1

MUHAMMAD AHSAN RAFI

0039-BH-BIO-T-19

COURSE NAME: BIOREMEDIATION

COURSE CODE: BIOTECH-3102

SUBMITTED TO: DR. HAMID MUKHTAR

SUBMISSION DATE: 30-08-2022

INSTITUTE OF INDUSTRIAL BIOTECHNOLOGY

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TABLE OF CONTENT

INTRODUCTION

FEEDSTOCK OF ANAEROBIC DEGRADATION

PRINCIPLES OF ANAEROBIC DEGRADATION

1. HYDROLYSIS
2. ACIDOGENESIS
3. ACETOGENESIS
4. METHANOGENESIS

IMPORTANT OPERATING FACTORS IN ANAEROBIC DEGRADATION OF ORGANIC COMPUNDS

A. TEMPERATURE
B. MOISTURE CONTENT
C. RETENTION TIME
D. PH

HOW TO ENHANCE ANAEROBIC DEGRADATION

• PHYSICAL TREATMENT
• CHEMICAL TREATMENT
• BIOLOGICAL TREATMENT
• THERMAL TREATMENT

ANAEROBIC DEGRADATION OF RURAL AREAS

CONCLUSION

REFRENCES
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Anaerobic Degradation of Organic Matter

Introduction:

Anaerobic degradation is a biochemical process of the breakdown of organic

content microorganisms without the availability of oxygen. This process results in the formation
of methane and carbon dioxide. Almost all of the materials high in organic content including
livestock waste, food waste, agricultural waste and Municipal waste goes through anaerobic
degradation. The resulting methane gas produced by this process can be collected and
consumed as a fuel source for heating production and cocking purposes. It can also be burned
to generate electricity which makes it one of the few renewable sources of energy.

Aerobic degradation, which uses molecular oxygen as the terminal electron acceptor, occurs for
the overwhelming majority of organic molecules created in nature or by human industry.
Oxygen is the go-to electron acceptor for natural microbial degradation processes as long as it
is present.

When compared to aerobic degradation, the rates and capabilities of anaerobic degradation
have traditionally been seen as subpar. Some kinds of substrates that are quite stable are
regarded to be particularly unsuitable for their sluggish and inefficient nature. However, in
certain anoxic settings like the cow's rumen, the turnover of, say, cellulose is considerably
quicker than in the presence of oxygen, with typical half-life periods in the range of one day.
Plant tissues rich in lignin, which generally resists destruction in the absence of oxygen,
represent the limit of the fermentative decomposition of fibers in the rumen.

Anaerobic processes have been shown to be effective and much cheaper than aerobic
treatment for waste management, particularly when dealing with large quantities of
biodegradable organic matter; these processes require only a negligible amount of energy input
in comparison to treatment in aeration basins, and they can generate a mixture of methane and
carbon dioxide ('biogas') that can be used for energy production. This is the case with most
types of trash that may be broken down without the presence of oxygen, including
polysaccharides, proteins, lipids, nucleic acids, etc. Certain extracellular enzymes catalyze the
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hydrolysis of these polymers into oligo- and monomers, which may then be destroyed in the
cell through enzyme processes analogous to those seen in aerobic metabolism. Transformation
rates per unit biomass should be comparable in aerobic and anaerobic cultures because the
specific activity of such enzymes are in the same range (0.1-1 mol substrate per min and mg cell
protein (Schink, 2005).

Feedstocks for Anaerobic Degradation:

Anaerobic degradation may use a variety of feedstocks,

including municipal and animal wastewaters, agricultural and food wastes, and other organic
materials. In addition to aerobic degradation (degradation with oxygen present) and
sedimentation, anaerobic degradation is a common treatment method for municipal
wastewater. Anaerobic degradation may lessen the quantity of solid waste created during
wastewater treatment, hence cutting down on disposal expenses. Animal waste contains the
potential to serve as a feedstock for anaerobic degradation, much like human waste.

