Perancangan Bioreaktor

TEKNOLOGI BIOPROSES
Teknologi Bioproses telah secara luas digunakan dalam
produksi enzyme dan protein yang digunakan dalam bidang
1)Pengolahan Pangan (Asam Glutamat)
2)Pengelolaan Limbah (EM4)
3)Penelitian Kedokteran (Vacin, serum, testkit)
4)Pertanian/Perikanan/Peternakan (Pupuk organic)
5)Pengembangan Bidang Farmasi. (Bahan dasar obat obatan)
6)Sejumlah bidang ilmu pengetahuan yang luas (Kosmetik,
bahan untuk militer dll)
 Hasil Hasil dari Teknologi Bioproses

Benih Berkualitas Produk Makanan

Obat-obatan

Pestisida Alami Pakan Ternak

POHON INDUSTRI UBI KAYU
Lanjutan
Rulide is used to treat infections in different parts of the body
caused by bacteria. For example: acute pharyngitis (sore throat and
discomfort when swallowing)

Sterile CLAFORAN (cefotaxime sodium) is a semisynthetic, broad spectrum

cephalosporin antibiotic for parenteral administration. It is the sodium salt of
7-[2-(2-amino-4-thiazolyl) glyoxylamido]-3-(hydroxymethyl)-8-oxo-5-thia-1-
azabicyclo [4.2.0] oct-2-ene-2-carboxylate 72 (Z)-(o-methyloxime), acetate
(ester). CLAFORAN contains approximately 50.5 mg (2.2 mEq) of sodium per
gram of cefotaxime activity. Solutions of CLAFORAN range from very pale
yellow to light amber depending on the concentration and the diluent used.
The pH of the injectable solutions usually ranges from 5.0 to 7.5. The CAS
Registry Number is 64485-93-4.
 BIOTEKNOLOGI DAN TEKNOLOGI BIOPROSES
Bioteknologi : Ilmu yang mendesain atau memodifikasi atau melakukan
perakitan materi yang memiliki orientasi hayati.
Obyek yang dipelajari : Protein, Enzim, mikroorganisma, sel
tumbuhan/hewan, jaringan, sel induk dll.
Tujuan : untuk menghasilkan produk yang berharga untuk
kesejahteraan dan bermanfaat bagi manusia untuk pangan dan
kesehatan.
Peran Bioteknologi meningkat pesat
Awalnya Teknologi pasteurisasi oleh Louis Pasteur.
Produknya : Ragi roti, keju, yoghurt, tape, tempe, protein sel tunggal,
pengembengan enzim amilase, glukosa dehydrogenase, antibiotic,
vitamin, asam amino, asam organic, etanol, metana, aplikasi
lingkungan
Teknologi Bioproses : Masih jarang didengar
Bagian tak terpisahkan dan sangat berperan penting dalam
aplikasi bioteknologi secara massif dalam industry.
TBP adalah teknologi yang berkaitan dengan segala operasi dan
proses yang memanfaatkan mikroba, baik dalam fasa hidupnya
maupun produk-produk enzimnya

Bioprocess engineers are engineers working to applythe

principles of various disciplines, such as chemical, mechanical,
electrical, and industrial, to processes based on using living cells
or subcomonents of such cells.
Teknologi Bioproses menggunakan system hayati atau biomolekul dalam
pembuatan produk pada skala komersial dengan memperhatikan efektivitas
biaya atau nilai ekonomi. Sistem hayati yg banyak digunakan : sel atau
jaringan, sedangkan system biomolekul yang digunakan : enzim, protein,
antibody dan DNA/ RNA

Teknologi Bioproses lahir dari berbagai disiplin ilmu, pada awalnya ilmu dasar :
biologi dan kimia melairkan teori teori dasar mikrobiologi dan biokimia.
Perkembangan ilmu rekayasa, seperti matematika dan fisika turut mendorong
berkembangnya bioteknologi sehingga melahirkan TEKNOLOGI BIOPROSES.

