobic digestion in sewage sludge treatment ..........................................18

iv


2.3 Summary ..............................................................................................................20
PART III: MATERIALS AND METHODS .............................................................21
3.1 Materials ...............................................................................................................21
3.1.1 Inoculum ........................................................................................................21
3.1.2 Substrates .......................................................................................................21
3.2 Experimental methods ..........................................................................................21
3.2.1 Biomethane potential test equipment ............................................................21
3.2.2 Experiment protocols .....................................................................................22
3.2.2.1 Analyzing total solid (TS), volatile solid (VS) and organic loading rate
(OLR) ..................................................................................................................22
3.2.2.2. Preparing solution...................................................................................23
3.2.2.3 Biomethane Potential (BMP) experiment ...............................................25
PART IV. RESULT .....................................................................................................28
4.1 Characteristics of inoculum and materials ...........................................................28

4.2 Operation results ..................................................................................................28
4.3 Methane potential per mass of substrate ..............................................................30
PART V. DISCUSSION AND CONCLUSION ........................................................32
5.1 Discussion ............................................................................................................32
5.2 Conclusion............................................................................................................33
REFERENCES ............................................................................................................34

v


LIST OF FIGURES

Figure 1. Anaerobic digestion process of organic maters (Thanh, N. 2014) ................10
Figure 2. Factors influencing AD performance .............................................................13
Figure 3. Analysimg CH4 concentration .......................................................................26
Figure 4. Monitoring biogas production ........................................................................26
Figure 5. Methane production curve of materials .........................................................30

vi


LIST OF TABLE
Table 1. Typical constituents of different types of sludge .............................................8
Table 2. Summarises the optimal concentration and effects of these nutrients in the
anaerobic digestion process ...........................................................................................17
Table 3. Advantages and disadvantages of anaerobic sludge digestion ........................19
Table 4. Preparation of nutrient solution .......................................................................23
Table 5. Preparation of sodium hydrate solution ..........................................................24
Table 6. Characteristics of inoculum and materials ......................................................28
Table 7. Bio-methane potential of substrates ................................................................29

Table 8. Weighted average methane potential per kg of substrate ................................30

vii


LIST OF ABBREVIATION

AD

:

Anaerobic digestion

BMP

:

Biomethane potential test

ORL

:

Organic loading rate

TS

:

Total solids


VFAs

:

Volatile fatty acids

VS

:

Volatile solids

Ww

:

Wet weight

WWTs

:

Wastewater treatment

WWTPs :

Wastewater treatment plans

viii



PART I: INTRODUCTION
1.1. Research rationale
In recent years, there are several techniques for the treatment and management of
sewage sludge, including landfill, incineration, composting, and anaerobic digestion
(AD) process. Among them, AD is the most commonly used technique since biogas,
which is a valuable form of bio-energy, can be extracted from sewage waste.
Anaerobic digestion is a process by which organic materials are naturally broken down
into biogas and bio-fertilizer. In this process involving several sequential biochemical
stages, many different microorganisms participate in a complex web of interacting
processes which result in the decomposition of complex organic compounds as
carbohydrates, fats and proteins to the final products as methane and carbon dioxide.
This process occurs naturally in many environments with limited access to oxygen, for
example in bogs and marshes, rice paddies and in the stomach of ruminants, such as
cows.
Besides, it happens in the absent of oxygen naturally, or in sealed, free-oxygen tanks
called anaerobic digesters. AD of solid organic waste such as bio-waste, sludge, cattle
manure, energy crops, and other biomasses, for bio-energy production is widely
applied technologies. The production of biogas in AD offers several advantages over
the other alternatives. These include biogas production, nutrient recovery, and
reduction of waste organic content and pathogen agents.
Sewage sludge can be described as a byproduct mixture of solid and water from
wastewater treatment (CIWEM, 1995). By applying several different treatment