More than 300 cattle, 600 pigs or sheep, or 30,000 chickens are often housed in concentrated
animal feeding operations (CAFOs) and confined feeding operations (CFOs) which are big
animal-feeding facilities. There may be serious consequences to the environment if manure and
wastewater from a farm with that many animals are simply allowed to wash off the land and
into storm drains and open bodies of water. As the trash breaks down in the water, it removes
oxygen, which may harm fish and other aquatic organisms. Water quality may be maintained if
the animal excrement is contained. Waste volume is reduced, usable methane is produced, and
a fertilizer-useable byproduct is obtained via anaerobic degradation.

Anaerobic degradation is not limited to the processing of animal waste, but may also be used
on agricultural plant waste. Bio gas facilities in Europe use energy crops produced specifically
for their anaerobic degradation processes. (The facility is considered a co-degradation plant if it
processes more than one kind of agricultural feedstock.) Feedstock for anaerobic degradation
might also include crops that have been damaged by insects or diseases.
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With the exception of woody wastes, most organic materials are amenable to anaerobic
degradation. Most anaerobic microbes have trouble breaking down lignin, a compound found
in wood. Anaerobes that are able to degrade cellulose for the production of ethanol from waste
wood have been the focus of biofuel industry research since the early 21st century. ("anaerobic
degradation | chemical process", 2022).

Principle of Anaerobic degradation:

The process of anaerobic degradation occurs in 4 series of

reactions including, methanogenesis, acetogenesis, acidogenesis and hydrolysis.

Figure 1 (Mechanism of degradation through organic matter)

Let’s discuss all these processes in detail:

1. Hydrolysis:
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Hydrolysis is the first step in the process of degradation causing the breaks down of the
complex organic molecules (lipids, proteins and carbohydrates) into their component
parts. Hydrolases are basically extracellular enzymes that catalyze hydrolysis processes.
Esterase, glycosidases, and peptidases are all examples of hydrolases that break down
different types of bonds, including those in lipids, carbohydrates, and proteins (enzymes
that cause the hydrolysis of peptide bond in proteins). Hydrolytic bacteria are
microorganisms responsible for the production of these enzymes. It has been stated
that common types of hydrolytic bacteria include Proteus Vulgaris, Clostridium,
streptococcus, bacillus, acetivibrio cellulolyiticus, vibrio, , Bacteroides, , micrococcus,
staphylococcus, and staphylococcus, micrococcus (Amani, Nosrati & Sreekrishnan,
2010).
The cellulase enzyme produced by the Cellulomonas bacterium can break down
polysaccharides into simple sugar; the protease enzyme produced by the Bacillus
bacterium can break down proteins into amino acids; and the lipase enzyme produced
by the Mycobacterium bacterium can break down lipids into fatty acids (Hobson, 1983).

2. Acidogenesis:
In this step, fermentation causing bacteria transform soluble compounds
generated in the hydrolysis stage into volatile fatty acids (propionate, b), alcohols, ,
carbon dioxide and lactate. Different bacterial species are responsible for each of the
many fermentation processes. A type of bacteria termed acetate-forming fermentative
bacteria also make acetate at this stage. Typical acetate-forming fermentative bacterial
species include Acetobacterium, Clostridium, Eubacterium, and Sporomusa (Li, Park &
Zhu, 2011).

3. Acetogenesis:
In acidogenesis step, fermentative bacteria causes the transformation of
soluble compounds being generated during hydrolysis into volatile fatty
acids(propionate, b) alcohol, lactate and carbon dioxide. Different bacterial species are
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responsible for each of the many fermentation processes. Bacterial species including
Saccharomyces (alcohol fermentation), Lactobacillus and Streptococcus (lactate
fermentation), Clostridium and Butyribacterium (butyrate fermentation),and
Clostridium (anaerobic degradation) are examples of genera that is responsible to carry
out fermentation pathways during anaerobic degradation(propionate fermentation). A
type of bacteria termed acetate-forming fermentative bacteria also make acetate at this
stage. Typical acetate-forming fermentative bacterial species include Acetobacterium,
Clostridium, Eubacterium, and Sporomusa.
4. Methanogenesis:
Methane gas is formed from acetate and molecular hydrogen in the last
step of anaerobic degradation, known as methanogenesis. As the last consumers of the
hydrogen created during acetogenesis, methanogens serve a crucial role in maintaining
the survival of the bacteria responsible for producing acetate.