Teknologi Bioproses lahir dan berkembang untuk memenuhi kebutuhan hidup

manusiadan mengatasi keterbatasan manusiadengan memanfaatkan aktivitas
mikroorganisma
 Material : Farmasi,
Industri Gas, minyak, Kedokteran/Pert
kosmetik, Pangan, Pakan,
petrokimia anian
Energi, Lingk

Rekayasa Biokimia, kimia Tekonologi

Bioteknologi
phisik Bioproses

Thermodinamika, unit
Ilmu Terapan operasi proses, desain Mikrobiologi, genetika
kendali dll molekuler

Akar/dasar ilmu Kimia Fisika/matematik Biologi

a
 Beda Fokus Perhatian Ahli Ilmu Dasar Biologi/Kimia
dengan Teknologi Bioproses

Ahli Biologi/ Kimia Ahli Teknologi Bioproses

1. Klasifikasi, Fisiologi, dan 1. Fisika dan kimia terkait
Struktur fenomena hayati.
2. Sifat unsur dan senyawa. 2. Modeling dan pendekatan
3. Mekanisme peralatan matematik
laboratorium. 3. Desain proses, control
4. Analisis dan interpretasi proses dan peralatan.
data lab dari system 4. Perancangan produk dan
komplek pabrik
Kebutuhan Produk Produk Utama dari Proses Hayati :

Produk Fermentasi Organisme Umum yang dipakai Perkiraan Pasar Dunia (kg/tahun)

Etanol (non beverage) Saccharomyces cerevisiae 2 x 10 10

Aseton/Butanol Clostridium acetobutylicum 2 x 10 6

Asam amino Asam L glutamat Corynebacterium glutamicum 2 x 10 8

Asam Amino L-lisina Brevibacterium flavum 3 x 10 7

Asam sitrat Aspergilus niger 2 x 10 7

Asam glukonat Apergilus niger 5 x 10 7

Asam Laktat Laktobacilus delbrueckii 2 x 10 7

Penisilin Penicilium chrysogenum 4 x 10 6

Tetrasiklin Streptomyces aureofaciens 1 x 10 6

Vitamin B12 Pseudomonas denitrificans 1 x 10 4

 Basic Fermenter Design Criteria
• Microbiological and biochemical characteristics of the
cell system (microbial, mammalian, plant)
• Hydrodynamic characteristics of the bioreactor
• Mass and heat transfer characteristics of the bioreactor
• Kinetics of the cell growth and product formation
• Genetic stability characteristics of the cell system
• Aseptic equipment design
• Control of bioreactor environment (both macro and
micro-environment)
• Implications of bioreactor design on downstream products
separation
• Capital and operating costs of the bioreactor
• Potential for bioreactor scale-up
 Requirements of Bioreactors
• There is no universal bioreactor.
• The general requirements of the bioreactor are as follows:
– The design and construction of bioreactors must keep sterility
from the start point to end of the process.
– Optimal mixing with low, uniform shear.
– Adequate mass transfer, oxygen.
– Clearly defined flow conditions.
– Feeding substrate with prevention of under or overdosing.
– Suspension of solids.
– Gentle heat transfer.
– Compliance with design requirements such as: ability to be
sterilized; simple construction; simple measuring, control,
regulating techniques; scale-up; flexibility; long term stability;
compatibility with up- downstream processes; antifoaming
measures.
 Fermenter Design
• The basic points of consideration while designing a fermenter:
– Productivity and yield
– Fermenter operability and reliability
– Product purification
– Water management
– Energy requirements
– Waste treatment
• Other few significant things to be taken in account:
– Design in features so that process control will be possible over
reasonable ranges of process variables.
– Operation should be reliable
– Operation should be contamination free
 Fermenter design
• To achieve these the fermenter should have:
– Heat and oxygen transfer configuration
– Sterilization procedures
– Foam control
– Fast and thorough cleaning system
– Proper monitoring and control system
• Traditional design is open cylindrical or rectangular vessels made
from wood or stone.
• Most fermentations are now performed in close system to avoid
contamination.
• It should be constructed from non-toxic, corrosion-resistant
materials.
• Small fermentation vessels of a few liters capacity are constructed
from glass and/or stainless steel.
 Fermenter design
• Pilot scale and many production vessels are normally made of
stainless steel with polished internal surfaces
• Very large fermenters are often constructed from mild steel lined with
glass or plastic, in order to reduce the cost.
• If aseptic operation is required, all associated pipelines transporting
air, inoculum and nutrients for the fermentation need to be sterilizable,
usually by steam.
• Most vessel cleaning operations are now automated using spray jets,
and called cleaning in place CIP. And located within the vessel.
• Associated pipe work must also be designed to reduce the risk of
microbial contamination. There should be no horizontal pipes or
unnecessary joints and dead stagnant spaces where material can
accumulate; otherwise this may lead to ineffective sterilization.
 Fermenter design
• Normally, fermenters up to 1000 L capacity have an external jacket,
and larger vessels have internal coils.
• Pressure gauges and safety pressure valves must be incorporated,
(required during sterilization and operation).
• For transfer of media pumps are used. Centrifugal pumps (generate
high shear forces and path for easy contaminations), magnetically
coupled, jet and peristaltic pumps.
• Alternate methods of liquid transfer are gravity feeding or vessel
pressurization.
• In fermentations operating at high temperatures or containing
volatile compounds, a sterilizable condenser may be required to
prevent evaporation loss.
• Fermenters are often operated under positive pressure to prevent
entry of contaminants.
 Summary of bioreactor systems
Bioreactor design Cell systems used Products
Air-lift bioreactors Bacteria, yeast and fungi SCP, Enzymes, Secondary
metabolites, Surfactants

