FACULTY OF ENGINEERING

DEPARTMENT OF CHEMICAL AND ENVIRONMENTAL ENGINEERING

UNIVERSITI PUTRA MALAYSIA

ECH4202 BIOREACTOR ENGINEERING DESIGN

ASSIGNMENT 1

ROTATING OR STIRRED-DRUM SOLID STATE BIOREACTOR

LECTURER: DR HALIMATUN SAKDIAH BINTI ZAINUDDIN

PREPARED BY GROUP 6:

SANGITHA A/P BATHUMALY 203691

NURUL FARIHA BINTI ISMAIL 201110

DATE SUBMITTED: 20/4/2023

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 1.0 Introduction

1.1 Type of Solid State Fermentation Bioreactors

Solid state fermentation (SSF) is a fermentation where the culture is introduced

into the solid substrate, and cultivation is often done in a controlled environment with
regulated, light, humidity, and temperature. It also includes controllable factors such as
nutrient levels, feedstock-to-inoculum ratios, C/N ratios, and pH. This fermentation's
characteristic is that it occurs in the absence or almost complete utter lack of free water
(Steudler et al., 2019). Due to the high demand of biologically active secondary
metabolites like pharmaceutical products, enzymes and antibiotics, SSF has become an
essential alternative to submerged fermentation (SmF) (Pandey et al., 2001; Thomas et
al., 2013).

According to Solid State Fermentation - Tech4Biowaste (2023), depending on

the aeration and mixing type, the design of SSF bioreactors can be categorized into four
groups. Group 1 consists of bioreactors with unforced aeration and no mixing or
agitation (static). Group 2 includes bioreactors with forced aeration and no mixing
(static). Group 3 includes bioreactors with unforced aeration and continuous or
intermittent mixing/agitation. Finally, Group 4 includes bioreactors with forced aeration
and continuous or intermittent mixing/agitation. However, this report will focus on
rotating or stirred-drum bioreactor which is categorised in Group 3. Figure 1 below
shows the basic design features of the various SSF bioreactors.

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 Figure 1.1: Basic design features of varioud type of SSF bioreactors depend on the
mixing and aeration (Mitchell et al., 2002)

1.2 General Design of Rotating or Stirred-drum Solid State Bioreactor

Arora et al. (2018) mentioned that the rotating drum bioreactor is composed of a
horizontal drum-shaped container that can be mounted on a roller for rotation. The
headspace, drum wall, and substrate are its usual three components. The headspace
above the substrate bed, which is tossed as the bioreactor rotates irregularly or
constantly, is blown with air. The most common method for mixing the substrate bed is
for the drum to rotate around its axis (Figure 1.2a), although stirred drum bioreactors
may also be employed, where mixing is facilitated by a paddle mounted on a central
shaft when the drum is stationary (Figure 1.2b). In order to aid in mixing, the drum may
feature lifters or baffles that are internally screwed and come in a variety of sizes
(Figure 1.2c).

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 Figure 1.2: a) schematic diagram of rotating drum bioreactor, b. Stirred drum
bioreactor, c. Cross section of RDB showing arrangement of baffles (Arora et al., 2018)

In terms of novelty and innovation, the internally screwed baffles or lifters, which
are used in rotating or stirred drum solid state fermentation bioreactors to encourage
mixing of the substrate bed, are new designs in this bioreactor that are not present in
other bioreactors. The use of a roller for rotation, which enables occasional mixing
without forced aeration, is another distinctive design feature of these bioreactors. A
paddle positioned on a central shaft that mixes the substrate when the drum is still can
also be found in stirred drum bioreactors. Conventional bioreactors for liquid
fermentation do not frequently utilise these design elements.

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 2.0 Mechanism and Detailed Purpose

2.1 Specific Purpose of Rotating or Stirred Drum SSF Bioreactor

The purpose of a SSF bioreactor is to create an optimal environment for the

growth and biological activity of microorganisms. It is necessary for bioreactors to have
the ability to contain the media and maintain a secure seal to prevent harmful
substances from entering. Through the circular movement of the vessel, rotating drum
bioreactors enable aeration and heat removal. This improved aeration and mixing
encourage microbial development, which raises metabolic heat (Robinson & Nigam,
2003).

2.2 Mechanism of Rotating or Stirred Drum SSF Bioreactor

Rotating drum bioreactors use moderate agitation to optimize mass and heat
transfer and encourage microbial growth. Convective transport is facilitated by mixing
because it increases the surface area of the substrate exposed to moist air or cooling
fluid (Arora et al., 2018). The substrate acts as a support matrix for the microorganisms.
Generally, the substrate bed partially fills the bioreactor, and in order to ensure optimal
oxygen and carbon dioxide transfer, the height of the fermented bed is recommended to
be moderate. For temperature regulation, the solid substrate's mixing action is essential
(Solid State Fermentation - Tech4Biowaste, 2023).

