Bio-Process KineticsBio-Process KineticsProcess BiotechnologyProcess Biotechnology

Anondho WIJANARKOAnondho WIJANARKOUniversity of IndonesiaUniversity of Indonesia

Simplified Lecture NotesSimplified Lecture NotesFor InstructorFor Instructor

To Dwi ImantiTo Dwi Imantiandand

Adam Aulia, Musa Maulana, Isa Dzulqarnain Adam Aulia, Musa Maulana, Isa Dzulqarnain and Lovely Muhammad Al Muhammad and Lovely Muhammad Al Muhammad

CONTENTSCONTENTS

Microbial GrowthMicrobial Growth– Microbial Cell GrowthMicrobial Cell Growth– Classical & Empirical Growth KineticsClassical & Empirical Growth Kinetics

Environmental Alteration StudiesEnvironmental Alteration Studies– Light Illumination EffectLight Illumination Effect– Temperature EffectTemperature Effect

Microbial Kinetic StudiesMicrobial Kinetic Studies– Non Elementer ReactionNon Elementer Reaction– Microbial Growth Reaction KineticsMicrobial Growth Reaction Kinetics

Microbial GrowthMicrobial Growth

Microbial Cell GrowthMicrobial Cell Growth Mode of GrowthMode of Growth

– Selective assimilation of nutrients and Selective assimilation of nutrients and convert into and also include Chemical convert into and also include Chemical rearrangement of protoplasmic material rearrangement of protoplasmic material characteristic of the particular organismcharacteristic of the particular organism

– Production of an increased amount of Production of an increased amount of nuclear substance and cell divisionnuclear substance and cell division

Growth PhaseGrowth Phase

Growth PhaseGrowth Phase

Induction Phase (Lag Phase)Induction Phase (Lag Phase) Transient Phase (Acceleration Phase)Transient Phase (Acceleration Phase) Exponential PhaseExponential Phase Stationary Phase (Declining Phase)Stationary Phase (Declining Phase) Death PhaseDeath Phase

Question SheetQuestion Sheet

Why microbial growth have an lag phase?Why microbial growth have an lag phase? Why death phase could be occurred in microbial Why death phase could be occurred in microbial

growth?growth? What is essential nutrient for growth of What is essential nutrient for growth of

organism especially organism especially prokaryotesprokaryotes?? What is important factor for cell division?What is important factor for cell division?

Growth ApproximationGrowth Approximation

Growth ConstantsGrowth ConstantsExponential – Stationary Growth PhaseExponential – Stationary Growth PhaseTotal Biomass Production(G)Total Biomass Production(G)

Incident growth rate, Incident mean division rate (Incident growth rate, Incident mean division rate ())

Specific growth rate, Beginning mean division rate (Specific growth rate, Beginning mean division rate ())

Doubling time of population (Doubling time of population (DD); exponential growth ); exponential growth phasephase

dt

dX

Xtt

X

X

1

12

ln1

2

dt

dX

Xt

1lim

0

)( im XXG

2ln

D

Classical Growth KineticsClassical Growth KineticsEmpirical ApproximationEmpirical Approximation

Monod Monod Growth KineticGrowth Kinetic

Tessier Tessier Growth KineticGrowth Kinetic

MoserMoser Growth Kinetic Growth Kinetic

ContoisContois Growth Kinetic Growth Kinetic

sΚ

s

s μμ

11

sK s

sKse /1

sXB

s

Empirical Growth KineticsEmpirical Growth KineticsMedium constituent InhibitionMedium constituent Inhibition

AndrewsAndrews Growth Kinetic Growth Kinetic

AibaAiba Growth Kinetic Growth Kinetic

i

2

Ks

sK

s

s

pKsK

Ks

ps

p

Growth KineticsGrowth KineticsMultiple essential nutrientMultiple essential nutrient

Bailey Growth KineticBailey Growth Kinetic

...33

3

22

2

11

1

sK

s

sK

s

sK

s

sss

Home WorkHome Work

Which kinetic approximation do you choose in case of Which kinetic approximation do you choose in case of microbial growth of Hepatotoxin produced microbial growth of Hepatotoxin produced Oscilatoria Oscilatoria AgardhiiAgardhii NIVA CYA 97 in low temperature? NIVA CYA 97 in low temperature?

