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AGITATION AND AERATION

Faculty of Engineering & Built Environment
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Typical Fermentation Process

Stock Culture Raw Materials

Shake Flasks Medium Formulation

Seed Sterilization
Fermenter

Fermenter

Air

Recovery

Purification Products
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INTRODUCTION
Provision for adequate mixing of fermentation contents
Agitation Aeration

Motion by shaking or Process of introducing

stirring to achieve air to increase oxygen
mixing concentration in liquids
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Aeration
• Dissolved oxygen (DO) is important for aerobic fermentation and it is a substrate
limiting due to its low solubility in the medium/water.

• At high concentration of cell, the rate of oxygen uptake is more than the rate of
the oxygen supplied and it is a major limiting step for cell growth and metabolism.

• When the oxygen becomes a rate-limiting factor, the rate of growth changes with
dissolved oxygen concentration

• The critical oxygen level for bacteria and yeast is 5-10% from saturated DO, while
for fungi is 10-15% depends on the size of pellets.

• The existence of dissolved salt and organic material can change saturated value
of DO, and also by increasing the temperature could lower the saturated DO.

• Oxygen is being introduced into fermentation broth by air sparging. The rate of
transfer oxygen from bubbles to the cells is limited by oxygen transfer through
liquid film around the gas bubble.
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Agitation
• Mixing is usually provided by mechanical agitation with or
without aeration. The most widely used arrangement is the
radial-flow impeller with six flat blades mounted on a disk

• Why mixing is important?:

• to disperse the air bubbles
• to suspend the cells
• to enhance heat and mass transfer in the medium

• relate to Gas-liquid mass transfer

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Solubility of Gases in Biological Media

• The solubility of gases in liquid media for slightly moderate
soluble gas is describe by Henry’s law.

pA P
CA   yA 6
H H
Where
• CA = concentration in the liquid phase
• pA = gas phase partial pressure of A (atm)
• H = Henry’s law coefficient
• yA = mole fraction of A in gas phase

• Solubility of gas depends on:

• temperature,
• the partial pressure of the gas over the liquid,
• the nature of the solvent and the nature of the gas
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• Oxygen solubility: oxygen in water

T (deg C) Oxygen concentration Henry’s law

(Co2* mg/L) in constant (atm-liter-
equilibrium with air (po2 mg-1 )
= 0.209 atm)
25 8.10 0.0258
35 6.99 0.0299

• Oxygen solubility in water can be correlated with

temperature

CO2  14.16  0.349T  0.00774T 2  0.000646T 3

where CO2is in mg/L and T is in deg C.

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MASS-TRANSFER COEFFICIENT
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Mass Transfer Coefficient

• Is a function of physical properties and vessel geometry

• The mass flux is proportional to a concentration

difference. If a solute transfer from the gas to the liquid
phase, its mass flux from the gas phase (Eq 7) and liquid
phase (Eq 8) to interface are:

qG
NG   k G  CG  CGi  7
a
qL
NL   k L  CLi  CL  8 
a
Where,
• CG and CGi = gas-side concentration at the bulk and interface
• kG, kL = individual mass transfer coefficient
• a = interfacial area
• qG, qL = rate of mass transfer
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• Since the amount of solute transferred from gas phase to

the interface must equal to the interface to the liquid
phase, then
NG  NL (10)

• Substition of Eqs (7) and (8) into Eq (10) gives

CG  CGi k
 L 11
CL  CLi kG
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• It is hard to determine the mass transfer coefficient

according to Eq (7) or (8)

• Because the interfacial concentrations, CLi or CGi cannot

be measure

• Define the overall mass-transfer coefficient as Eq. (12)

N G  N L  K G  CG  C*G   K L  C*L  CL  12 

where,
CG* is the gas side concentration which in equilibrium with the existing liquid phase concentration
CL* is the liquid side concentration.
K G and K Lare overall mass transfer coefficient for gas and liquid side
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Correlation for Mass Transfer Coefficient

• In mass transfer correlations of gas-liquid dispersion, the
following dimensionless group are often employed
• Since they are dimensionally consistent and also useful
for scale-up processes

total mass transfer k D

Sherwood number, NSh   L 32 13
diffusion mass transfer D AB
momentum diffusivity c
Schmidt number, NSc   14 
mass diffusivity C D AB
gravitation forces D332
Grashof number, N Gr   2 15
viscous force c
subscript c is referred to continuous phase. D32 is sauter mean
diameter
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• Mass transfer between the gas-liquid phase reported the

volumetric mass-transfer coefficient kLa.