As previously mentioned, mixing in rotating or stirred drum SSF bioreactors is

achieved by blowing air through the headspace above the substrate bed, which
provides forced aeration and promotes heat and mass transfer and bacterial growth.
The rotation or stirring of the drum facilitates the mixing of the substrate bed, which
improved the substrate’s surface area that is exposed to cooling water or humid air,
leading to enhanced convective transport. The use of internally screwed baffles or lifters
inside the drum further facilitates mixing of the substrate bed.

On the other hand, for sterilization of this bioreactor, high-pressure steam

injection can be used for in situ sterilization, and this process can be carried out in a
closed system. Besides, Sánchez et al. (2006) reported that the manufacture of

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 amylase by Aspergillus oryzae on wheat bran and the manufacturing of penicillin on a
big scale both utilized rotation at 24 rpm and direct injection of steam at a pressure of 1
atm above ambient pressure.

In terms of other parameters like scalability, rotating drum bioreactors typically

operate with a working volume of 30% from the total reactor volume. However, scaling
up the bioreactor may not be a practical option. Large bioreactor requires intensive
maintenance compared to regular rotating drum bioreactor.

2.3 Commercialization of Rotating or Stirred Drum SSF Bioreactor

According to Prado Barragán et al. (2016) rotating drum bioreactors were initially
employed in the early 1900s to produce amylase by Aspergillus oryzae in SSF with
wheat bran as a substrate. Subsequently, in the 1940s, advancements were made in
the design of rotating drum bioreactors, leading to their application in commercial-scale
penicillin production. Since then, technological enhancements have been made to this
type of bioreactor. Table 2.1 depicts the usage of rotating drum bioreactor for various
products.

Table 2.1: Examples of SSF bioreactors used for enzymes and chemical products

Microorganism Production
Substrate Product References
used level
Soybean meal Amylase and Aspergillus 85000- (Sukumprasertsri,
Protease oryzae 110000 U g- 2013)
ds−1
Palm oil Cellulase Trichoderma 8.2 FPA g-ds−1 (Alam et al.,
lignocellulosic harzanium 2009)
biomass
(Rodríguez-
Autohydrolyzed
Fucoidanase Mucor sp. 3P 9.62 U L−1 Jasso et al.,
algae
2013)

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Apple pomace Citric acid Aspergillus niger 220.6 g kg-ds−1 (Dhillon et al.,
2011)
Sugarcane Ethanol Kluveromyces 24 g L−1 (Lin et al., 2013)
bagasse marxianus
Rice by- Ethanol Aspergillus niger 11.7 g L−1 (Rocha et al.,
product, whey 2013)
and sugarcane
bagasse

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 3.0 Design Diagram

3.1 Process Flow Diagram

Based on research by Díaz et al. (2009), the rotating drum bioreactor used for
the production of hydrolytic enzymes consists of a glass roller bottle with a volume of
250 mL and a diameter of 7 cm, which is connected to a filtered air supply (shown in
Figure 3.1). A 0.45 m cellulose filter is used to sterilize the air after the air flow rate is
determined using a rotameter. The humidifier process utilizes a glass column filled with
sterilized distilled water and 3 mm glass beads. A syringe connected to a barbed wire
introduces air into the roller bottle in order to flush out any solids that may have become
lodged there.

Figure 3.1: Laboratory scale rotating drum bioreactor (Díaz et al., 2009)

Besides, a study by E.-Q. Wang et al. (2010) utilized rotating drum bioreactor for
ethanol production from sweet sorghum stalks. Utilizing batch fermentation studies,
sweet sorghum stalks were converted into fuel ethanol using yeast TSH-SC-1 in a 5 m3
spinning stainless steel drum. The drum rotated for 20 minutes every five hours, the
trials were run for a hold period of 48 hours. Finally, the samples were collected for cell,
sugar, and ethanol assays. The rotating drum bioreactor is used to turn sugar into
ethanol, and the steaming bucket unit is used to extract both ethanol and water from the
solid substrate (see Figure 3.2 below). The process starts with a belt conveyor (1) which
feeds the disintegrator (2). The disintegrated material is then transferred to the seeding
tank (3). The rotating drum bioreactor (RBD) (4) is where the fermentation takes place,

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with gas outlet (5) for exhaust gases. Steam is injected into the RDB through a
steaming bucket (6) with vapor (7) released through a heat exchanger (8). The ethanol-
water mixture is then transferred to an ethanol dehydration unit (9), while the solid
residues are sent to a waste treatment or downstream process (10).