Which empirical equation that you choose of Which empirical equation that you choose of inoculation of microorganism in case of multiple inoculation of microorganism in case of multiple content limitation of nutrients, such as Mgcontent limitation of nutrients, such as Mg2+2+, phosphate, , phosphate, Nitrate and organic compound?Nitrate and organic compound?

Which kinetic approximation do you choose of Which kinetic approximation do you choose of cultivation photosynthetic microorganism that did not cultivation photosynthetic microorganism that did not grew up in pH above 7.8? grew up in pH above 7.8?

Simple Bio-Production KineticSimple Bio-Production Kinetic Cellular growth rateCellular growth rate

– Monod approximationMonod approximation

– Yield factorYield factor

Substrate UtilizationSubstrate Utilization

Product FormationProduct Formation

(Beginning of (Beginning of Stationary PhaseStationary Phase))

sΚ

s

s μμ

dt

ds

dtdX

sX s

XY

/

Xμdt

Xd

dt

dX

dtdP

XP X

PY

/

sK

Xsμ

dt

Xd

s

sK

Xs

Y

μ

dt

sd

ssX

/

sK

XsY

dt

Pd

sXP

/

EnvironmentalEnvironmental Alteration Studies Alteration Studies

Microbial Growth KineticMicrobial Growth KineticEnviromental ConditionEnviromental Condition

Direct EffectsDirect Effects– Light Illumination (Energy Source)Light Illumination (Energy Source)– TemperatureTemperature– Essential nutrients contentEssential nutrients content

Indirect EffectsIndirect Effects– Gas inlet volumetric rateGas inlet volumetric rate– Gas inlet content Gas inlet content – Liquid circulation rateLiquid circulation rate– Non essential nutrients contentNon essential nutrients content

Light Illumination EffectLight Illumination Effect Oscillatoria agardhiOscillatoria agardhi Gomont Gomont

– (Post AF, R de Witt, LC Mur, J. Plank. Res., 7 (1985) 487-495)(Post AF, R de Witt, LC Mur, J. Plank. Res., 7 (1985) 487-495)

Temperature effectTemperature effect Modified Modified ArheniusArhenius Model Model

RdH

RdS

a

T

RTE

e

TeA

1

/

Temperature EffectTemperature EffectClassification of MicroorganismClassification of Microorganism

Question SheetQuestion Sheet

What is happen if microorganism is at 90What is happen if microorganism is at 90ooC? Why?C? Why? In case of decreasing of temperature about 20In case of decreasing of temperature about 20ooC from C from

optimum temperature, what is happen in case of optimum temperature, what is happen in case of microbial growth rate?microbial growth rate?

In case of ethanol production that was In case of ethanol production that was S. sakeS. sake have have ethanol tolerance around 10%, what do you do to make ethanol tolerance around 10%, what do you do to make an whisky industry?an whisky industry?

Why a shade microbe does not grew well in high light Why a shade microbe does not grew well in high light illumination and commonly have not high temperature illumination and commonly have not high temperature resistance?resistance?

Temperature EffectTemperature EffectCellular ConsiderationCellular Consideration

Psychrophile Psychrophile – ObligateObligate

Protococcus Agardh SS 100-3Protococcus Agardh SS 100-3 Oscillatoria redekei Van GoorOscillatoria redekei Van Goor Oscillatoria sp. SS 100-5Oscillatoria sp. SS 100-5

– FacultativeFacultative Anabena cylindricaAnabena cylindrica Lemmerman Lemmerman Oscillatoria AgardhiOscillatoria Agardhi Gomont Gomont Nostoc communeNostoc commune Antartica Antartica

MesophileMesophile– Synechococcus leopoliensisSynechococcus leopoliensis– Anabaena variabilisAnabaena variabilis IAM M3 IAM M3– Microcystis AeruginosaMicrocystis Aeruginosa IAM M228 IAM M228

ThermophileThermophile– Mastigocladus laminosusMastigocladus laminosus HTF HTF– Synechococcus lividusSynechococcus lividus OH75S OH75S– Synechcocus elongatus It 7SSynechcocus elongatus It 7S

Home WorkHome Work

Why optimum specific growth rate values of Why optimum specific growth rate values of psychrophile factually, lower than thermophile?psychrophile factually, lower than thermophile?