• k La
• the combination mass transfer coefficient and interfacial area.
• difficult to identify which parameter is responsible for the change of
k La

• Calderbank and Moo Young (1961) separate kLa by

measuring interfacial area and correlated mass transfer
coefficients in gas-liquid dispersions
• In mixing vessels and sieve and sintered plate column
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1. For small bubbles < 2.5 mm in diameter

1/3
  c g 
k L  0.31N Re 2/3   (16)
  2
c 
or NSh  0.31NSc1/3 N Gr1/3 (17)

• The more general forms which can be applied for both small
rigid sphere bubble and suspended solid particle are:

2D AB 2/3   c g 
kL   0.31NSc   (18)
D32   2
c 
or NSh  2.0  0.31NSc1/3 N Gr1/3 (19)

• Eq. 18 and 19 for mass transfer to and from dispersion of low-density

solid particles in agitated liquids
• to simulate mass transfer to microorganisms in fermenter
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2. For bubble > 2.5 mm in diameter

1/3
  c g 
1/2
k L  0.42 NSc   (20)
  2
c 
or NSh  0.42 NSc1/2 N Gr1/3 (21)

• kL varies from 7.3x 10-4 to 3.4 x 10-3 m/s depending on the

power dissipation by impeller per unit volume (Pm /v) (Lopez De
Figueiredo and Calderbank (1978).
0.33 0.25
Dimensionally consistent
 Pm   Pm 
k L    k L  0.592D AB
1/2
  (22)
 v   v 
• Based on the absorption of CO2 into water in an agitated vessel
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• kL based on the O2 absorption into several liquids of

different physical properties w/o mechanical agitation
Akita and Yoshida (1974)

1/2 1/4 3/8

k L D32  vc   g D   g D  
3 3
 0.5   
32
 
32 c
 (23)
D AB D v 2
 AB   c    

• νc and σ are the kinematic viscosity of the continuous

phase and interfacial tension

• Applicable for column diameter up to 0.6m, superficial

velocity up to 25m/s and gas hold-up to 30%
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Measurement of Interfacial Area

• Can be measured by employing several techniques:
• photography, light transmission, and laser optics.

• Can be calculated as in Eq. (24):

6H
a (24)
D32

volume fraction of gas-phase

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Empirical Correlation for a and D32

1. Gas sparging with no mechanical Agitation (Akita and Yoshida
1974)
0.50 0.12 0.12
D32  g D c  2
 gD  3  Vs 
 26  
C
 
C
  (25)
DC     v 
2
c  gDC 
and for the interfacial area
0.50 0.1
1  g D L 2
 gD 
3
aDC   
C C
 2  H1.13
(26)
3    vL 
• DC is bubble column diameter and Vs is superficial gas velocity, which is gas flow
rate Q divided by the tank cross-sectional area.

• Eqs. (25) and (26) are based on data in columns of up to 0.3 m in diameter and
up to superficial gas velocities of about 0.07 m/s.
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2. Gas sparging with Mechanical Agitation

A. For VS < 0.02 m/s. Interfacial areas for the gas-liquid

dispersion agitated by a flat-blade disk turbine as
follows:
  Pm / v 0.4 0.2   Vs 
1/2

ao  1.44  c
  (27)
  0.6
  Vt 
0.3
 N DI 
  20, 000
0.7
For N Re 
 VS 
where N Re is the impeller Reynolds number defined as
D 2I Nc
N Re  (28)
c
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• For
0.3
 N DI 
  20,000
0.7
N Re 
 VS 
the interfacial area a 0
0.3
 2.3a  5 0.7  ND I 
log10    1.95 x10 N Rei   (29)
 o 
a  s 
V

Obtained from the Eq. (27)

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B. For VS > 0.02 m/s

ao at Vs > 0.02, Eq (27) is modified by Miller (1974) as

  Pe / v 0.4 c 0.2   V 1/2

a o  1.44   
s
 (30)
  0.6
  t
V  Vs 

Where,
• Pm is power input by mechanical agitation replace by Pe
effective power input
• Vt by (Vt+Vs).
• The effective power input Pe combines both with gas
sparging and mechanical agitation energy.
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Sauter mean diameter for the gas liquid dispersion agitated by a

flat-blade turbine impeller

1. For dispersion of air in pure water

 0.6  0.5 4
D32  4.15   H  9.0x10 (31)
  Pm / v   
0.4 0.2

2. In electrolyte solution (NaCl, Na2SO4 and Na3PO4)

  0.6  0.4   d 
0.25

D32  2.25  0.4 0.2 

H   (32)
  Pm / v     c 

3. In alcoholic solution (aliphatic alcohols)

  0.6  0.65   d 
0.25

D32  1.90  0.4 0.2 

H   (33)
  Pm / v     c 
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The preceding 3 Eqs can be

modified for high gas flow rate
(Vs> 0.02m/s) by replacing Pm
by Pe as suggested by Miller
(1974)
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Correlation for a and