Fluidized bed bioreactor Immobilized bacteria, yeast Ethanol, Secondary

and other fungi, Activated metabolites, Wastewater
sludge
Microcarrier bioreactor Immobilized (anchored), Interferons, Growth factors,
mammalian cells on solid Blood factors, Monoclonal
antibodies, Vaccines,
particles Proteases, Hormones

Surface Tissue Mammalian, tissue growth Interferons, Growth factors,

on solid surface, tissue Blood factors, Propagator,
Monoclonal antibodies,
engineering Vaccines, Proteases,
Hormones
 Summary of bioreactor systems
(cont’d)
Bioreactor design Cell systems used Products
Membrane Bioreactors, Bacteria, yeast Mammalian Ethanol, Monoclonal anti-
Hollow fibers and cells, Plant cells bodies, Interferons, Growth
membranes used,
Rotorfermentor factors, Medicinal products
Modified stirred tank Immobilized bacteria, yeast Ethanol, Monoclonal anti-
bioreactor and plant cells bodies, Interferons, Growth
factors,
Modified packed bed Immobilized bacteria, yeast Ethanol, Enzymes,
bioreactor and other fungi Medicinal products

Tower and loop Bioreactors Bacteria and yeasts Single cell proteins
 Summary of bioreactor systems
(cont’d)
Bioreactor design Cell systems used Products
Vacuum bioreactors Bacteria, yeast and fungi Ethanol, volatile products

Cyclone bioreactor Bacteria, yeast and fungi Commodity products and

SCP
Photochemical bioreactor Photosynthetic bacteria, Interferons, Growth factors,
Algae, Cyano bacteria, Blood factors, Monoclonal
antibodies, Vaccines,
plant cell culture, r-DNA Proteases, Hormones
plant cells
Surface Tissue Mammalian, tissue growth SCP, algae, medicinal plant
on solid surface, tissue products, monoclonal anti-
bodies, vaccines and
engineering interferons
Bioreactor configuration
1. Stirred tank bioreactors

Mixing method: Mechanical

agitation
•Baffles are usually used to
reduce vortexing
• Applications: free and
immobilized enzyme reactions
•High shear forces may damage
cells
•Require high energy input
Schematic diagram of a tower bioreactor system with
multiple impellers and liquid down comer and
counter-current air liquid flow
Air Feed

Perforated
plate

Downcomer

Baffle

Impeller

Product
Air
2. Bubble column bioreactors

Mixing method: Gas

sparging
• Simple design
•Good heat and mass transfer
•Low energy input

Gas-liquid mass transfer

coefficients depend largely on
bubble diameter and gas hold-
up.
 Schematic diagram of a tower bioreactor
system with perforated plates and co-current
air liquid flow
Air exhoust