Why GC content of microbial DNA is important Why GC content of microbial DNA is important for classification of organism in terms of growth for classification of organism in terms of growth rate dependence on temperature?rate dependence on temperature?

What is DBI?What is DBI?

Microbial Kinetic StudiesMicrobial Kinetic Studies

Non Elementer ReactionNon Elementer Reaction

Common reaction rateCommon reaction rate

– N = integer N = integer ElementerElementer

– N = non integer N = non integer Non ElementerNon Elementer

Non Elementer Example :Non Elementer Example :

nskdt

ds

HBrBrH 222 22

22

3

212

'][

'

BrkHBr

HBrk

dt

Brd

COCHCHOCH 43 23

33 CHOCHkdt

CHOCHd

Reaction MechanismReaction Mechanism

22

1

1

2 ONNOk

k

2222 / OHON

022

dt

ONd

OHOHHk

2222 23

022

dt

OHd

OHNHNO 222 222

222222

2

OHNHONk

022232222 HOHkHONk 223

222 ON

k

kOH

2

22

21

212

2

2

21

122222

2

11 H

HNO

kk

kkH

H

NO

kk

kkHONk

dt

Nd

022222212

1 HONkONkNOk 2

2

21

122 1 H

NO

kk

kON

Question SheetQuestion Sheet

What is mechanism path?What is mechanism path? What definition of intermediate species?What definition of intermediate species? What was become determining factor of reaction What was become determining factor of reaction

rate?rate?

Microbial GrowthMicrobial GrowthEnzymatic Reaction/Kinetic considerationEnzymatic Reaction/Kinetic consideration

Michaelis-MentenMichaelis-Menten Kinetics Kinetics– Reaction mechanismReaction mechanism

– Kinetic equationKinetic equation

Substrate Actvation and InhibitionSubstrate Actvation and Inhibition– Reaction mechanismReaction mechanism

– Kinetic equationKinetic equation

Product Activation and InhibitionProduct Activation and Inhibition– Reaction mechanismReaction mechanism

– Kinetic equationKinetic equation

sK

s

m

ESSEk

k

1

1

EPESk

2

ESSEk

k

1

1

2

2

2

ESSESk

k

EPESk

3

im KssK

s

/2

ESSEk

k

1

1

EPESk

2

ESPPESk

k

3

3

pKsK

Ks

ps

p

Michaelis-Menten KineticsMichaelis-Menten Kinetics

Reaction mechanismReaction mechanism

Kinetic derivationKinetic derivation

ESSEk

k

1

1

EPESk

2

ESEE 0

ESESkk

kES

kk

kES

ESkkESkES

ES

dtd

dtd

021

1

21

1

211

0

00 ES

m

dtd

dtd

KS

SP

kkk

S

SEkESkP

kkk

S

SEES

max

1

21

022

1

21

0

Home WorkHome Work

Please exhibit kinetic derivation of substrate Please exhibit kinetic derivation of substrate activation and inhibition?activation and inhibition?

Please exhibit kinetic derivation of product Please exhibit kinetic derivation of product activation and inhibition?activation and inhibition?

What do you think about reaction kinetic if KWhat do you think about reaction kinetic if Kmm is is

high that was indicated in bioremediation of high that was indicated in bioremediation of toluene by toluene by C. nivalisC. nivalis??

LiteratureLiterature

1.1. Aruga Y., Aruga Y., Ecological Studies of Photosynthesis and Matter Production of Ecological Studies of Photosynthesis and Matter Production of Phytoplankton I: Seasonal Changes in Photosynthesis of Natural Phytoplankton I: Seasonal Changes in Photosynthesis of Natural PhytoplanktonPhytoplankton, Bot. Mag. Tokyo, 78 (1965) 280-288, Bot. Mag. Tokyo, 78 (1965) 280-288

2.2. Bailey JE and DF Ollis, Bailey JE and DF Ollis, Biochemical Engineering FundamentalsBiochemical Engineering Fundamentals, McGraw-, McGraw-Hill Book Co., New York, 1986Hill Book Co., New York, 1986