D32

Gas sparging with no Gas sparging with

Sauter mean diameter
mechanical Agitation Mechanical Agitation

Columns of up to 0.3 m
in diameter alcoholic solution
VS > 0.02 m/s
Superficial gas velocities (aliphatic alcohols)
of about 0.07 m/s.

electrolyte solution
VS < 0.02 m/s (NaCl, Na2SO4 and
Na3PO4)

dispersion of air in pure

water
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Determination of O2 Absorption Rate

• The O2 absorption rate per unit volume qa/v can be
estimated by

 K L a  C*L  CL   k L a  C*L  C L 
qa
(40)
v
Since the O2 is sparingly soluble gas, the overall mass-transfer
coefficient KL is equal to the invidual mass-transfer mass transfer
coefficient, kL.

• Objective in fermenter design:

• to maximise the O2 transfer rate
• to minimum power consumption necessary to agitate the medium
• to minimum air flow rate
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• To maximize the oxygen transfer rate (OTR), have to maximize

i. kL the main design parameter
ii. a
iii. C*L-CL
 concentration difference is quite limited to control because the value of
CL* is very low maximum solubility.

• For the use of air to supply the O2 demand, maximum

concentration of O2 in water which is in equilibrium with CL* at atm
pressure is about one fifth of total solubility (0.209 atm), according
to Hendry’s law

po 2
C *
(41)
Ho2  T 
L
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O2 Absorption
Rate

Sodium Dynamic
Dynamic Direct
Sulfite Gassing-out
technique measurement
Oxidation Technique
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Sodium Sulfite Oxidation Method

• Based on the oxidation of sodium sulfite to sodium sulfate in
presence of catalyst
Cu   or Co  
Na 2SO3  O 2 
1
2  Na 2SO 4

• The reaction has following characteristics:

1. Reaction rate is independent of the concentration of sodium sulfite within
the range of 0.04 to 1 N
2. Reaction rate faster than OTR, therefore rate of oxidation is controlled by
rate of mass transfer alone.

• Limitation of the method

1. the solution cannot estimate the physical & chemical properties of
fermentation broth
2. High ionic strength affects the interfacial area, hence lower the mass
transfer coefficient.

• This technique is helpful in comparing the performance of fermenters

and studying the effect of scale-up and operating conditions
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Dynamic Gassing-out Technique

• Monitors the change of the O2 concentration by polarographic
electrode
• O2-rich liquid is deoxygenated by passing nitrogen through it.

dCL  t 
 k L a C*L  C L  t   (42)
dt

• Integration,  C*L  CL  t1  
ln  * 
 C  C  t 
2 
k La 
L L
(43)
t 2  t1

• kLa can be calculated based on the measured values of CL(t1)

and CL(t2)
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Direct measurement
• Directly measure the gas stream entering and leaving the
fermenter by gaseous oxygen analyser. The O2 uptake can be
q a   Qin CO2,in  Qout CO2,out 
calculated as
(44)

 k L a  C*L  CL 
qa
(40)
v
q  mass transfer rate, kmol / s
• If the size of fermenter is small (<50 L), variation (CL*-CL) is
fairly small

• For larger fermenter size > 50 L, the log-mean of CL*-CL can

be used

(C  CL ) LM
*

 C  C   C  C 
*
L L in
*
L L out
(45)
ln  C  C  /  C  C  
L
* *
 L L in L L out 
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Dynamic technique
• Based on material balance in an aerated batch fermenter
while microorganism are actively growing as
Cell respiration rate
(gO2/g cell h), /Yx/o X
 k L a  C*L  CL   rO2 CX
dCL
(46)
dt
• While the O2 level is steady, the air supply is turned off,
and the O2 will be decreased with this Eq.

dCL
 rO2 CX kLa is equal zero (47)
dt

the slope from plot CL vs t

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• Then the O2 is turned on again, O2 concentration will be

increased according to Eq

1  dCL 
CL  CL  *
  rO2 C x  (48)
k L a  dt 

The plot CL vs (dCL/dt+ro2Cx) will result in a straight line which has

the slope of -1/(kLa) and y-intercept of CL*