OBJECTIVES

The objectives of this experiment are to study the dynamic simulation of fed-batch

penicillin production by using a web-based simulator. Besides, the objective of this experiment is

to study the effect of aeration rate, agitator power, substrate feed rate, dissolved oxygen

concentration, substrate concentration, biomass concentration and Penicillin concentration over

time.
INTRODUCTION

Fermentation is a technique used for the production of penicillin. There are three types of

fermentation process such as batch fermentation, continuous fermentation and fed-batch

fermentation. For the production of penicillin, commercially, the fed-batch fermentation was

used. There are several factors need to be consider for the optimal design of fed-batch cultivation

such as substrate concentration in the feed, the rate of addition, the time to start the fresh nutrient

feed and when to finish the nutrient feeding so that the highest concentration of product is

produced and no unconverted substrate. All these factor could lead to the effect of substrate

concentration, biomass concentration, product concentration and others. These can be determine

by a web-based simulator such as Pensim simulator. This Pensim simulator will come out with

several graphs which can be examined the pattern of lines. Therefore, by key in the simulation

condition such as substrate concentration and biomass concentration, this web-based will directly

shows the results in graphical method.

From the results or graphs obtained, the effect of parameters can be examined and

explained by the penicillin production. All these effect such as aeration rate, agitator power,

substrate feed rate, dissolved oxygen concentration, substrate concentration, biomass

concentration and penicillin concentration over time can be evaluated on the efficiency of fed-

batch condition.
METHODOLOGY

This experiment was simulated the fed-batch penicillin production by using PenSim

simulator. The supplied default values were used for the simulation conditions except for

substrate and biomass concentration as shown in Table 1.0. Then, the output data was observed

and examined such as aeration rate, agitation power, substrate feed rate, dissolved oxygen

concentration, substrate concentration, biomass concentration and penicillin concentration.

Substrate concentration 5 50 15 15
Biomass concentration 0.1 0.1 10 20
Table 1.0: Simulation condition
RESULTS

Aeration Rate:

Figure 1.0: The graph of aeration rate against time for simulation condition of substrate

concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1 g/L (right)

respectively.

Figure 2.0: The graph of aeration rate against time for simulation condition of substrate

concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and 20 g/L (right)

respectively.
Agitation Power:

Figure 3.0: The graph of agitation power against time for simulation condition of substrate

concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1 g/L (right)

respectively.

Figure 4.0: The graph

of

agitation power against time for

simulation condition of substrate concentration and biomass concentration of 15 g/L and 10 g/L

(left) and 15 g/L and 20 g/L (right) respectively.

Dissolved Oxygen Concentration:

Figure 5.0: The graph of dissolved oxygen concentration against time for simulation condition

of substrate concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and

0.1 g/L (right) respectively.

Figure 6.0: The graph of dissolved oxygen concentration against time for simulation condition

of substrate concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and

20 g/L (right) respectively.

Substrate Concentration:

Figure 7.0: The graph of substrate concentration against time for simulation condition of

substrate concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1

g/L (right) respectively.

Figure 8.0: The graph of substrate concentration against time for simulation condition of

substrate concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and 20

g/L (right) respectively.

Biomass Concentration:

Figure 9.0: The graph of biomass concentration against time for simulation condition of

substrate concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1

g/L (right) respectively.

Figure 10.0: The graph of biomass concentration against time for simulation condition of

substrate concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and 20

g/L (right) respectively.

Penicillin Concentration:

Figure 11.0: The graph of penicillin concentration against time for simulation condition of

substrate concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1

g/L (right) respectively.

Figure 12.0: The graph of penicillin concentration against time for simulation condition of

substrate concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and 20

g/L (right) respectively.

Substrate Feed Rate:

Figure 13.0: The graph of substrate feed rate against time for simulation condition of substrate

concentration and biomass concentration of 5 g/L and 0.1 g/L (left) and 50 g/L and 0.1 g/L (right)

respectively.

Figure 14.0: The graph of substrate feed rate against time for simulation condition of substrate

concentration and biomass concentration of 15 g/L and 10 g/L (left) and 15 g/L and 20 g/L (right)

respectively.
DISCUSSION

Based on this experiment, the simulation condition was set up into two types which are

the different value for substrate concentration and the similar value for biomass concentration,

and the similar value for substrate concentration and the different value for biomass

concentration. For this experiment, the production of penicillin was using a fed-batch condition.

