THE VOLUMETRIC MASS TRANSFER COEFFICIENT kLa AND METHODS OF THE VOLUMETRIC MASS TRANSFER COEFFICIENT kLa AND METHODS OF MEASUREMENT

Mass Balance of Oxygen in Unit Liquid Volume FIG. 2.7 Schematic diagram of the mass balance of oxygen transfer in unit liquid volume

Mass Balance of Oxygen in Unit Liquid Volume (Cont’d)

Mass Balance of Oxygen in Unit Liquid Volume (Cont’d)

Mass Balance of Oxygen in Unit Liquid Volume (Cont’d)

Mass Balance of Oxygen in Unit Liquid Volume (Cont’d)

Mass Balance of Oxygen in Unit Liquid Volume (Cont’d)

Methods of Measurement of KLa in a Bioreactor

Chemical Methods of KLa Measurement FIG. 2.8. Schematic diagram of a stirred tank batch reactor

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) The Bioreactor Vessel is Equipped with: ● The D.O. Probe, Connected to a D.O. Analyzer. ● Chart Recorder: To Measure Signal from D.O. Probe and Measure On-line the D.O. Concentration in the liquid phase of the Bioreactor.

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) ● The D.O. Probe Measures the PyO2 Partial Pressure (PyO2) of dissolved O2 in the liquid phase, which means that it measures HO2CL. Where: HO2 = Henry’s Constant for O2 in Water CL = D.O. Concentration In the Liquid Phase (Mass of O2/L)

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) Fig. 2.10 Set up of a Stirred tank Bioreactor with Dissolved Oxygen Probe, pH probe and accessories.

● Turning air ON and OFF while Maintaining the same R.P.M. we can: In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) ● Turning air ON and OFF while Maintaining the same R.P.M. we can: Record the D.O. Probe Output in the Chart Recorder. From these Data, we can get KLa, QO2, CL* at given in-situ Bioreactor Conditions.

● The ON-OFF Operation takes 5 min, during which time: In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) ● The ON-OFF Operation takes 5 min, during which time: Cell Concentration X (g /L)  Constant. We make sure that the D.O. Concentration CL never falls below the critical oxygen concentration CCRT, which means that the respiration rate coefficient QO2 = QO2Max = Constant. ● Using the D.O. probe output and a recorder we measure directly the D.O. concentration as a function of time, t.

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor While we maintain the same R.P.M. of the bioreactor impeller, we turn the AIR-OFF. During the AIR-OFF period the following conditions apply: ● Rate of Supply of O2 = 0 ● No Air Present in the Bioreactor ● KLa = 0 because a = 0, no air bubbles present ● Using Eq. 2.2 for O2 Mass Balance, we have: ● We know cell concentration X by measuring it. Therefore, we calculate QO2 because we also measure the slope – QO2X.

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor ● Fig.1 Shows D.O. concentration CL inside the bioreactor = f(t) when Air is turned Off and On, always keeping the R.P.M. of the impeller the same to provide good mixing of the liquid phase. ● After a period of about 5 min, a liquid sample is taken from the bioreactor to measure the cell concentration X (g dry wt./L). ● The KLa, QO2, and CL* values correspond to that specific fermentation time and given cell growth conditions. ● We can do many AIR-OFF and AIR-ON measurements to get all three parameters KLa, QO2, and CL* as a function of total batch fermentation time.

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor FIG.1 Transient Air-Off, Air-On Experiment in a Bioreactor System

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor ● During the AIR-OFF period the D.O. concentration CL is plotted as a function of time t from which we get the slope = - QO2X, as shown in Fig. 2. FIG. 2 D.O. concentration CL as function of time during AIR-OFF period.

 From the CL vs. time (t) data we can get In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor  AIR-ON Period During this period the following oxygen mass balance equation applies:  From the CL vs. time (t) data we can get

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor ● Re-arranging Eq. 2.2 and solving for CL we get Eq. 2.9 ● By plotting CL vs. at a given fermentation time, t, we can get the slope which is equal to

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor (Cont’d) ● and therefore, the value of KLa is found, and the intercept also gives the value of ● During the Air-On Period: CL* = Constant QO2 = Constant KLa = Constant CL, dCL/dt vary with time t

In Situ Measurement of KLa, QO2, and CL In Situ Measurement of KLa, QO2, and CL* During Cell Growth in a Bioreactor FIG. 2.13. D.O. concentration CL as function of [dCL/dt + QO2X] during AIR-ON period.

GENERALIZED VIEW OF BIOPROCESS

TYPICAL BIOPROCESS FLOW SHEET