EFFECTS OF CARBON SOURCES

ON THE POLYSACCHARIDE AND

ENZYME PRODUCTION
DURING GROWTH OF
GANODERMA LUCIDUM

By: Redzuan B.Hussin (150957)

Supervised by: Dr. Helmi B. Wasoh @ Mohamed Isa

OVERVIEW

Introduction

Objectives

Problem statement and motivation

Literature review

Methodology

Result and discussion

Conclusion

Recommendation

References

INTRODUCTION

Ganoderma lucidum

It is a medicinal mushroom

It is well known as Lingzhi in Chinese, Reishi

in Japanese, and Youngzhi in Korean

It has been intensively studied because of its wide

range of biological activities

It can be cultivated in both solid-state and

submerged fermentation

Objectives

To study the effect of types of carbon source on

mycelial growth of Ganoderma lucidum and
production of EPS and IPS.
To study the effect of substrate variability on
cellulase enzyme activity in Ganoderma lucidum
culture.

Problem
statement

Currently, the role of different factors for the

growth of Ganoderma lucidum and the production
of its metabolites under the conditions of
submerged culture is poorly understood.

Motivation

The study will enhance the understanding about

several factors in submerged cultivation of
Ganoderma lucidum that will gradually increase
the cell growth and led to the highest production of
cell biomass, polysaccharides (EPS and IPS) and
cellulase enzymes of Ganoderma lucidum.

LITERATURE REVIEW

Individual compounds isolated from Ganoderma lucidum

exhibit hypolipidemic and bactericidal activities,
stimulate immunity, and are effective in treating diseases
of the cardiovascular and nervous systems (Stamets,
2000)

Submerged cultivation of Ganoderma lucidum has been

developed to obtain mycelial biomass, ganoderic acid
and polysaccharides which can be used to produce
medicinal products (Yang and Liau, 1998).

CONTINUE..

In recent studies on Ganoderma lucidum, bioactive

polysaccharides were isolated from the basidiocarps (Bao et
al., 2002; Zhang and Lin, 2004) and from the mycelial
biomass that cultivated in liquid culture (Kim et al., 1993).

It is widely accepted that cellulases secreted by fungi

consist of three major components which are
endoglucanases, (EGs) cellobiohydrolases (exocellulases,
CBHs) and -glucosidases (Zhou, Chen and Li, 2004).

METHODOLOGY
The mycelia of Ganoderma lucidum was cultured on the
agar plate
Ganoderma lucidum was cultivated in submerged
condition with using different types of carbon sources and
substrates
Ganoderma lucidum culture was cultivated for 30 days

Mycelial Growth

Polysaccharides Analysis
IPS - extracted from mycelia
EPS- extracted from culture
media

Cellulase Analysis
Fpase
CMCase
- glucosidase

Firstly the mycelia was cultivated on the agar plate for

10-15 days which is the period for the mycelia to fill the
entire plate.

For submerged cultivation of ganoderma lucidum, about

6 plug of the agar containing mycelia were inoculated
into the flask that contain standard medium with various
type of carbon source.

The ganoderma lucidum culture was cultivated for 30

days on rotary shaker at150rpm and 26 degree. Then,
several types of analysis were carried out on the
ganoderma lucidum culture during the cultivation period.

There are three main analyses that were done

Mycelial growth of ganoderma lucidum

Polysaccharides analysis

Cellulase analysis

RESULT AND DISCUSSION

Figure 1: Mycelial Growth of Ganoderma lucidum with Different Types of
Carbon Sources
18
16
14
12
Gluco
se
Lacto
se

10
Mycelial8Dry Weight of G. Lucidum (g/l)
6
4
2
0
0

10

15

20

25

30

Cultivation Time (Days)

Lactose as carbon source showed the highest mycelial growth of

Ganodema lucidum

Based on this figure 1, mycelial growth of Ganoderma

lucidum was proportionally increased with time until the day
10 of cultivation time. It reached the maximum growth at
day 10 and started to decrease after that.

The decreases of the mycelial growth were occurred due to

carbon source exhaustion so that the cellular energy that
needed for the mycelial growth cannot be generated

Lactose was more preferable rather than other carbon

compounds to be utilized by the mycelia because of higher
metabolization rate of lactose by ganoderma lucidum to
produce cellular energy that needed for the mycelial growth.

CONTINUE..
Figure 2: The Production of Extracellular Polysaccharides of Ganoderma
Lucidum Culture with Different Types of Carbon Sources
0.8
0.7

0.69
0.63

0.6

0.51

0.5

Lactose
Glucose
Sucrose

0.4
Production of EPS (g/l)
0.3
0.2
0.1
0.0
0

10

15

20

25

30

Cultivation Time (days)

Sucrose as a carbon source showed the highest production of EPS in

Ganodema lucidum culture

According to Figure 2, the production of EPS was proportionally increased

with time until the day 10 of cultivation time. At day ten, the production of
EPS reached the stationary phase and has not significantly changed until
the day 30 of the cultivation time. The maximum production of EPS was
obtained when sucrose was used as a carbon source at day 30 of cultivation
time

It shows that sucrose was favorable for production of EPS since it

produces maximum amount of EPS during cultivation of ganoderma
lucidum.

It is believed that, many short mycelia were observed when sucrose was
used as carbon source. It is because sucrose is not good for mycelial
growth and exhibit the morphology changes of mycelia. This marphology
changes will inhibit the mycelial growth and increase the amount of
polysaccharide secreted out into the medium. So, the production of EPS is
higher.