Then, a series of graph produced were examined and identify. For the aeration rate as shown in

Figure 1.0, Aeration rate is the rate of air being transfer or circulated through the liquid or

substance. As the substrate concentration was 5 g/L, the aeration rate was slightly decreased

before reached 50 hours while as the substrate concentration was 50 g/L, the aeration rate was

increased before reached 50 hours. Based on the pattern of the graph, it shows unstable

increasing and decreasing with the time. Comparing the aeration rate for the process when both

of the substrate concentration is 15 g/L while the biomass concentration are 10 g/L and 20 g/L,

the comparison were shown in Figure 2.0. The aeration rate is directly affected by the

concentration of the materials and products in the vessel in a process. The graph shown that the

highest aeration rate between the two is recorded at 8.623 L when the concentration of the

biomass is 20 g/L, while the lowest aeration rate between the two is recorded at 8.573 when the

concentration of biomass is 10 g/L. For graph from left (substrate concentration is 15 g/L and

biomass of 10 g/L), the highest peak recorded is at 100 hour and the lowest peak at 60 hour. For

graph from right (substrate concentration of 15 g/L and biomass of 20 g/L) the highest peaks is

recorded at the 30 hour, while the lowest reading is recorded at the 180 hour.

Besides, for the agitator power, agitator power is the power needed to mix all the

materials contained inside the vessel so that the product can be produced. The trend of the graph
was a bit similar for both with different substrate concentration condition which at 150 hours as

shown in Figure 3.0, the agitator power slightly drop and slow the motor rotation. But, increasing

suddenly when reach to 200 hours. This is due to the fed-batch condition where the at 5 g/L of

substrate concentration, the aeration rate was increased which affected the agitation power while

at 50 g/L of substrate concentration, the aeration rate was drop to the lowest as shown in Figure

1.0. Next, based on Figure 4.0, the highest peak recorded in graph from right (substrate

concentration of 15 g/L and biomass concentration of 10 g/L) is recorded at 30.2 at 48 hour. The

lowest peak recorded for this graph is at 29.67 at 190 hour. Therefore, to compare between

aeration rate and agitator power, the aeration of producing product in fermentation is undertaken

primarily to supply oxygen requirements which it is necessary oxygen for growth and production

of culture can be ensured by aeration and agitation of the culture. Therefore, the efficiency of

aeration can be improved by agitation which resulting in an increased interface between gas and

liquid.

Besides, the dissolve oxygen concentration refers to the microscopic bubbles formed

inside the vessels, which containing oxygen (O 2) gas mixed in the water and available for the

respiration of the organisms. Based on the Figure 5.0, for the dissolved oxygen concentration at

substrate concentration of 5 g/L is decreasing slowly with a period of time while the dissolved

oxygen concentration at substrate concentration of 50 g/L is slightly decreasing until 50 hours

and slightly increasing and remain constant. Next, based on Figure 6.0, the lowest reading

between the two is recorded at 0.85 when the biomass concentration is 20 g/L while the highest

peaks between the two is recorded at 1.12 when the biomass concentration is 10 g/L. For graph

from left (the substrate concentration is 15 g/L while the biomass concentration is 10 g/L) the

highest peak recorded is at the 400 hour. While for graph from right, the highest peak recorded is
at the 400 hour. The readings are not constant as the process undergone fed-batch process. Where

the fed is added bit by bit, thus resulting in the fluctuation in the reading of the graphs. These

three (aeration rate, agitation power and dissolved oxygen) are directly affecting one and the

other in the process. If one of the system failure, both the other two will be affected as well,

resulting in the failure for the whole process.

For 5 g/L and 50 g/L of substrate concentration, the graph pattern as shown in Figure 7.0

was a bit similar which it is slightly decreasing to the lowest until none of the substrate

concentration was detect as shown in Figure 7.0. This is due to the microorganism that consumed

all the substrate in the medium which is after 50 hours, the substrate have been consumed by the

microorganism. By referring to the graph against time for substrate concentration and biomass

concentration of 15 g/L and 10 g/L, the pattern showed that the concentration of substrate was

plunged from 14 to 0 g/L at initial time. This pattern also occurred for simulation condition of 15

g/L substrate condition and 20 g/L biomass concentration. Generally, according to study made by

Saarela U., 2003, penicillin is a secondary metabolite which were produced by inhibition of

growth of fungus by stress. There were several method to inhibit fungus by stress and one of

them is by reducing the substrate concentration which act as nutrient sources rapidly. Optimum

level of substrates condition were really crucial in order to perform any fermentation process

because it will affect the product and biomass concentration at the end of fermentation process

but in order to allow high amount of penicillin produced, the growth of fungus need to be inhibit.

This is also one of the reason of using fed-batch fermenter because the amount of substrate fed

can be adjusted by time to time.