1


processes, the resulting sewage sludge types extremely differ in their characteristics.
Constituents of sewage sludge regarding to carbonhydrate, proteins, lipids are highly

depended on their origin. The presence of significant concentration of nitrogen,
phosphorus, and potassium in sewage sludge make it possible for fertilizing soil
because these elements are essential for plant growth.
Anaerobic digestion instability is resulted from the fluctuation in organic loading rate,
heterogeneity of waste or excessive inhibitors. Towards improving AD performance in
biogas production and accelerating the microbial activities for higher quality of biosolids, several environmental conditions should be meticulously controlled.
Additionally, various studied have demonstrated that hydrolysis phase is a ratelimiting stage, and seriously impacts on the performance of AD.
At present, end-users in Vietnam, often have difficulties in controlling the technology
efficiently, due to poor management competence (Jiang et al., 2011). This leads to
production being inadequate in periods of high demand in low temperature regions
during winter, and excessive during periods of high temperature and high production
of excreta (Cu et al., 2012). There is thus a need to improve knowledge about biogas
production potential using local biomass, in order to develop digesters adapted to the
local environment and individual management schemes, thus ensuring production of
the gas needed for cooking, heating and light (Vu et al., 2007; Cu et al., 2012). Hence,
there is an associated need to review, develop and validate methods to assess biogas
production which can be used in laboratories with limited access to analytical
instruments. Research carried out at laboratories in regions with limited access to high-

2


tech instruments must be of international standard, so as to ensure useful results and
contribute to progress in development of the technology.
1.2 Research’s objectives
This paper aims to assess and screen potential substrates from three major waste
streams for a proposed anaerobic digestion facility using the biochemical methane
potential (BMP) test which can be carried out in simple laboratories. The BMP test is
also used to assess the level of variability of biomethane potential (methane
concentration in biogas) within the waste streams by using Dr. Einhorn’s fermentation

saccharometer with dilution tube and absorption of CO2 in alkaline liquid (7M NaOH)
. The objective is to recognize substrates with a high methane production per unit mass
in lab-scale with limited access to analytical equipment, which will lead to an
economic digester design in future.
1.3 Research questions
This study is operated to investigate these following issues:
o How much biogas can be produced from the substrates?
o What is methane potential of the substrates?
1.4 Limitation
The increasing demand of renewable source of energy and quality of bio-solids has
determined as a great deal to formulate the feasible treatment processes applied in
WWTPs. In addition to sewage sludge stabilization, AD has been known to produce
biogas, which is renewable fuel. Using organic materials is expected to enhance the
efficiency of anaerobic digesters. Furthermore, a more comprehensive understanding
3


of key physiochemical properties of the substrate, operational conditions, and biogas
potential is of great necessary prior to any large-scale opperations.
The BMP assay is designed to provide ideal anaerobic conditions and prevent any
form of biochemical inhibition. To ensure this, three important conditions should be
met throughout the BMP assay (Labatut, et al, 2010): (1) appropriate microbial
community, enzyme pool, and nutrients are present; (2) environmental conditions are
optimal; and (3) substrate and intermediate product concentrations are well below
inhibitory/toxic levels. BMP results should be limited to a relative interpretation of the
substrate’s methane potential, and not for an absolute estimation of daily biomethane
yields or the overall performance and stability of large-scale digesters, it is best suited
when used to elucidate what types of substrates, from an array of potential substrates,
have the highest biomethane potential.


4


PART II: LITERATURE REVIEW

This chapter provides an overview of the current knowledge regarding bio-methane
potential test, including anaerobic digestion of sludge sewage and other organic waste
materials. The AD process is firstly presented and discussed. This is followed by a
comprehensive review of CH4 production by anaerobically digesting sewage sludge
with other substrates.
2.1 Sewage sludge
In the effort of improve effluent quality, waste water treatments (WWTs) are built and
upgraded. While these plants can produce high effluent quality, sludge disposal
remains underlying issues. These include the expensive cost of sludge treatment,
which makes up more than 50% of total WWTs cost (Rulkens, W., 2007), and
potential risks associated with the sludge disposal for the environment and human
health.
Sewage sludge is a mixture of solids and semi-solid removed from the liquid stream of
WWPs. A more restricted definition is “a residual solid from sewage plants treating
domestic and urban waste water and from other sewage plants treating waste water of
a composition similar to domestic and urban waste water”
2.1.1 Type of sewage sludge
To assess options for sludge treatment and disposal, it is necessary to investigate
different kind of sludge and origins. A typical sewage treatment plants includes
primary, secondary, and tertiary processes (Fytili, D., et al, 2008).
5