CONTINUE..
Figure 3: The Production of Intracellular Polysaccharides of Ganoderma
Lucidum Culture with Different Types of Carbon Sources
1.4

1.26

1.2
0.99

1.0

Lactos
e
Sucros
e

0.8
Production of IPS (g/l)
0.6
0.4

0.25

0.2
0.0
0

10

15

20

25

30

Cultivation Time (days)

Lactose as a carbon source showed the highest production of IPS in

Ganodema lucidum culture

Production of IPS is slightly related with mycelial growth

Ganoderma lucidum since it was extracted from the mycelia
itself. So, when the mycelial growth is higher, the production
of IPS is higher as well.

It shows that production of IPS was increased with time until

the day 10 and started to decrease after that. This because of
carbon source exhaustion so that the production of IPS
became slower

The preference of lactose to be utilized by the mycelia rather

than other carbon source may be due to the fast
metabolization of lactose by the mycelia to produce
polysaccharides easily.

CONTINUE..
Figure 4: Fpase activity of Ganoderma Lucidum Culture with
Different Types of Substrates
3.0
2.5
2.0
OPEFB

1.5
Fpase Activity (U/ml)

Cellulose
Wheat

1.0

Control

0.5
0.0
0

10

15

20

25

30

Cultivation Time (days)

Wheat as a substrate showed the highest Fpase activity

in Ganodema lucidum culture

According to Figure 4, it showed that Fpase

activity was proportionally increased with time
and reach the maximum point at day 20. After
that, the Fpase activity was decreased.
The production of cellulase is directly related to
mycelial growth higher mycelial growth,
higher cellulase production

The pattern for all cellulase activity were almost

the same where the optimum activity of the
enzyme were obtained during the day 20. This is
probably due to highest mycelial growth during
that period.

CONTINUE..
Figure 5: CMCase activity of Ganoderma Lucidum Culture
with Different Types of Substrates
9.0
8.0
7.0
6.0
5.0

OPEFB

CMCase
4.0 Activity (U/ml)

Cellulose
Wheat

3.0

Control

2.0
1.0
0.0
0

10

15

20

25

30

Cultivation Time (days)

Wheat as a substrate showed the highest CMCase

activity in Ganodema lucidum culture

The fact that the wheat yielded highest Fpase

and CMCase activities, as compared with OPEFB
and pure cellulose, may be related to their
composition, namely to the presence of high
concentrations of soluble sugars and the easy
availability of their polysaccharides which
promoted an abundant growth of the mycelia.

High mycelial growth will exhibit more enzyme

inducer that will promote the secretion of
cellulase enzyme from the Ganoderma lucidum
culture.

CONTINUE..
Figure 6: -glucosidase activity of Ganoderma Lucidum
Culture with Different Types of Substrates
0.40
0.35
0.30
0.25
OPEFB
0.20
-Glucosidase Activity (U/ml)

Cellulose
Wheat

0.15

Control

0.10
0.05
0.00
0

10

15

20

25

30

Cultivation Time (days)

OPEFB as a substrate showed the highest glucosidase activity in Ganodema lucidum culture

According to figure 6

It was proven that the lignocellulosic substrates

such as OPEFB contain specific

aromatic compounds or

microelements liberating during fermentation

or

some specific compounds appeared during

substrates fermentation

which stimulated cellulolytic enzyme synthesis.

The presence of those extractive substances,

derived from OPEFB, was essential for the
production of B-glucosidase by Ganoderma
lucidum.

Consequently, according the results obtained for

the analysis of cellulase production in
Ganoderma lucidum culture, it showed that the
production of CMCase during the cultivation
period was the highest followed by Fpase while
B-glucosidase was the lowest one .

CONCLUSION

The usage of suitable types of carbon sources and

substrates gradually increase the cell growth and led to
the highest production of cell biomass, polysaccharides
(EPS and IPS) and cellulase enzymes of Ganoderma
lucidum culture.

Lactose was determined as the best carbon source for

mycelial growth and the production of IPS while sucrose
was more favorable for the production of EPS.

For cellulase activity analysis, wheat yielded the

highest activity of Fpase (2.783 U/ml) and CMCase
(7.805 U/ml) while OPEFB produce highest activity of glucosidase (0.348 U/ml).

RECOMMENDATION

There are several factors could be studied in order

to enhance the knowledge/ understanding about
the growth of Ganoderma lucidum under
submerged cultivation condition. (e.g. pH effect,
nitrogen source effect, temperature effect,
agitation effect and etc.)

The productivity of valuable metabolites from

Ganoderma lucidum culture can be increased
maximally by optimizing the culture condition for
the growth of Ganoderma lucidum.

REFERENCES

Stamets, P. (2000). Growing Gourmet and Medicinal

Mushrooms. Stamets, P., Ed., Berkley: Ten Speed
Press .

Yang, F.C. and Liau, C.B. (1998). The influence of

environmental conditions on polysaccharide
formation by Ganoderma lucidum in submerged
cultures. Process Biochemistry , 547-553.

Bao X.F., Wang X.S., Dong Q, Fang J.N., Li X.Y.

(2002). Structural features of immunologically
active polysaccharides from Ganoderma lucidum
(italic). Phytochemistry , 175-181.

Zhou, X., Chen, H. and Li, Z. (2004). CMCase activity

assay as a method for adsorption analysis.
Enzyme and Microbial Technology , 455-459.

THANK YOU!