Broth
outlet

Sampling
nozzles
Jacket

Constant temp.
water bath Perforated
Pump
plate

Compressed
air Orifice
Sparger

Air filter
Peristaltic Medium
pump reservior
Flow
Medium meter
inlet
3. Air lift reactors
Mixing method: airlift
• Compared to bubble
column reactors, in an
airlift reactors, there
are two liquid steams:
up-flowing and down-
flowing steams. Liquid
circulates in an airlift
reactor as a result of
density difference
between riser and
downcomer.
ICI Deep shaft unit
 EMLICHHEIM FLOWSHEET
CONDENSATE,
MAE-UP WATER, AND
CLARIFIER FLOCCULATING AGENT
WASH
WATER
SETTLEMENT RECYCLED
TANT WATER
SAND B RECYCLE SLUDGE
B

DECANTER
CENTRIFUGE FLOATATION

SOIL AND
SLUDGE
DEEP
SHAFT

AIR

COMPRESSOR

LAGOON
4. Packed bed bioreactor
Packed-bed
reactors are used
with immobilized
or particulate
biocatalysts.

Medium can be
fed either at the
top or bottom and
forms a
continuous liquid
phase.
Packed bed perfusion bioreactor for
mammalian cell culture
5. Trickle-bed bioreactor
The trickle-bed
reactor is another
variation of the
packed bed
reactors.

Liquid is sprayed
onto the top of the
packing and
trickles down
through the bed in
small rivulets.
Trickle bed reactor for treating waste
water
6. Fluidized bed reactor

When the packed beds

are operated in upflow
mode, the bed
expands at high liquid
flow rates due to
upward motion of the
particles.
Internal circulation patterns of fluidized Ca-alginate
beads containing immobilized cells of Z. mobilis. All
dimensions in cm.
Outer draft tube
Inner draft tube

1.176 2.620 4.530

4 Jets

28.40

21.30 26.43

6.895
2.876

0.1
0.953
 Miscellaneous reactors
• Vacuum fermenter
• Tray fermenter
• Stirred cascade reactor
 Vacuum Fermenter
Chilled
water

Condenser
Vacuum
Level pump
control
Vacuum
control
Receiving
tank
(product)

Dry ice
bath

Heating Fermenter
water
Filter

Medium
reservoir
Metering Receiving
Rheostat pump tank
Filter
(bleed)

Air or O2
Tray fermenter
Stirred cascade reactor
 Bioreactor Operation Modes
1. Batch Operation
•A foam breaker may be installed to disperse foam
A batch bioreactor is
normally equipped
with an agitator to
mix the reactant, and dC s rmax CS
r= =
the pH of the reactant dt K m + CS
is maintained by
employing either
buffer solution or a
Batch operation
pH controller
with stirring

+ (C s 0 − C s ) = rmax t
Cs 0
Change of Cs
K m ln
with time, t
Cs
 Bioreactor Operation Modes
-2. Plug-flow mode
An ideal plug-flow reactor can
approximate the long tube,
In a plug-flow packed-bed and hollow fiber or
reactor, the substrate multistaged reactor
enters one end of a
cylindrical tube with F, Cs0 V F, Cs
is packed with
immobilized enzyme V
and the product t=0 = Residence
time
steam leaves at the F
Continuous operation
other end.
without stirring

+ (C s 0 − C s ) = rmax t
Cs 0
K m ln
Cs
 Bioreactor Operation Modes
3. Continuous stirred-tank
A continuous stirred-
F, Cs0
tank reactor (CSTR)
is an ideal reactor
which is based on the
F, Cs
assumption that the V
reactants are well
mixed.
Continuous
operation with
stirring
 Bioreactor Operation Modes
3. Continuous stirred-tank reactor-Con.
Mass balance of substrate:
F, Cs0
Input - Output − Consumptio n = Accumulation