Important operating factors in Anaerobic degradation of organic compounds:

The breakdown of organic matter by anaerobic degradation is a multi-step process

that requires a lot of time and energy. For each stage of degradation, the microorganisms
involved in the process may have distinct needs in terms of the surrounding environment.
Anaerobic degradation is sensitive to changes in moisture, temperature and retention duration,
organic loading rate, pH and carbon to nitrogen (C/N) ratio (Khalid, Arshad, Anjum, Mahmood &
Dawson, 2011).

1. Temperature:

Because it is necessary for the microorganisms' survival and development,

active temperature is the most crucial aspect in the determination of anaerobic degradation's
results. The rate of the microbial process is also established by the values of the primary kinetic
parameters involved in the process. Maximum gas generation, bacterial growth, and substrate
breakdown rates during anaerobic degradation occur in two temperature ranges mesophilic
(30-40°C) and thermophilic (50-60°C) (Mata-Alvarez, 2015).
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Biogas generation from a broad variety of waste has made widespread use of both mesophilic
and thermophilic anaerobic degradation, with the findings demonstrating that both methods
have distinct benefits and drawbacks, as shown below.

Figure 2: Comparison of mesophilic and thermophilic anaerobic

degradation

The anaerobic degradation of OMSW (organic municipal solid waste) under thermophilic (50°C)
and mesophilic (35°C) conditions yielded specific growth rate and methane yields, which
indicates that microbial activity is preferred operating at the thermophilic range of temperature
(Kim, Oh, Chun & Kim, 2006). Anaerobic degradation of food generated waste showed that
thermophilic digesters removed more soluble chemical oxygen demand (sCOD) and produced
more methane than mesophilic digesters Anaerobic degradation of the organic part of
municipal solid waste yields values for microbial activity (highest specific growth rate) and
methane generation (specific methane output).

2. Moisture Content:
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One of the most important aspects of anaerobic degradation is the amount of

moisture present. It has been suggested that moisture aids degradation by regulating turgidity
of cell; (ii) carrying nutrients, products, enzymes, intermediates and microbes; (iii) ability to
response to complex organic materials; and (iv) altering the enzymatic structure. Due to the
likelihood of water contents affecting process efficiency by dissolving easily degradable organic
matter, high moisture concentrations often aid the anaerobic degradation.

Low solids or wet degradation (containing less than 10% TS), medium solids or semidry (10-20%
TS), and high solids degradation (more than 20% TS) describe the three types of anaerobic
degradation processes. OSMW contains high solid content leading to the majority of research
on its degradation has focused on the dry anaerobic degradation process. To make OMSW
more suited for the semidry anaerobic degradation process, however, the moisture content
may be raised by adding water or by co-digesting with low content of solid wastes like manure
and sewage sludge(Fernández, Pérez & Romero, 2010).

In high-solids sludge degradation Methanogenic activity was enhanced by raising the starting
moisture availability of mesophilic anaerobic digesters from 96 to 90%. Different research
found that mesophilic anaerobic degradation of OMSW yielded a higher methane generation
rate and a greater DOC (dissolved organic carbon) removal efficiency when the digesters were
run at a higher starting moisture content. Methane generation in anaerobic digesters with
intermittent cycles of water addition and leachate drainage was observed to be reduced when
the moisture content of OMSW was increased. As water reductions into the bioreactors might
wash out of nutrients and microbes, those running at 80% moisture content showed a worse
volatile solids (VS) content than those running at 70% moisture content (Hernández-Berriel,
Márquez-Benavides, González-Pérez & Buenrostro-Delgado, 2008).

3. Retention Time:

Determining the retention time for a wastewater treatment plant involves

comparing the BOD (biochemical oxygen demand) and COD (chemical oxygen demand) of the
influent and the effluent. Longer retention duration will cause increased breakdown of organic
materials. The retention time necessary for full AD is governed by the technology used, the
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temperature of the process, and the makeup of the waste. The retention duration for
mesophilic anaerobic degradation is 8-40 days, whereas that for less thermophilic AD is shorter.
Dry thermophilic anaerobic degradation of OMSW was studied in relation to solids retention
time (SRT), with times ranging from 8 days to 40 days. The SRT of 15 days was found to achieve
the maximum VS elimination and methane output.