Other than that, based on Figure 9.0, for the biomass concentration at 5 g/L of substrate

concentration is increasing with period of time until it reaches equilibrium at 300 hours to 400
hours while for the biomass concentration at 50 g/L of substrate concentration is slightly

increasing until 60 hours but it slowly decreasing until 400 hours. Furthermore, by studying the

graph pattern, it clearly showed that the biomass concentration were rapidly decrease from time

to time. For Figure 10.0, the initial concentration of biomass was 10 g/L and it increased rapidly

to 16 g/L at early time before decrease gradually to below 13 g/L at the end of fermentation time.

While for the second simulation condition, the biomass concentration also highly increased from

20 g/L to 26 g/L at early stage before decrease until reached 20 g/L at the end of the process.

Firstly, the rapid increase of the biomass concentration at early stage was strongly influenced by

the amount of substrate concentration fed to the bioreactor. By comparing to the substrate

concentration graph at initial stage, the substrate concentration that been introduced was 15 g/L

for both simulation condition. Due to that, the biomass inside the bioreactor had obtained their

nutritious for microbial growth thus increased the biomass concentrations at initial stage. When

the amount of substrate introduced into the reactor for both simulation condition were stopped

until the concentration was 0 g/L, the amount of biomass concentration also decreased for both

conditions. As mention above, penicillin was produced by inhibition of fungus by stress, so when

there were no more substrate in the medium, the biomass will start to stop growth and some of

them were dead due to nutritious lack. By that, the production of penicillin started to increase

rapidly.

The penicillin concentration graphs as shown in Figure 11.0 are slightly different because

the production of penicillin at 5 g/L of substrate concentration is increasing until 400 hours while

for the penicillin concentration at 50 g/L of substrate concentration is decreasing to the negative

value until 300 hours and continue to increased. Besides, penicillin concentration graph provided

by the Pensim Simulator showed that for first simulation condition, the amount of penicillin
produced increased rapidly from 0 to 1 g/L at the first 100 hours and keep increased gradually

until 350 hours. This condition happened because this was the stage where the fungus started to

inhibit due to insufficient substrate that act as nutrients for growth. Therefore, here was the stage

where penicillin was produced effectively. At the last 50 hours of fermentation time, the

production of penicillin started to plateau and slightly decreased. For second simulation

condition as shown in Figure 12.0, the penicillin concentration graph pattern was quite dispersed.

Therefore, the substrate concentration and biomass concentration will affected the penicillin

concentration.

Last but not least, Figure 13.0 shows the graph for substrate feed rate where the substrate

feed rate increasing suddenly before reach 50 hours for the condition of substrate concentration

of 5 g/L while the substrate feed rate increasing suddenly after 50 hours for substrate

concentration of 50 g/L. But, there are no differences shows for different biomass concentration

of 10 g/L and 20 g/L as shown in Figure 14.0. This can be explained by the fed-batch

fermentation condition where normally, for fed-batch fermenter, substrates were added at high

rate initially to promote growth at early phase and later will be adjusted to lower levels in order

to promote product promotion. Therefore, as the substrate feed rate increased suddenly, the

substrate concentration was initiated increasing while the biomass concentration also increasing.
CONCLUSION

Therefore, the dynamic simulation of fed-batch penicillin production was studied by

using Pensim simulator. Besides, there is a correlation between aeration rate and agitator power

where the efficiency of aeration in production of penicillin can be improved by agitator power

which resulting in an increased interface between gas and liquid. Next, when the amount of

substrate introduced into the reactor for both simulation condition were stopped until the

concentration was 0 g/L, the amount of biomass concentration also decreased for all conditions

except for the condition of substrate concentration of 5 g/L. This is because of the biomass will

start to stop growth and some of them were dead due to nutritious lack. By that, the production of

penicillin started to increase rapidly. This is also one of the reason of using fed-batch fermenter

because the amount of substrate fed can be adjusted by time to time. Lastly, for the substrate feed

rate, the graphs are a bit similar for all condition which the lines are increased suddenly. This can

be explained by the fed-batch fermentation condition where normally, for fed-batch fermenter,

substrates were added at high rate initially to promote growth at early phase. Therefore, as the

substrate feed rate increased suddenly, the substrate concentration was initiated increasing while

the biomass concentration also increasing.

REFERENCES

Vince F. (2014). Fermentation Phases. [Online]. [Accessed 10-03-2017]. Available

from world wide web: http://www.homebrewtalk.com/yeast-femrnetation-

phases.html

Saarela U., Leiviska K. et al., (2003). Modelling of a Fed-Batch Fermentation

Process. [Online]. [Accessed 10-03-2017]. Available from world wide web:

http://jultika.oulu.fi/files/isbn9514275144.pdf

Mestrovic T. and Chow S. (2015). Penicillin Production. [Online]. [Accessed

10-03-2017]. Available from world wide web: http://www.news-

medical.net/health/Penicillin-Production.aspx