F, Cs
dCs
V
FC s 0 − FC s − rsV = V
dt
dCs
Steady state: =0
dt
Michaelis- rmax C S
r=
Menten rate: K m + CS

rmax Cs
FC s 0 − FC s − V =0
K m + Cs
 Bioreactor Operation Modes
3. Continuous stirred-tank reactor-Con.
Mass balance of substrate:
F, Cs0

rmax Cs
FC s 0 − FC s + V =0
F, Cs K m + Cs
V

F rmax C s
=
(
V (C s 0 − C s ) K m + C s )
F 1
=
V 
rmax C s
Cs = − K m +
Cs 0 − Cs
 Aeration and oxygen transfer in
bioreactor systems
• Bio-oxidation of substrate with molecular oxygen as
the final electron acceptor
 Oxygen transfer in fermentation
system
GAS FILM
GAS-LIQUD
INTERFACE

O2
CELL-LIQUD
O2 INTERFACE INTERNAL
CELL
AIR BUBBLE RESISTANCE
O2O2 CELL
LIQUID FILM
Dissolved O2
O2 in liquid phase,
nutrients
Electron
(medium mostly
Transport
water)
System +
TCA cycle
LIQUID FILM enzymes

The oxygen transport path to the microorganism. Generalized path of oxygen from the gas
bubble to the microorganism suspended in a liquid is shown. The various regions where a
transport resistance may be encountered are as indicated
 Oxygen transfer

• At Steady-state with no O2 accumulation in the liquid

phase:

• What are the O2 requirements of microorganisms?

– We define: QO2 = Respiration rate coefficient for a given
microorganism.
 Oxygen transfer
In general:
QO2 = f(microbial species and type of cell, age
of cell, nutrient conc. in liquid medium,
dissolved O2 conc., temperature, pH, etc.)
• For a given: 1) type of species of cell
2) age of cell
3) nutrient concentration
4) temperature
5) pH
 Oxygen transfer
and if O2 concentration, CL, is the limiting factor in cell
growth, then QO2 is a strong function of dissolved O2
concentration CL (= mg O2/L). The relationship between QO2
and CL is of the Monod type.
1
2
Q
O2ma
x

1
0

Q
O2
8
QO 2

Q
O2ma
x
6
/
2

2
K
O2
C
L C
RIT.
0
0 2 4 6 8 1
0 1
2 1
4 1
6 1
8 2
0
O
xyg
enC
ONC
. (C)
L

Respiration coefficient QO2 as a function of the dissolved oxygen concentration CL.

• where: KO2 = O2 conc. at QO2 max/2
CL CRIT. = Critical O2 conc. beyond which O2 is
not limiting
QO2 = QO2max = constant
• At CLCRIT. respiration enzymes of Electron Transport System are saturated with O2.

• When O2 conc. is the “limiting substrate” then

analogous to the Monod equation:

µmax.S
µ = ________ (S = substrate conc. (g/L)
KS + S

µ = 1 dX (h-1) [Ks = S (g/L), at µmax/2]

X dt
 Volumetric mass transfer coefficient
kLa and methods of measurement
GAS FILM
GAS-LIQUD
INTERFACE

a
UNIT LIQUID
VOLUME

AIR BUBBLE
O2 CL

O2 TRANSFER CELLS
(CONC. X)
kL OXYGEN
C *L BULK
(CONC. C )
L
LIQUID
LIQUID FILM
PHASE

Schematic diagram of the mass balance of oxygen transfer in unit liquid volume
Mass balance of oxygen in unit liquid
volume
Mass balance of oxygen in unit liquid
volume (cont’d)
Mass balance of oxygen in unit liquid
volume (cont’d)

At all the times CL = constant

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
The Sulphite Batch Oxidation Method.

2-
F, SO
3
In flu e n t

A ir o u tle t
M o to r

A ir flo w , ra te
rp m

W a te r o u t

W a te r in

Schematic diagram of a stirred tank batch reactor

 Chemical Methods of KLa Measurement
(Cont’d)
● Liquid Solution = 0.5 M Na2SO3 (Sodium
++
sulphite), with Cu as catalyst.

● Sparge air through the bioreactor vessel at a

given volumetric flow rate Q and impeller
speed (R.P.M.)