Lowering the retention value decreases the volume needed for the reactor and subsequently
minimizes the capital expenses of anaerobic process of degradation. (Forster-Carneiro, Pérez &
Romero, 2008).

Proper mixing guarantee that bacteria have immediate access to as many numbers of digestible
surface as they want and that environmental variables are uniform throughout the digester.
Recirculation of water and bio gas into the chamber to keep material flowing has been
employed as a potential mixing approach to increase anaerobic degradation. Reusing water in a
two-phase thermophilic anaerobic process with a 3-day hydraulic retention period was shown
to increase methane output. Decreasing solid content may lower the retention period, since
bacteria can more readily access liquid substrate and because the key processes need water. In
addition, a decreased solids concentration leads to better overall mixing. Retention time may
be decreased by splitting the phases of the degradation into different chambers such that the
bacterial population in each chamber is optimized for its function. From a kinetics perspective,
two-phase anaerobic degradation has proven useful for treating wastes like municipal solid
waste that have a high percentage of solids (Mata-Alvarez, 1987).

4. pH:

Methanogenic bacteria, their development, and methane generation are all sensitive to
acidic operating pH, therefore this is another critical consideration. The pH of anaerobic
digestate is altered by biological processes occurring at various stages. Since methanogens can't
survive in an environment when the pH is below 5, digester failure is possible during the
acetogenesis phase.
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Nutrient deficiency, poor mixing, Overloading temperature change, and loss of bacteria in the
effluent may all lead to elevated VFA levels. Buffering reagents may not be required if there is
sufficient alkalinity to neutralize the acids. However, ammonium ions may offer buffering when
they combine with bicarbonate ions to generate ammonium bicarbonate.

Organic acids are produced during the beginning of the process, when new waste is supplied,
but before the methanogenesis stage begins. Since this causes the pH to drop, it is the case.
This makes early-stage pH regulation particularly challenging. For a higher pH, buffers like lime
or calcium carbonate must be added to the system.

Anaerobic degradation is said to work best within a pH range of 6.8 and 7.2. They also said that
between pH 5.5 and 6.5 is excellent for hydrolysis and acidogenesis, and that pH 7.0 is optimal
for methanogenesis. This is a key consideration for those who favor two-stage procedures that
split hydrolysis/acidification from acetogenesis/methanogenesis. OMSW and manure anaerobic
codegradation performance may be improved by the recirculation of process liquid by
maintaining a constant pH, as has been observed.

Anaerobic degradation of food waste's initial step (hydrolysis and acidogenesis) was studied by
varying pH from 5 to 7 to 9 to 11. They discovered that adjusting the pH of the two-phase
anaerobic degradation of food scraps increased the rates of hydrolysis and acidogenesis, as well
as the elimination of TS and the formation of biogas (Zhang, Zhang, Zhang, Shi & Cai, 2005).

How to enhance anaerobic degradation:

A number of pretreatment techniques has been applied

to increase the rate of anaerobic degradation of organic method. This serves the basic purpose
of faster production of biogas. A few noticeable techniques among these are:
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Physical Pretreatment:

Mechanical pretreatment, or particle size reduction, is a time-honored

strategy for maximizing the effectiveness of the anaerobic degradation process. By providing
more accessible specific surface for the microorganisms, this treatment expedites the biological
process and enhances degradation. Also, the feedstock shouldn't be too big, otherwise the
digester can become clogged up. According to the literature, the methane output from the wet
anaerobic degradation system was unaffected by the particle size of the OMSW used. Further,
they found that the quick acidification brought on by the finest materials present in the dry
digesters led to the failure of the operation at high rates of loading of organic content.
Mechanical pretreatment has several benefits, including low energy use, low water usage, little
odor production, and improved dewaterability of the final anaerobic waste. The findings
demonstrated that the methane production might be improved by decreasing the particle size.
However, the author stated that the surface area does not significantly affect the kinetics of
degradation for the particle sizes evaluated. When the average particle size of food waste was
reduced from 0.888 to 0.718 mm by pretreatment through bead milling the methane
generation rate of the mesophilic anaerobic degradation process rose by a figure of 28%. The
scientists also noted that an increase in volatile fatty acid (FFA) buildup caused by substrate size
reduction led to a decrease in methane generation (Izumi et al., 2010).