3.3. Boney AD, Boney AD, PhytoplanktonPhytoplankton, Edward Arnold Publ. Ltd., London, 1975, Edward Arnold Publ. Ltd., London, 19754.4. Contois DE, Contois DE, Kinetics of Bacterial Growth: Relationship between Population Kinetics of Bacterial Growth: Relationship between Population

Density and Specific Growth Rate of Continuous CulturesDensity and Specific Growth Rate of Continuous Cultures , J. Gen. , J. Gen. Microbiol., 21 (1959) 40-50Microbiol., 21 (1959) 40-50

5.5. Castenholz RW, Castenholz RW, Laboratory Cultures of Thermophilic CyanophyteLaboratory Cultures of Thermophilic Cyanophyte , , Symposium uber Cyanophytensystematik, 32 (1970) 538551Symposium uber Cyanophytensystematik, 32 (1970) 538551

6.6. Foy RH, CE Gibson and RV Smith, Foy RH, CE Gibson and RV Smith, The Influence of Day-length, Light The Influence of Day-length, Light Intensity and Temperature, on the Growth of Planktonic Blue-Green AlgaeIntensity and Temperature, on the Growth of Planktonic Blue-Green Algae , J , J Phycol., 11 (1976) 151-163Phycol., 11 (1976) 151-163

7.7. Frohlich BT, IA Webster, MM Ataai and ML Shuler, Frohlich BT, IA Webster, MM Ataai and ML Shuler, Photobioreactors: Photobioreactors: Model of Interaction of Light Intensity, Reactor Design and Algal PhysiologyModel of Interaction of Light Intensity, Reactor Design and Algal Physiology , , Biotechnol. Bioeng. Symp., 13 (1983) 331-350Biotechnol. Bioeng. Symp., 13 (1983) 331-350

8.8. Leopold AC and PE Kriedmann, Leopold AC and PE Kriedmann, Plant Growth and DevelopmentPlant Growth and Development, Tata , Tata McGraw-Hill Publ. Co., New Delhi, 1975McGraw-Hill Publ. Co., New Delhi, 1975

9.9. Pilling MJ and PW Seakins, Pilling MJ and PW Seakins, Reaction KineticsReaction Kinetics, Oxford Sc. Publ., Tokyo, Oxford Sc. Publ., Tokyo10.10. Mosser JL and TD Brock, Temperature Optima for Algae Inhabiting Cold Mosser JL and TD Brock, Temperature Optima for Algae Inhabiting Cold

Mountains Stream, Arctic and Alpine Research, 8 (1976) 111-114Mountains Stream, Arctic and Alpine Research, 8 (1976) 111-11411.11. Murata N, Murata N, Low Temperature effects on Cyanobacterial MembranesLow Temperature effects on Cyanobacterial Membranes, J. , J.

Bioenerg. Biomembr., 21 (1989) 61-75 Bioenerg. Biomembr., 21 (1989) 61-75 12.12. Post AF, R de Witt and LR Muur, Post AF, R de Witt and LR Muur, Interaction between Temperature and Interaction between Temperature and

Light Intensity on Growth and Photosynthesis of the Cyanobacterium Light Intensity on Growth and Photosynthesis of the Cyanobacterium Oscilatoria agardhiiOscilatoria agardhii, J. Plankton Research, 7 (1985) 487-495 , J. Plankton Research, 7 (1985) 487-495

13.13. Sato N, N Murata, Y Miura and N Ueta, Sato N, N Murata, Y Miura and N Ueta, Effect of Growth Temperature on Effect of Growth Temperature on Lipid and Fatty Acid Composition in the Blue Green Algae, Lipid and Fatty Acid Composition in the Blue Green Algae, Anabaena Anabaena variabilisvariabilis and and Anacystis nidulansAnacystis nidulans, Biochim. Biophys. Act., 572 (1979) 19-, Biochim. Biophys. Act., 572 (1979) 19-28 28

14.14. Seaburg KG, BC Parker, RA Wharton Jr. and GM Simmons Jr., Seaburg KG, BC Parker, RA Wharton Jr. and GM Simmons Jr., Temperature-Growth Responses of Algal isolates from Antartica OasesTemperature-Growth Responses of Algal isolates from Antartica Oases, J. , J. Phycol., 17 (1981) 353-360Phycol., 17 (1981) 353-360