● -2
Make sure that [SO3 ] is in excess (i.e. 0.5 M
Na2SO3)
Chemical Methods of KLa Measurement (Cont’d)
Chemical Methods of KLa Measurement (Cont’d)
Chemical Methods of KLa Measurement (Cont’d)
 Chemical Methods of KLa Measurement (Cont’d)

-2
● The reaction with [SO3 ] is extremely
fast.
● As a result, the O2 gas molecules are
consumed as soon as they diffuse into
the liquid phase.
● Therefore, the D.O. concentration in the
liquid phase, CL  0.
Chemical Methods of KLa Measurement (Cont’d)
 Chemical Methods of KLa Measurement (Cont’d)

● Use iodometric titration to measure

-2
[SO3 ] as a function of time, t, as the air
bubbles pass through the bioreactor
vessel at a given R.P.M.
 Chemical Methods of KLa Measurement (Cont’d)

8
0
7
0 d[SO -2] ]2 [SO3-2]
SLOPE =
d -
[ -
SO
3 2
]
SLO PE =~- -
3
[ -
SO
3 ~-
d t dt t t
6
0
5
0
[SO3 ]
-2

4
0
3
0
2
0
1
0
0
0 2 4 6 8
T
IME,t
,(min
)
Concentration of SO3-2 as a function of oxidation time
Chemical Methods of KLa Measurement (Cont’d)
In Situ Measurement of KLa, QO2, and CL* During Cell
Growth in a Bioreactor

Consider a Stirred Tank Bioreactor System,

Where Cell Growth takes Place at a Given Set of
Conditions:
Aeration
Agitation
pH
Temperature
Medium Composition
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* 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* During
Cell Growth in a Bioreactor (Cont’d)

15 10

1. Feed
DO2 pH rpm 2. Flow meter
3. Ring sparger
5 4. Impeller
Alkali Acid 11 12 13 5. Motor
6
6. Shaft
7. pH probe
Water out 8. D.O. probe
2
7 8 9. Baffle
15 10. To Condenser
11. D.O. meter
1 12. pH meter
14
30 deg. 13. Speed controller
4 9
water in 14. Water Jacket
3 16 15. Thermometer
16. Chart recorder

Set up of a Stirred tank Bioreactor with Dissolved Oxygen Probe, pH probe and accessories.
 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.
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* During Cell
Growth in a Bioreactor (Cont’d)

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* During Cell
Growth in a Bioreactor (Cont’d)

• Fig. 2.11 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* During
Cell Growth in a Bioreactor (Cont’d)

AIR-OFF
CL STEADY-STATE
DO2 CONC. C L (mM O2/L)

CL,CRIT AIR-ON

TIME (MIN) 3-5

Transient Air-Off, Air-On Experiment in a Bioreactor System
 In Situ Measurement of KLa, QO2, and CL* During
Cell Growth in a Bioreactor (Cont’d)

● 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. 2.12.
4

SLOPE = - QO2 X
3
CL (mMO2/L)

0
0 1 2 3 4 5 6 7 8 9 10
Time, t (min)

D.O. concentration CL as function of time during AIR-OFF period.

In Situ Measurement of KLa, QO2, and CL* During Cell
Growth in a Bioreactor (Cont’d)

• 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* During Cell
Growth in a Bioreactor (Cont’d)
● Re-arranging KLa equation and solving for CL we get

● 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* 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* During
Cell Growth in a Bioreactor (Cont’d)

4.4
*
Intercept = CL
3.8
CL (mgO2/L)

3.2
SLOPE = -1/kLa
2.6
2.0
1.4
0.8
0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8
[dCL/dt+QO2 X]

D.O. concentration CL as function of [dCL/dt + QO2X] during AIR-ON period.

 In Situ Measurement of KLa, QO2, and CL*
During Cell Growth in a Bioreactor (Cont’d)

Dynamic air-on/air-off data during Poly(glutamic acid (PGA) production by Bacillus subtilis IFO 3335 (fermentation time =
26 h). Dissolved oxygen concentration CL ( ) as a function of time.