Primary sludge is collected from primary treatment process containing high total solids
(TS) content. The characteristics of primary sludge vary considerably depending on

the initial compositions of wastewater, the efficiency of primary sedimentation and the
usage of chemicals in sedimentation (Guyer, J.P., 2011)). Primary sludge may contain
oil, grease, vegetable materials, paper, faecal materials, sanitary and medical waste,
kitchen waste.
Treatment process such as activated sludge process, or rotating biological contactors
results in humus sludge or biological sludge (Arnaiz, C., et al, 2006). Humus sludge is
the settled product from soluble waste in the primary effluent. This is a mixture of
microorganism: sloughed bacteria and fungus under living or dead remains. Humus
sludge has earthly smell and color of dark brown.
Humus sludge from biological aerated filters and their variation, which have different
types of biological media, share certain characteristics with activated sludge. In
practice, humus sludge is returned to co-settle with primary sludge in the primary
settle.
Activated sludge is removed from the activated sludge process. Main components of
activated sludge are flocculated and synthesised solids and microorganism (CIWEM,
1995). Due to the rate of recycling and other factors, activated sludge has low TS (1%)
with the color ranging from grey, dark brown to black.
In the tertiary treatment step, the product is called tertiary sludge. It has fractions in
common with secondary sludge, which remains in the effluent of the secondary
treatment step and removed in the tertiary step. This sludge is normally transferred to
primary tanks and co-settle with primary sludge due to its small amount.
6


Digested sludge, as bio-solids, is the product of biological digestion. This process
can be performed in the reactor with or without the presence of oxygen,
corresponding in the aerobic or anaerobic digestion processes. Bio-solids contain
nutrient (Jarrell, K.F., et al, 1992) thus they should be considered as a resource. They
could also contain pathogens, which must be carefully managed because of their
impacts on public health. Bio-solids are classified due to the level of their

contaminant and stabilization. As these levels are assessed, the beneficial use of biosolids will be divided into three sectors: Unrestricted, Restricted, and Not Suitable
for Use. (Kostenberg, D., et al, 1993).
Combination of different sludge type is commonly utilized in sludge treatment. This
could be clarified with diverse characteristics and compositions of mixed sludge.
Regarding AD, the composition of sewage sludge is the mixture of primary and
secondary sludge. (Yamada, T., et al, 2005; Noutsopoulos, C., et al, 2012).
2.1.2 Component of sewage sludge
It is important to know characteristics of sewage sludge for its effective treatment and
disposal. Generally, sludge includes volatile, organic solids, nutrients, metals, organic
pollutants, and water (Rulkens, W., 2007; Zaha, C., et al, 2008). Table 1 summarizes
some analyses of sledge components from the literature.
Total solid (TS) content influences the ability of sludge transference in the sewerage
system. The higher amount of TS, the more difficultly sludge flows. Thus, it is
necessary to maintain sludge in liquid stage, which makes sludge flow easier from
vessels and through pipes. Sewage sludge should be qualified for TS prior to any
sludge treatment processes.
7


The value of TS content after being treated can change basing on different treatment
methods. After thickening, TS content of sludge will increase up to 9%, and reach 25 –
35% after mechanical dewatering. (Milieux, Z.d., 2003)
Table 1. Typical constituents of different types of sludge (Fytili, D., et al, 2008;
CIWEM, 1995)

Type of sludge
Constituent

Unit


Untreated

Digested

Activated

primary sludge

primary sludge

sludge

Total solids (TS)

%

2.0 – 8.0

6.0 – 12.0

0.83 – 1.16

Volatile solids (VS)

% TS

60 -80

30 – 60


59 - 88

5.0 – 8.0

6.5 – 7.5

6.5 – 8.0

pH

The solid content of has 59 – 88% of volatile solids (VS) on dry weight basis. VS
content mainly contains organic compounds of animal or pant origin. It is defined as
the mas of solid materials that can be lost through evaporation or oxidation at 550oC.
VC is an important character of the odour problem of sludge; thereupon, the reduction
of VS is one of the main objectives in sludge treatment. A series of treatment methods,
including AD, aerobic digestion, composting, and incineration are used to minimize
the VS content (Thanh, N., 2014). AD can biologically convert around 50% of VS to
biogas.

