The experiment was done to study the growth kinetics of microorganism in

bioreactor experiment, construct a growth curve and to determine the Monod

parameter. The objective of this experiment were concluded in this discussion.
The table 6.2 and 6.3 shows all the data needed to construct the growth curve
of E.coli being fermented in the shake flask. From these data, Figures 6.1 and 6.2
were constructed, and these graphs were used to determine the Monod
parameters.
Figure 6.1 and 6.2 illustrate the absorbance analysis and cell dry weight of
the bacteria against time. The absorbance and cell biomass against time graphs
however does not resemble the theoretical growth curve of most common
microorganism but compare to the previous experiment the fermentation in
conical flask, this experiment the growth curve are much more close to the
theoretical growth curve. Roughly, it does not exhibit any lag phase, which
proves that the broth from the seed culture flask poured into the bioreactor did
contain microorganisms already in its exponential phase. After a couple of
minutes adjusting to its new vessel, the broth quickly shoots up and started
growing, showing an apparent exponential phase that went on until the
experiment ends, where it can be observed that stationary phase does not exist
in the growth curve figure 6.2. However, it can be assume at around 9 th hour the
stationary phase is observe based on previous experiment done by Olympia
et.al. (2010) and other group result. This result obtains may not accurate due to
error reading when weighting cell dry mass and initial tube reading. During
drying, the cells in tube are over dry which lead mistakes in obtaining cell dry
weight. Other factor like present of air moisture, sample taken does not
homogenously mix and measure apparatus failure also contributes to the result
gain. From this result, the death phase cannot be observed as the experiment
stops during stationary phase based on theoretically. It is assume death phase
take place in 48th hour upwards.
The figure 6.2 was constructed based on the results of the cell dry weight.
This graph however shows logical results and correlates with the growth curve of
the microorganism as it showed an increase in value, which indicates that the
microorganisms are starting to get out of its dormant state and start replicating,
thus the increase in mass as it moves along the horizontal axis. After reaching a
peak at 10th hour, the graph is slowly increase and from result in table 6.3 some

value show a decrease in cell mass concentration after reaching 10 th hour. This
can correspond to the stationary phase of the microorganism in its growth curve.
Figure 6.1 shows that absorbance reading increase from 0 hour to 10 th
hour which indicates that the cell concentration increase. At 10 th hour upward the
absorbance reading are nearly constant which indicates stationary phase is
surpass the lag phase.
From the result gain, this shows that the fermentation in the bioreactor is
successful and can be acquitted to a few facts. It showed its nutrient is sufficient
for it to survive as well as the conditions during the fermentation was optimal for
the microorganism to survive. However, the analysis from the absorbance
reading is not highly accurate due to the fact that absorbency tests not only
measure the cells, but also dead cells and other unwanted molecules. However
the absorbance reading shows a familiar trait with the cell mass concentration.
It can also be noted that absorbency test are most suitable for all the phases
except the death phase. This is because there are more dead cells in the death
phase, and the readings have a higher chance to be inaccurate.
Compared to the shake-flask experiment, the glucose concentration graph
was not able to be constructed since the substrate used in this bioreactor
fermentation is glycerol. Unfortunately, there is no available equipment to
analyse glycerol concentration in the laboratory. Therefore, glycerol analysis was
not able to be carried out. Consequently, the saturation constant, which is
dependent on the substrate, could not be determined. If present, more
microorganism activity could be monitored.
By utilizing a simple conversion method, the cell biomass from the cell
mass concentration was determined, and the graph in Figure 6.3 was plotted. As
mentioned previously, the readings from the absorbance test may not be very
accurate due to the possible presence of other molecules in the sample.
Therefore, it is advisable to convert its values to the cell biomass form. Similar to
the growth curve, this curve too, displays its two phases which is exponential
phase and stationary phase. By comparing three growth curve graph which are
plotted by using three different data, it can be simplified that, the exponential
phases of the growth were between 1st to 16th hour which is familiar with
previous study conducted by Chung et. al, (2015), Olympia et.al. (2010)

and

Yang et. al (2013) . For the conversion calculation, the highest approximate

values given by the data points were used to determine its gradient, which in
turn produced the Monod parameters that was the objective of the experiment.
From the exponential phase gradient, it was found that the net specific cell
growth rate, net 0.0437 hr-1. This value represents the specific growth rate of the
microorganisms under the conditions of this experiment. Thus the doubling time
can be obtain by calculation mention in above and the doubling time is 15.8615
hr. Compared to the shake flask experiment done prior to this lab, the Monod
parameters in bioreactor suppose report a higher net. However, in this
experiment, the value selection, result gain and calculation might affect the net
obtained. From other group, the net gain is 1.1350 hr-1 and doubling time is
7.4451 h which is more correct value compare to our group. The higher value of
net show that fermentations carried out in bioreactors are much more desirable.
Some of the advantages of using the bioreactor instead of shake flask
fermentation is that the parameters can be controlled more accurately, and
automatically. If there is any depletion or change in, for example, dissolved O 2
levels, the instrumentation and control system of the bioreactor have been
designed to adjust the levels accordingly. This is the function of the cascade
settings done during the setup of the bioreactor. From this system, the software
installed with the bioreactor is able to record all the level changes during the
reaction.
From this experiment, few error was observer in the experiment. During
drying and weighting for cell dry weight analysis, it was observed that there are
few moisture in tube and it was manually removed. This condition may probably
effect the initial weight even it had been dying using oven and put inside the
desiccator. Due to ineffective time management, the inoculum stage was done
more than 4 hours exceeding the procedure suggested time. Extending time for
inoculum stage may lead to dead of few cell as the depletion on nutrient. Thus it
may affect the experiment, During experiment, the lack of equipment available
in laboratory lead to failure in obtaining correct data for initial reading of ph
level, dissolved oxygen and antifoam which available on computer data.The
computer software for the bioreactor failed to be installed. However, during on
4th reading the software is successful working and start takes reading. During
early of experiment, there was error on base drop. The pump does not move the
base into the bioreactor as the pipe position not allow the base to be pump.
Thus, this condition lead to the bioreactor to be in acidic condition. It was realize
during on 3rd - 4th reading that the bioreactor has low ph. Thus this problem was

overcome by manually add base until the desire pH value reach and the pipe was
adjusted so that it works perfectly.