Chemical Pretreatment:

Acidic pretreatment, alkaline pretreatment, and ozonation are all

chemical pretreatment processes used to break down organic molecules, which increases
biogas output and speeds up the hydrolysis rate. Using plant materials rich in lignin has been
said to benefit greatly from alkali treatment under anaerobic conditions. Torres et al. looked at
how lime addition (Ca(OH)2) influences anaerobic degradation of OMSW. Results showed that
pretreatment of alkaline enhanced anaerobic degradation by elevating soluble COD (by
enhancing the COD of solubilization). The result was a rise in methane production from 0.055 to
0.15 m3 CH4/kgVS.

Biological pretreatment:
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Biogas generation and VS reduction may be increased by using aerobic

and anaerobic processes before anaerobic degradation. The first phase of a two-phase AD
process is a biological type of pretreatment approach that eliminates the need for oxygen.
These systems have several benefits, such as it improved stability by providing better pH
management, (ii) a higher rate of loading (iii) increased specific activity of methanogens, leading
to a larger output of methane (iv) increased reduction in VS, and (v) a strong potential for
eliminating pathogens. Substrate digestibility is enhanced by the inclusion of microbial strains.
It has been shown that certain bacterial and fungal strains may increase gas generation by
increasing the activity of enzymes which is involved in cellulose breakdown (Yadvika, Santosh,
Sreekrishnan, Kohli & Rana, 2004).

Thermal pretreatment:

Pretreatment may also be accomplished by subjecting the material to

very high temperatures. Dissolving cell membranes is the primary consequence of heat
pretreatment, which allows for the release of organic molecules. Furthermore, pathogens are
eliminated, dewatering efficiency is increased, and the digestate's viscosity is decreased thanks
to the thermal pretreatment. Thermal pretreatment at 175°C increased the rate of methane
generation from sludge, kitchen trash, and fruit/vegetable waste, according to research. In
contrast, 8 and 12% reductions in methane generation were achieved with thermal
pretreatment of kitchen and fruit/vegetable waste, respectively. To improve anaerobic
degradation of OMSW, researchers heated the waste in microwave ovens at temperatures
ranging from 115 to 175 degrees Celsius. Pretreatment at 115 and 145°C resulted in a 4-7%
increase in biogas output, but pretreatment at 175°C resulted in a reduction owing to the
creation of refractory compounds, which inhibited the degradation (Liu, Wang, Gao, Zhou &
Shen, 2012).

Anaerobic degradation for rural areas:

Aerobic and anaerobic composting are advantageous

because of the need for soil conditioners and fertilizers in rural regions and the appeal of
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biofertilizer in comparison to artificial goods. Anaerobic degradation is the ideal alternative for
treating organic waste in both urban and rural regions because of its benefits, which include
lowering emission of greenhouse gas, killing pathogens, and producing sustainable energy. The
organic component of rural areas' solid waste comes from a variety of sources, including animal
feed by products and manure from animals. By modifying the C/N ratio and the moisture
content, as previously discussed, co-degradation becomes a real option, which is great for
boosting biogas production. Also, the methane output from anaerobic degradation may be
increased by using "energy crops" (such as maize, grass, and grains) as substrate. The use of
animal manure in small-scale or household biogas plants allows for anaerobic degradation to
occur in rural areas. Reportedly, the use of biogas in rural homes has helped with agricultural
structural adjustment, increased rural incomes, improved rural ecological conditions, and
resulted in higher quality of living and agricultural outputs. Central anaerobic degradation (CAD)
is the second option for anaerobic degradation in rural settings. In CAD, several farms work
together to feed a single big degradation plant with a wide range of substrates. The biogas
generation of CAD plants might be maximized by co-degradation, which would have far-
reaching effects for the local population. Typically constructed underground, small-scale
residential digesters create biogas that is utilized mostly in the kitchen. It is in China and India
that you will find them utilized the most. (Chen, Yang, Sweeney & Feng, 2010).