Taken from A. Richard and A. Margaritis, “Rheology, Oxygen Transfer, and Molecular Weight Characteristics of
Poly(glutamic acid) Fermentation by Bacillus subtilis”, Biotechnology and Bioengineering, Vol. 82, No. 3, p. 299-305
(2003).
Agitation requirement in fermentation systems

The dimensions of what is called a “standard” stirred tank

bioreactor vessel with Baffles.
 Geometric Ratios for a Standard Bioreactor
Vessel
Impeller Di/Dt HL/Dt Li/Di Wi/Di Hb/Di Wb/Dt No. Baffles
Type

Flat-Blade 0.33 1.0 0.25 0.2 1.0 0.1 4

Turbine

Paddle 0. 3 3 1.0 - 0.25 1.0 0.1 4

impeller

Marine 0.33 1.0 pitch = Di 1.0 0.1 4

Propeller

Where:
Dt = tank diameter,
HL = liquid height
Di = impeller diameter
Hb = impeller distance from bottom of vessel
Wb = baffle width
Li = impeller blade length
Wi = impeller blade height
Different Impeller Types. (a) Marine-type propellers; (b) Flat-blade turbine, Wi =
Di/5. © Disk flat-blade turbine, Wi = Di/5, Di = 2Dt/3, Li = Di/4; (d) Curved-blade
turbine, Wi = Di/3; (e) Pitched-blade turbine, Wi = Di/8; and (f) Shrouded turbine,
Wi = Di/8.
 Macam Gambar Agitator

Curved blade turbine Pitch Blade Turbine

 Lanjutan

Marine Type Propeller Flat Blade Turbine

 Lanjutan

Disk flat blade turbine Shrouded Turbine

Mixing Patterns for Flat-Blade Turbine Impeller. Effect of Baffles. Liquid agitation in presence
of a gas-liquid interface, with and without wail baffles: (a) Marine impeller and (b) Disk flat-
blade turbines; (c) in full vessels without a gas-liquid interface (continuous flow) and without
baffles.
 Mixing and Power Requirements for Newtonian
Fluids in a Stirred Tank

NP vs. NRe; the power characteristics are shown by the power number, NP, and the modified
Reynolds number, NRe, of single impellers on a shaft. [Adopted from S. Aiba, A.E. Humphrey
and N.F. Millis. “Bubble Aeration and Mechanical Agitation”. In Biochemical Engineering, 2nd
Ed., Academic Press, Inc., New York (1973) 174].
Last slides figure shows relationship between NP
and NRe at three different flow regimes:
● Laminar
● Transient
● Fully Turbulent
for three different impeller types:
● Six-bladed flat blade turbine
● Paddle impeller
● Marine Propeller
The power number is given by

NP = Pgc/n3Di5

The impeller Reynolds number is given by

NRe = nDi2/

Where:
NRe = dimensionless Reynolds number
NP = dimensionless Power number
P = Un-gassed power for liquid (no air), W
gc = 1, for SI units system
n = Impeller rotational speed, revolutions per
sec., (s-1)
Di = Impeller diameter, m
 = Density of liquid, kg/m3
 = Viscosity of liquid, (N.m)/(s)

For six-bladed flat-blade turbine impeller (cf.

Fig. 3.3), the mixing becomes fully turbulent at
an impeller Reynolds number NRe = 3,000.

Power number NP = 6 (constant) at NRe > 3,000

Different Types of impellers have different power
characteristics

For six-bladed flat turbine and for turbulent conditions:

NP = 6 = Pgc/n3Di5

or P = (6)(n3Di5)/(gc)

At NRe = 3,000 the corresponding impeller speed is:

n = (3,000)()/(Di2)()
● Earlier equation is an estimate of the minimum
impeller speed, n, of a 6-flat blade turbine impeller
for the on-set of turbulent flow within the stirred
tank bioreactor vessel.
● Equation also shows that for a fluid of a given
density, :
P  n3Di5

This is an important consideration for bioreactor

vessel scale-up.
Calculation of the Required Volumetric
Mass Transfer Coefficient, KLa, During
Fermentation, and Gassed Power, Pg.

At Steady-State Operation of an Aerobic

Fermentation:
OTR = OUR
KLa[CL* - CL] = QO2X
For a given QO2, X, and (CL* - CL), KLa can
be calculated using earlier equation

For a given VL and Ug, Pg can be calculated

using the empirical correlation for KLa given

KLa = C [Pg/VL]m [Ug]k