Figure 3.2: Schematic process design for the process

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 4.0 Organism, Substrate and Products

4.1 Type of organism grow within stirred-drum solid state bioreactor

The organism that grow in rotating or stirred-drum solid state bioreactor needs to
fulfill several criteria so that they will be able to grow there. The microorganism must be
able to use the particular substrate that is offered in the bioreactor. Depending on the
application, the substrate may change, however lignocellulosic materials, agricultural
wastes, and other organic wastes are often used substrates (Li et al., 2011).

The water activity level offered in the bioreactor should allow the microorganism
to proliferate. In stirred-drum solid state bioreactors, water activity (aw) is a crucial
parameter to take into account since it influences microbial growth, metabolism, and
product generation (Antolli & Liu, 2012). The ratio of the vapour pressure of the water in
a sample to the vapour pressure of pure water at the same temperature and pressure is
known as the water activity (Herrington & Vernier, n.d.). By altering the moisture content
of the solid substrate, it is possible to regulate the water activity level in stirred-drum
solid state bioreactors.

Depending on the microbe and the substrate, different solid state bioreactors
have different ideal water activity levels for microbial development. While certain fungi
may grow at lower water activity levels of 0.6 to 0.7, most microorganisms generally
need a minimum water activity level of 0.85 to flourish (Brown, n.d.). Nevertheless, too
much moisture might result in inadequate aeration and more competition amongst
microorganisms, which lowers the yield of the final product.

Water activity indicators such as monitoring relative humidity inside the

bioreactor, measuring the water content of the solid substrate, and other methods can
be used to monitor and manage the water activity level (Krishna, 2005). Depending on
the desired water activity level, either more water may be added or the solid substrate
can be dried to change the water activity level. Overall, stirring-drum solid state
bioreactors require a certain degree of water activity to sustain optimal microbial growth
and product generation.

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 Next, the microbe must to be able to develop within the bioreactor's pH range. By
adding buffers or changing the culture medium, the pH may be managed. Depending on
the microbe and the substrate, there are several pH ranges where microbial growth is
best suited in solid state bioreactors. Using tools like pH electrodes, colorimetric
indicators, or titration, the pH of the bioreactor may be monitored and managed. By
adding acids or bases to the culture medium, or by utilising buffer solutions, the pH can
be changed (Shipman & Dziewiatkowski, n.d.). For optimum microbial growth and
product generation, the ideal pH range must be maintained. Microbial growth may be
restricted and product yields may decline if the pH is too low or high. pH can also alter
enzyme stability and activity, which can have an impact on product yields.

To maximise microbial growth and product generation, the pH range in stirred-

drum solid state bioreactors needs to be closely watched and managed. To create the
best possible circumstances for the microorganisms, the pH range may need to be
modified during the fermentation process.

Furthermore, in stirred-drum solid state bioreactors, temperature is a crucial

factor because it influences microbial growth, metabolism, and product synthesis
(Prabhu et al., 2022). Depending on the microbe and the substrate, there are several
temperature ranges that are ideal for microbial development in solid state bioreactors.

Thermocouples, infrared thermometers, and temperature probes are just a few of

the tools that may be used to measure and regulate the temperature inside the
bioreactor. By controlling the environment inside the bioreactor, such as changing the
agitation speed or applying external heating or cooling, the temperature may be
changed. For optimal microbial growth and product generation, the ideal temperature
range must be maintained. A temperature that is too low may prevent microbial
development and lower product output. Too much heat can hinder microbial
development and cause the germs to die or create undesired byproducts (Gregg et al.,
2009).

In order to maximise microbial growth and product generation, the temperature

range in stirred-drum solid state bioreactors needs to be closely monitored and

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managed. To provide the best environment for the microorganisms during the
fermentation, the temperature range may need to be modified.

Moreover, as oxygen is needed for respiration and energy generation, it is crucial

for microbial development in stirred-drum solid state bioreactors. Microbial growth,
metabolism, and product synthesis can all be strongly impacted by the oxygen levels in
the bioreactor (S.-J. Wang & Zhong, 2007). In stirred-drum solid state bioreactors,
oxygen can be delivered to the bacteria through aeration, agitation, or a mixture of both.

Depending on the microbe and the substrate, different solid state bioreactors
have different ideal oxygen needs for microbial development. In general, certain
anaerobic microbes may thrive in environments with little or no oxygen. Oxygen sensors
or mass spectrometry are only two examples of the different methods that may be used
to monitor and regulate the oxygen content in the bioreactor. By altering the bioreactor's
architecture to improve oxygen transmission, agitation speed, aeration rate, or oxygen
concentration may all be changed (Mete et al., 2012).