Conclusion:

Anaerobic degradation is often regarded as the most effective method for

processing the organic component of municipal trash at present. The environment, energy, and
economy may all benefit from this technology.

There are several steps involved in this biological process. Biogas is produced from the organic
part of municipal solid waste via a series of steps, each of which is dependent on a unique set of
microbes. There are a variety of environmental parameters, including moisture, organic loading
rate, temperature, pH, and carbon/nitrogen ratio, that affect the efficacy of the process as a
whole. It has been hypothesized that a C/N ratio of 25-30 for the feedstock would provide the
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most efficiency. Co-degradation of OMSW and other organic wastes may be used to enhance
nutritional value and carbon to nitrogen ratios.

In addition to the requirement for biofertilizer, the availability of various waste products (which
may be co-digested) in rural regions also makes anaerobic degradation interesting. Individual
homes equipped with digesters or a community-wide anaerobic degradation facility are both
viable options for producing biogas.
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https://www.britannica.com/science/anaerobic-degradation

2. Schink, B. (2005). Principles of Anaerobic Degradation of Organic

Compounds. Environmental Biotechnology, 229-257.

3. Amani, T., Nosrati, M., & Sreekrishnan, T. (2010). Anaerobic degradation from the
viewpoint of microbiological, chemical, and operational aspects — a
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4. Hobson, P. (1983). Anaerobic digesters. (A code of practice on safety In and around

anaerobic digesters). Edition 1. Agricultural Wastes, 6(4), 250-251.

5. Li, Y., Park, S., & Zhu, J. (2011). Solid-state anaerobic degradation for methane
production from organic waste. Renewable And Sustainable Energy Reviews, 15(1), 821-
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6. Khalid, A., Arshad, M., Anjum, M., Mahmood, T., & Dawson, L. (2011). The anaerobic
degradation of solid organic waste. Waste Management, 31(8), 1737-1744.

7. Mata-Alvarez, J. (2015). Biomethanization of the Organic Fraction of Municipal Solid

Wastes. Water Intelligence Online,
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8. Kim, J., Oh, B., Chun, Y., & Kim, S. (2006). Effects of temperature and hydraulic retention
time on anaerobic degradation of food waste. Journal Of Bioscience And
Bioengineering, 102(4), 328-332.

9. Fernández, J., Pérez, M., & Romero, L. (2010). Kinetics of mesophilic anaerobic
degradation of the organic fraction of municipal solid waste: Influence of initial total
solid concentration. Bioresource Technology, 101(16), 6322-6328.

10. Hernández-Berriel, M., Márquez-Benavides, L., González-Pérez, D., & Buenrostro-

Delgado, O. (2008). The effect of moisture regimes on the anaerobic degradation of
municipal solid waste from Metepec (México). Waste Management, 28, S14-S20.

11. Forster-Carneiro, T., Pérez, M., & Romero, L. (2008). Anaerobic degradation of municipal
solid wastes: Dry thermophilic performance. Bioresource Technology, 99(17), 8180-
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12. Mata-Alvarez, J. (1987). A dynamic simulation of a two-phase anaerobic degradation

system for solid wastes. Biotechnology And Bioengineering, 30(7), 844-851.
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Hydrolysis and Acidogenesis of Kitchen Wastes in Two-phase Anaerobic
Degradation. Environmental Technology, 26(3), 329-340.

14. Izumi, K., Okishio, Y., Nagao, N., Niwa, C., Yamamoto, S., & Toda, T. (2010). Effects of
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15. Yadvika, Santosh, Sreekrishnan, T., Kohli, S., & Rana, V. (2004). Enhancement of biogas
production from solid substrates using different techniques––a review. Bioresource
Technology, 95(1), 1-10.

16. Liu, X., Wang, W., Gao, X., Zhou, Y., & Shen, R. (2012). Effect of thermal pretreatment on
the physical and chemical properties of municipal biomass waste. Waste
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17. Chen, Y., Yang, G., Sweeney, S., & Feng, Y. (2010). Household biogas use in rural China: A
study of opportunities and constraints. Renewable And Sustainable Energy
Reviews, 14(1), 545-549.