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Wednesday, November 28, 2007

Day Three Part III

11:01:00 PM

Introducing to you the History Plot!

(click to view enlarged image)

A colourful graph was presented to us. From the lab technician, we found out that it was actually called the History Plot.

What the hell is a History PLOT??? The History Plot (from the title that is already so obvious) covers the past events of the fermentation process in the bioreactor. It records the major operational parameters during our fermentation process.

In our case, it consists of 4 major operational parameters. They are the pH level in red, dissolved partial oxygen level (PO2) in light blue, stirring speed in green, and temperature in dark blue.

The values of these parameters fluctuate due to the computer system trying to negate the changes in the parameters so that the optimal growth condition is maintained.

Overview of the graph:

• The lag phase for our group can be said to be extremely short which amounts to about 1.3 hours
• By cross referencing with the cell growth graph, the log phase starts, but the log phase is begins 1.3 hour after inoculation and lasts for 3.7 hours till the 5th hour.
• The stationary phase begins at around the 5th hour and ends around 17.5th hours.
• The death phase then follows.

Temperature:

The optimal temperature for cell growth (from 31.5˚C to 32.5˚C) was kept constant throughout by the automated system.

pH:

It is rather consistent throughout the lag and log phase but there is a slight increase in pH value as it transits from log phase to stationary phase. After which it remain relatively unchanged/constant. The sudden increase may be attributed to the toxins and the secondary metabolite.

pO2 level and Stirrer speed:

From Lag phase to Log phase pO2 level decreases rather consistently with a slight irregularity (a sudden increase)
As pO2 level hits the 24.0% saturation mark, the stirrer speed increases to keep the pO2 level from going any lower. This results in a relatively low change in the pO2 level, during the log phase, after hitting the 24% saturation.
As the fermenter enters the stationary phase, a great amount of fluctuation in pO2 level occurs. To negate changes and maintain the minimum of 24% saturation, the system adjusts the stirrer speed. This resulted in the coordinated peaks and lows between 5th hour to 17.5th hour.
In the death phase (17.5 hours onwards), cell death rate is higher then growth rate and there is a decrease in cell O2 demand. This causes the pO2 level increase as the rate at which O2 dissolves is higher then the O2 up take rate. This in turn causes the stirrer speed to decrease to the minimum of 200rpm.

Since it is said that both the pO2 level and the stirrer speed are inversely proportional, we shall explain the non-coordinated graph of pO2 and stirrer speed between time 2.5h to 5h.

As observed, the stirrer speed is increasing rather consistently while the pO2 level remains constant. The reason being that this is the log phase.

During the log phase O2 demand level increases without the stirrer the pO2 level will drop consistently, however the automated system deems that the lower limit for pO2 level is 24% saturation. Therefore, to keep the pO2 at least at 24% saturation, the stirrer speed increases to make the dissolving rate of O2 equivalent to the O2 uptake rate. As the uptake rate and the dissolving rate is the same, the net change is zero. Hence the pO2 level remains constant during this period.

There are four sections to Day 3:

Part I (Procedures)

Part II (Growth Graph)

You are Here: Part III (History Plot)

Part IV (Futher Questions)

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Day 1, Experiment 1: Familiarization of bioreactor and equipments
Day 1, Experiment 2: Equipment and media preparation
Day 2: Inoculation, fermentation and monitoring
Day 4: Harvesting of the cells
Day 5: Isolation and purification of product