8


2.2 Anaerobic sludge digestion
2.2.1 Fundamentals of anaerobic digestion
AD is a process in which organic matter can be biodegraded in the absent of oxygen
by a consortium of microorganisms. An important product of AD is biogas, which
mainly contain CH4, carbon dioxide (CO2) and traces of other gases (Clemens, J., et al,
2006). AD involves series of biochemical reactions, which can be divided into four
stages, namely hydrolysis, acidogenesis, acetogenesis, and methanogenesis (Figure 3).
AD has been used to treat biodegradable organic and produce biogas (5). AD is a

sequential process involving several complex biochemical stages. Each stage is
consistently performed under activities of interaction of different bacteria. In
hydrolysis stage, hydrolytic microorganisms hydrolyse polumer materials to form
monomers, such as amino acids and glucose. These monomers are then converted to
H2, CO2 and short-chain fatty acids such as acetic, propionic acids in the next step,
namely acidogenesis. In aectogenesis phase, syntrophicacetogenic bacterial metabolize
these volatile fatty acids (VFAs) to produce precursors for the methanogenic
fermentation. In the end, CH4 is formed from either acetate or CO2 and H2 by
methanogenic bacterial in methanogenesis step.

9


Complex particulate organic matter

Hydrolysis

Soluble organics (simple sugars,
alcohol, organic acids, amino acids)

Acidogenesis
VFAs

Acetohenesis
Acetate

H2, CO2

Aceticlastic
Methanogenesis


Hydrogenotrophic

CH4

methanogenesis

Figure 1. Anaerobic digestion process of organic maters (Thanh, N. 2014)
2.2.2 Current status of anaerobic digestion process
Anaerobic digestion is a natural process which occurs in several environments, such as
wetland, rice fields, intestinal tracts of animals, marine or fresh water sediments.
Humans have applied this process to take benefits as energy, rapid decomposition of
organic waste, and stabilized residue for a long time.

10


Over the last two decades, a great deal of the literature has been published on feasible
applications of AD for solid waste and wastewater treatment. Apart from biogas
production, AD brings much greater potential due to more intrinsic merits, including
energy saving, nutrient recovery, reduction of waste organic content to the
conventional aerobic digestion (Wilkie, A.C., 2005; Nasir. I.M., et al, 2012; Demirel,
B., et al, 2005). As a result, extensive application of AD have been only revealed
recently with a number of developing designs by focusing on more complicated
devices and operational techniques, and increased understandings of microbiology and
biochemistry. There are various anaerobic reactor types in practice, of which batch
reactors are the simplest configuration. The one-stage continuously fed systems, the
two-stage and multistage continuously fed systems were more advanced reactors
applied for AD treatment (Ward, A.J., et al, 2008)
The evolution of AD applications was also confirmed by a broad range of potential

substrates for this process. Anaerobic technology such as single-phase (conventional)
and two-phase anaerobic digesters was often used in the treatment of dairy wastewater
for energy product and waste stabilization. In term of low content of suspended solids
in dairy wastewater, the conventional anaerobic reactors are generally nominated for
treatment. Currently, variety studies of dairy wastewater treatment have shown a wide
range of application of anaerobic reactor designs, such as down-flow film, anaerobic
filter, up-flow anaerobic sludge blanket. At laboratory scale, the efficient removal of
chemical oxygen demand (COD) of these reactors could reach up more than 90%
(Dermirel, B., et al, 2005). The authors also studied the two-phase anaerobic treatment,
which was applied for dairy wastewater comprising high concentrations of non-filtered
solids, and lipids. The prevalent reactor type was continuously stirred tank reactor
11


(CSTR), and up-flow filter, with CSTR used for acidogenesis stage while up-flow
responsible for methanogenesis. In comparison between these two processes, the twophase shows better outcomes with various kinds of industrial wastewater. Sludge from
WWTPs has also aroused much consideration since some strict rules of sludge
disposals were adopted (Scragg, A.H., 2005). Due to advantages of AD, it has become
one of the bright solutions for sludge stabilization and energy production (Arnaiz, C.,
et al, 2006; Rajagopal, R., et al, 2011; Tomei, M.C., et al, 2009). Current
improvements of high-rate anaerobic system have been drawing more attention on AD
performances in agricultural waste treatment, especially animal residue (Ward, A.J., et
al, 2008), which have different features from those of municipal and industrial
wastewater (Lorimor, J., et al, 2000). Anaerobic treatment of the poultry and livestock
manure waste, two kinds of agricultural waste, were also of interest due to increasing
concerns of their disposal (Sakar, S., et al, 2009; Demirer, G., et al, 2005), there are
more and more investigations of the AD process on them. The type of reactors used for
livestock manure waste treatment comprise: batch, continuous one stage, and
continuous two stage reactors, tubular reactors.
2.2.3 Factors influencing the anaerobic digestion process