For microbial growth and product synthesis to be maximised, the ideal oxygen
level must be maintained. Inadequate oxygen can result in poor development, but too
much oxygen can produce oxidative stress and limit growth. In order to maximise
microbial growth and product generation, the oxygen content in stirred-drum solid state
bioreactors must be carefully monitored and managed.

In order to achieve optimal development and product generation, it is important to

take into account a number of parameters while choosing the microorganisms to
cultivate in stirred-drum solid state bioreactors.

4.2 List of organism grow within stirred-drum solid state bioreactor

A variety of microorganisms have been grown in stirred-drum solid state

bioreactors for the manufacture of a variety of goods. Stirred-drum solid state
bioreactors have been used to cultivate a variety of microorganisms, including the
following:

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 1. Filamentous fungi, such as Aspergillus niger, Aspergillus oryzae, Trichoderma
reesei, and Penicillium chrysogenum, for the production of enzymes, antibiotics,
and organic acids.
2. Yeasts, such as Saccharomyces cerevisiae, Candida rugosa, and Candida utilis,
for the production of biofuels, organic acids, and enzymes.
3. Bacteria, such as Bacillus subtilis, Pseudomonas aeruginosa, and Escherichia
coli, for the production of enzymes, biofuels, and biopolymers.
4. Actinomycetes, such as Streptomyces griseus, for the production of antibiotics
and enzymes.
5. Algae, such as Chlorella vulgaris and Spirulina platensis, for the production of
biofuels, pigments, and nutraceuticals.
6. Protozoa, such as Tetrahymena thermophila, for the production of proteins and
enzymes.

4.3 Fermentation conditions in stirred-drum solid state bioreactor

The type of microorganism and the product being generated can affect the
fermentation conditions in stirred-drum solid state bioreactors. Nevertheless, some
typical fermentation conditions that are commonly managed and observed in stirred-
drum solid state bioreactors include:

1. Temperature: Depending on the microbe and the substrate, the ideal

temperature range for microbial growth and product synthesis varies. The typical
temperature range is 25°C to 45°C.
2. pH: Depending on the microbe and the substrate, the pH range for microbial
growth and product synthesis varies. The pH range is typically kept between 4.5
and 8.0.
3. Moisture content: Depending on the microbe and the substrate, the ideal
moisture content for microbial growth and product synthesis varies. The moisture
level is typically kept between 50 and 80%.
4. Oxygen concentration: Depending on the microbe and the substrate, the ideal
oxygen concentration for microbial growth and product synthesis varies. Aeration

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 or agitation is typically used to keep the oxygen concentration between 2-10%
(v/v).
5. Nutrient availability: The availability and concentration of nutrients, such as
carbon, nitrogen, and minerals, can have a big impact on how quickly microbes
grow and what kinds of products they produce. Based on the microorganism and
the substrate, the nutrient concentration must be tuned.
6. Agitation and mixing: By agitating and mixing the solid substrate, it is possible to
increase oxygen transmission, nutrient availability, and temperature distribution,
all of which will promote microbial growth and the production of more effective
products.

4.4 Production capacity in stirred-drum solid state bioreactor

The size and design of the bioreactor, the type of microbe and substrate utilised,
the fermentation conditions, and the product being generated are all variables that might
affect the production capacity of stirred-drum solid state bioreactors. Compared to other
types of bioreactors like stirred-tank reactors, stirred-drum solid state bioreactors
typically have lower output capacity. Depending on the bioreactor design and the
process parameters, the production capabilities of stirred-drum solid state bioreactors
can range from a few grammes to several kilos (Ashok et al., 2017).

The type of product being produced might also have an impact on the production
capacity. For instance, compared to processes that create organic acids, biopolymers,
or biofuels, the output capacity of solid-state fermentation systems that produce
enzymes can be rather high. It is important to note that the possibility for low-cost
production and simplicity of scaling-up are the key benefits of solid-state fermentation
processes in stirred-drum bioreactors (Chilakamarry et al., 2022). These bioreactors are
a practical choice for small-scale production or the creation of new goods due to their
cheap capital costs. Other bioreactor types, including stirred-tank reactors or airlift
reactors, can be preferable if bigger production capacities are needed.

