Kinetics of Microbial Growth and Synthesis of

Poly(3-hydroxybutyrate) Using Fed-batch

Culture of Alcaligenes Eutrophus with
Phosphorus Limitation
Author(s) : Li Hongqi ; Ryu Hee Wook; Chang Yong Keun;
Chang Ho Nam; Shen Zhongyao

Presentation By: Dalupo, Jillian A.

Abstract
Phosphorous concentration in the culture medium affects the
production of poly (3-hydroxybutyrate) by Alcaligines Eutrophus.

Phosphorus concentration in the initial culture medium influences

both the residual cell mass and the PHB production.
INTRODUCTION

Poly(3-Hydroxybutyrate)
Intracellular carbon and energy storage in many
microorganism
INTRODUCTION
Poly(3-Hydroxybutyrate) is a biodegradable thermoplastic
polyester if the cell growth is impaired by the depletion of
a specific nutrient such as N, P, S, Mg or O.

Study is all about the Phosphorus concentration on Poly(3-

Hydroxybutyrate) synthesis
INTRODUCTION

Kinetic model of PHB production was proposed to describe

the effect of phosphate concentration on a fed-batch
culture of Alcaligenes Eutrophus by the phosphorus
limitation method.
Materials and Methods

Alcaligenes
Eutrophus
Used to produce Poly(3-
Hydroxybutyrate
Materials and Methods

For the fed-culture, initial mediums contained (per Liter) :

o Glucose (C6 H12 O6 ): 20 g

o Ammonium Sulfate ((NH4 )2 SO4 ): 4 g
o Magnesium Sulfate Heptahydrate (MgSO4 ∙ 7H2 O): 1.2 g
o Citric Acid (C6 H8 O7 ): 1.7 g
o Trace element solution : 10 mL
o Potassium Di-hydrogen Phosphate (KH2 PO4 ): 1.5 g/L to 4.5 g/L
Materials and Methods
Prepare seed culture in flasks mounted om a reciprocal shaker
30◦C for 1 to 2 days.

100 mL of seed culture was used to inoculate the 2. 5L fermentor

equipped with two six-bladed disk-turbine impellers and three
baffles

Controlled Temperature at 30◦C

Materials and Methods
PH is at 6. 8 using 3 mol/L HCl solution and 28% NH4 OH solution

Dissolved oxygen concentration is above 10% of air saturation by

increasing the agitation speed up to 1000 r/min and the aeration
rate up to 5. 0 L/min

Glucose concentration in the broth was maintained at 10 g/L by

feeding 800 g/L glucose solution
Materials and Methods

Phosphate concentration was measured using Taussky's method

Computer was used to simulate and evaluate the unknown

parameters in the model using Runge Kutta Method

Add KH2 PO4 to the initial culture medium to adjust the phosphate
concentration in the fementer from 1.5 g/L to 4. 5 g/L
Results and Discussion

Fig. 1 Fed-Batch fermentation of

Alcaligenes Eutrophus with
Phosphate concentration of 4
g/L
Results and Discussion
Two phases can be clearly recognized:

o Cell growth phase

o PHB storage phase.

Experimental results showed that higher phosphate

concentration in the broth caused the residual cell rate to
increase but inhibited PHB production.

Ammonia was not completely exhausted during the fed-batch

fermentation
 Mathematical Modelling
𝑑(𝑅𝑉) 𝜇𝑚𝑎𝑥 𝑆
= (𝑅𝑉) Model proposed in this study was
𝑑𝑡 𝐾1 + 𝑆
obtained by augmenting a previous
𝑑(𝑅𝑉) 𝛽𝐾2 model for PHB production
= (𝑅𝑉)
𝑑𝑡 𝐾2 + 𝑆 developed using the nitrogen
limitation method.
𝑑 𝑆𝑉 1 𝑑(𝑅𝑉)
− =
𝑑𝑡 𝑌1 𝑑𝑡
𝑑 𝐺𝑉 1 𝑑 𝑅𝑉 1 𝑑 𝑅𝑉
− = + + 𝐾𝑚 𝑅𝑉 − 𝐹
𝑑𝑡 𝑌2 𝑑𝑡 𝑌3 𝑑𝑡

𝑋 =𝑅+𝑃
Model Simulation

μmax and K1 were determined by a goodness-of-fit method

Y1 , Y2 , Y3 and K m were obtained from experimental data

K 2 and β were evealuated by solving model equations

Model Simulation

Simulation results are in good

agreement with the
experimental data for the
entire process
Model Simulation

Model estimates did not

deviate substantially from the
experimental data when these
three parameters were
changed by ± 10%

Changes in K1 have the most

significant effect on the results
Conclusion
Model can be used to predict the optimum conditions for
achieving a higher Poly(3-Hydroxybutyrate) content and Poly(3-
Hydroxybutyrate) productivity
Reference:
Li, Ryu, H.Chang, Y.Chang and Sheng, Kinetics of Microbial
Growth and Synthesis of Poly(3-hydroxybutyrate) Using Fed-
batch Culture of Alcaligenes Eutrophus with Phosphorus
Limitation, Tsinghua Science and Technology, China, 1998,