AD can be sensitive to several operating factors, including pH, temperature, and
characteristics of the substrates (figure 2). To optimize the efficiency of AD, these
factors should be carefully regulated.

12


Substrate

Operating factors

pH

C/N ratio
AD processes

Temperature
VFAs

Alkalinity

Figure 2. Factors influencing AD performance
2.2.3.1 pH
pH fluctuation can effect biogas yield throughout AD. In the early stages such as
hydrolysis, acidogenenis, and acetogenesis, pH decrease due to the formation of
organic acids. Since the methanogenesis phase occurs, pH may increases slightly
because of the production of ammonia (Verma, S., 2002). Below pH 6, inhibition of
CH4-forming bacteria may occur, which can disrupt anaerobic process (Castro, H., et
al, 2002). The pH inside digesters is an important feature influencing the growth of
anaerobic microbes, especially methanogens, through its impact on enzyme activities.

This is because each group of microorganisms has its own appropriate pH for growth.
Methanogenis bacteria are seriously sensitive to pH and need a pH range from 6.5 to
7.8 (Sakar, S., et al, 2009) while acid forming bacteria can function in a wider range
between 4.0 and 8.5 (Hwang, M.H., et al, 2004) but prefer a pH of 5.5 to 6.5 (Ward,
A.J., et al, 2008; Khanal, S.K., 2009). In operation, it is necessary to keep pH close to
neutral since methanogenesis is the yeald-limiting step. Lime addition is a common
technique to overcome pH reduction.
13


2.2.3.2 Alkalinity
Alkalinity refers to the buffering capacity, which is important for regulating pH in AD.
Alkalinity originates from the degradation of organics in the form of CO2, bicarbonate
and ammonia (Hwang, M.H., et al, 2004). The equilibrium of CO2 and bicarbonate
will resist the dramatic changes in pH. Compared to pH, alkalinity or buffering
capacity gives more reliability for system stability because the possible accumulation
of VFAs can lead to a reduction in buffering capacity ans pH (Astls, S., et al, 2011).
The pH in an anaerobic system is adjusted by CO2 in gas phase, and bicarbonate in
liquid phase. Thereby, pH will decrease if there is a lack of bicarbonate and vice versa.
In practice, when pH of digester decreases a net strong base, either sodium hydroxide,
or calcium hydroxide (Saker, S., et al, 2009) or carbonate salts (Thanh, N., 2014), are
utilized. They are able to remove CO2 in the gas phase to convert into bicarbonate.
Bicarbonate can be directly added to reject the lag time and over organic dosing
(Ward, A.J., et al, 2008).
2.2.3.3 Temperature
AD strongly depends on temperature since it affects not only the physicochemical
properties of substrate in digesters, but also bacteria which is seriously sensitive with
any alterations in temperature. Thus, it is essential to maintain constant favorable
temperatures for the growth of anaerobic microbes (Castro, H., et al, 2002). Water
baths or passive solar heating are used for temperature maintenance; and heat can be

added by using heat exchanges in the recycled slurry or heating coils or steam
injection in the digester (Ostrem, K., 2004). Any fluctuation of temperature even small
change between 30 – 32oC (Ward, A.J., et al, 2008), may result in inactivation of
14