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 5.0 Advantages and Limitation

5.1 Advantages of stirred-drum solid state bioreactor

Compared to other types of bioreactors, stirred-drum solid state bioreactors

provide a number of benefits for microbial fermentation operations. Compared to other
types of bioreactors like stirred-tank reactors or airlift reactors, stirred-drum solid state
bioreactors are very easy to design and build, which results in cheaper capital
expenditures (Hansen et al., 2015). Moreover, stirred-drum bioreactors' solid-state
fermentation process doesn't need large quantities of liquid medium or complicated
apparatus, which might lead to cheaper operating costs. When compared to submerged
fermentation processes, the solid substrate in stirred-drum bioreactors can provide a
higher concentration of nutrients and promote better microbial growth, which can result
in higher volumetric productivities.

Due to the concentration of nutrients in the solid substrate and the existence of
suitable environmental conditions for microbial growth, solid-state fermentation
processes in stirred-drum bioreactors can provide high product yields. As contrast to
submerged fermentation processes, the solid substrate utilised in stirred-drum solid
state bioreactors can operate as a physical barrier against contamination from rival
bacteria (Arora et al., 2018). Stirred-drum solid state bioreactors are appropriate for
industrial production because they can be readily scaled up by expanding the reactor
without significantly altering the fermentation process. In comparison to submerged
fermentation operations, solid-state fermentation processes in stirred-drum bioreactors
produce less effluent and use less energy, making them more ecologically friendly
(Abdul Manan, 2014). Overall, stirred-drum solid state bioreactors are a potential choice
for the manufacture of a variety of products, including enzymes, organic acids, biofuels,
and biopolymers, and they provide various benefits for microbial fermentation processes.

5.2 Disadvantages of stirred-drum solid state bioreactor

Using stirred-drum solid state bioreactors for microbial fermentation has a

number of benefits, but there are also some possible drawbacks to take into account.
The restricted oxygen transfer is one of the main drawbacks of solid-state fermentation

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in stirred-drum bioreactors, which can lead to poorer product yields and slower
fermentation rates when compared to submerged fermentation procedures (Mitchell et
al., 2002). Stirred-drum solid state bioreactors employ solid substrates, which can make
it difficult to regulate process variables like temperature, pH, and water activity, which
can impact fermentation success. The physical and chemical characteristics of solid
substrates can vary widely, which may have an impact on microbial development and
product generation. It may be challenging to optimise the fermentation process and
obtain consistent product quality due to this unpredictability.

Due to the solid substrate's opaque nature, solid-state fermentation in stirred-

drum bioreactors can be tricky to monitor in real-time, making it difficult to see changes
in the fermentation process or spot possible problems. It might be difficult to separate
the product from the solid substrate because it may be firmly bonded to it, which results
in low yields for product recovery and purification (Rudakiya, 2019). Although solid-state
fermentation in stirred-drum bioreactors is appropriate for the production of some
products, including organic acids and enzymes, it may not be appropriate for the
production of other products, such as therapeutic proteins, which necessitate exact
control over the fermentation environment. Overall, there are a number of possible
drawbacks to using stirred-drum solid state bioreactors for microbial fermentation
processes that should be taken into account when choosing a bioreactor for a particular
application.

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 Question 2

C* = 7.3 mg/L

Time (min) DO (mg/L)

Air switch OFF -1 3.3
0 3.3
1 2.4
2 1.3
3 0.3
4 0.1
5 0.0
Air switch ON 6 0.0
7 0.3
8 1.0
9 1.6
10 2.0
11 2.4
12 2.7
13 2.9
14 3.0
15 3.1
16 3.2
17 3.2

3.5

2.5
DO (mg/L)

1.5

0.5

0
-5 0 5 10 15 20
-0.5
Time (min)
Time (min) CL (mg/L)
0 3.3
1 2.4
2 1.3
3 0.3

3.5

2.5
CL (mg/L)

y = -1.01x + 3.34
1.5
R² = 0.9986

0.5

0
0 0.5 1 1.5 2 2.5 3 3.5
Time (min)
Time (min) CL (mg/L)
8 1.0
9 1.6
10 2.0
11 2.4
12 2.7
13 2.9
14 3.0
15 3.1

3.5

3.0

2.5
CL (mg/L)

2.0

1.5

1.0
y = 0.0013x3 - 0.0846x2 + 1.728x - 8.0461
0.5
R² = 0.9993
0.0
0 2 4 6 8 10 12 14 16
Time (min)
t (min) CL (mg/L) dCL/dt
8 1.0 0.62336
9 1.6 0.52038
10 2.0 0.4252
11 2.4 0.33782
12 2.7 0.25824
13 2.9 0.18646
14 3.0 0.12248
15 3.1 0.0663

0.7

0.6

0.5

0.4
dCL/dt

0.3

0.2

0.1 y = -0.2577x + 0.9198

R² = 0.9688
0
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5
CL (mg/L)
CamScanner