bacteria, leading to a decrease in biogas production. Furthermore, process failure can
be reported at temperature alteration in excess of 1oC per day (Appels, L., et al, 2008).
There are three temperature ranges investigated for applications: psychrophilic
temperature from 10 to 20oC, mesophilic temperature between 20 and 40oC, and
thermophilic temperature between 40 and 60oC (Sakar, S., et al, 2009). Since sufficient
retention time for CH4-foring bacteria is provided, anaerobic sludge digestion could be
operated successfully at psychrophilic temperature as low as 20oC (Ward, A.J., et al,
2008). The main different between mesophilic and thermophilic digestion is CH4
yield. It is studied that CH4 produced by thermophilic digestion is higher than that by
mesophilic digestion in a given digester because of the fact that high temperature is a
conducive condition for methanogens growth (Castro, H., et al, 2002; Burton, C.H.T.,
2003). Another advantage of thermophilic digestion is to facilitate the balanced
fermentation in producing biogas (Del Borghi, A., et al, 1999). The application of high
temperature, however, has some disadvantages, such as the increase of free ammonia
or VFAs, which easily results in inhibiting the process. (Appels, L., et al, 2008)
2.2.3.4 VFAs
VFAs created during AD are important product and relates to the imbalance of AD.
High VFA concentration principally leads to the process failures with respect to an
imbalance among acidogenic, acetogenic, and methanogenic organisms (Boe, K.,
2006). Additionally, less effective removal of COD is reported with increased VFA
production (Sakar, S., et al, 2009). In the acetogenic stage, the VFA accumulation will
result in pH decrease, which adversely impacts on the growth of methanogens. If
inhabitations occur in long time, acetogens will predominate in digesters. As
15



mentioned, the addition of buffering is an effective deal since this can resist pH drop
and maintain sufficient VFA concentration (Ward, A.J., et al, 2008). While acetic acid
is the key substrate for methanogenesis, it if defined that propionic and butyric acids
are inhibitory to methanogenic bacteria. So as to avoid process failure, mornitoring of
VFA has been studied to stabilize the overall system (Ward, A.J., et al, 2008).
2.2.3.5 Ammonia
The present of ammonia in digester results from breakdown of nitrogen-containing
matter, mainly from protein and urea. Ammonia is one of inhibitory substrates to the
AD process (Chen, Y., et al, 2008). Between two form of ammonia, NH4+ and free
ammonia NH3 in liquid, free ammonia is defined as the main cause of inhabitation.
The reason is hydrophobic form of ammonia could easily penetrate through cell walls,
causing pH imbalance and enzyme malfunction (Chen, Y., et al, 2008). This inhibition
is generally reported in the methanogenesis stage. High concentration of ammonia in
digester could affect aciddogenic populations while methanogenic population may lose
56% of its activity (Lettinga, G., et al, 1980). On basis of CH4 production, ammonia
has stronger impact on aceticlastic than hydrogenotrophic methanogens (Thanh, N.,
2014). It is suggested that free ammonia concentration should be kept below 80 mg/L,
meanwhile ammonium could reach up to 1500 mg/L without making any inhibition
(Burtton, C.H.T. 2003). pH and temperature are determined as factors affecting the
ammonia inhibition capacity through ammonia concentrations (Chen, Y., et al, 2008).
The higher pH is, the more the amount of ammonia is and the less the amount of
ammonium is.

16


2.2.4. Nutrient requirements for anaerobic digestion
A well-balanced anaerobic digestion system requires a mix of nutrients to sustain all

groups of anaerobic microbes. The elements that are essential required in substantial
amounts. Many essential nutrients, however, can become inhibitory or toxic to the
anaerobic digestion process when present in high concentrations.
Nutrient requirements for anaerobic processes are much lower than the requirements
for aerobic processes due to the lower cell yield from the degradation of equal
quantities of substrate. Apart from the carbon source, the two major nutrients in
anaerobic processes are nitrogen and phosphorus. These must be available in the
digester. In addition to the two major nutrients, other elements such as sulphur,
calcium, magnesium, although required to a lesser extent, also have important roles in
anaerobic microorganisms.
Table 2. Summarises the optimal concentration and effects of these nutrients in the
anaerobic digestion process

Nutrient

Ca

Mg

Na

S

Concentration
required (mg L-1)
100-200
(Mara, et al, 2003)
75-150
(Mara, et al, 2003)
100-200

(Mara, et al, 2003)
0.001-1.0
(Speece, 1996)

Effects on digestion

Essential for cell growth (Murray, et al, 1985)

Increase cell activity and facilitate granulation

Increase cell activity and facilitate granulation

Sulphur source for protein sythesis

17