THAI NGUYEN UNIVERSITY
UNIVERSITY OF AGRICULTURE AND FORESTRY

HOANG THI MAI
Topic title:
ALGAL CELL CULTURE IN MICROFLUIDIC DEVICES AND
MICROENVIRONMENT

BACHELOR THESIS

Study Mode :

Full-time

Major

:

Biotechnology

Faculty

:

Biotechnology and Food Technology

Batch

:

2013 – 2017

Thai Nguyen, 12/6/2017


THAI NGUYEN UNIVERSITY
UNIVERSITY OF AGRICULTURE AND FORESTRY

HOANG THI MAI
Topic title:
ALGAL CELL CULTURE IN MICROFLUIDIC DEVICES AND
MICROENVIRONMENT

BACHELOR THESIS

Study Mode :

Full-time

Major

:

Biotechnology

Faculty

:

Biotechnology and Food Technology


Batch

:

2013 – 2017

Supervisors :

Dr. Panwong Kuntanawat
Dr. Nguyen Xuan Vu
Mr. Phongsakorn Kunhorm

Thai Nguyen, 12/6/2017


DOCUMENTATION PAGE WITH ABSTRACT
Thai Nguyen University of Agriculture and Forestry
Major

Biotechnology

Student name

Hoang Thi Mai

Student ID

DTN1353150021

Thesis title


Algal cell culture in microfluidic devices and
microenvironment

Supervisors

1. Dr. Panwong Kuntanawat
2. Dr. Nguyen Xuan Vu
3. Mr.Phongsakorn Kunhorm

Abstract:
Arthrospira platensis is a filamentous multicellular cyanobacterium that has
two distinct shapes: helical and straight filaments. They have high nutritional
value, chemical composition such as protein, pigments, antioxidant, fatty acids.
Microfluidics devices that were applied in various fields such as biological,
biomedical, biotechnology and chemical analyses. A.platensis was captured in
the microfluidics devices in order to observed activation, fragmentation time,
change color, life cycles. It was performed with total 20 filaments (10 filaments
of C005 str and 10 filaments of Central Lab str) in two different conditioned
medium. The result was based on measure length to comparison growth length,
fragmentation time, growth rate of filament and strain. In the standard
Zarrouk’s medium, length and growth rate of Central Lab str is faster than C005
str, fragmentation time is the same. In the stationary from cell culture:
Fragmentation was expressed with two filaments of C005 str (rate 40%) and
three filaments of Central Lab str (rate 60%). Moreover, the growth rate of
Central Lab str was faster than C005 str. The both strains of standard Zarrouk’s
medium were grew faster than Zarrouk’s stationary from cell culture.
Key words

C005 str, Central Lab str, microfluidic devices,

growth length, fragmentation time, growth rate.

Number of pages

38

i


ACKNOWLEDGEMENT
Foremost, I would like to express my deep and sincere gratitude to my
supervisor Dr. Panwong Kuntanawat from the School of Bioresources and
Technology, King Mongkut’s University of Technology Thonburi (KMUTT),
Thailand, for providing me the opportunity to conduct research in his lab and
giving me endless support in the past six months. His insights, wisdoms, advices
and enthusiasm for research have greatly influenced me and made the completion
of my dissertation possible.
I would also like to thank Dr. Nguyen Xuan Vu from the Faculty of
Biotechnology and Food of Thai Nguyen University of Agriculture and Forestry
(TUAF) who used to help, support and give me encouragements during this
thesis implementation. I would also like to extend my heartfelt thanks to my
lectures of Biotechnology and Food Department, TUAF who imparted me a lot
of knowledge through four years of university. The knowledge not only helped
me with my research, but also created a basic and soul foundation for me to start
the job in the future. Further, I would also like to express my sincere gratitude to
Ms. Trinh Thi Chung for providing me the opportunity to develop my skills by
doing an internship abroad.
I sincerely thank to the teachers, the laboratory staffs and students at the
laboratory for their regards and giving me an opportunity to do research in the
laboratory. I would also especially thank Mr. Phongsakorn Kunhorm who always

helped, cared, instructed and taught me during my practicing in Thailand.
Finally, I would like to thank my family and my friends for their love and
support. I could not have done this without you.
Many thank and best regards
Student

Hoang Thi Mai

ii


TABLE OF CONTENT

PART 1. INTRODUCTION ................................................................................ 1
1.1. Background ........................................................................................................................2
1.1.1. Microalgae ................................................................................................... 2
1.1.2. General Arthrospira platensis ..................................................................... 4
1.1.2.1. Morphology and taxonomy for Arthrospira platensis ............................. 5
1.1.2.2. Effect of temperatures .............................................................................. 6
1.1.2.3. Effect of pH .............................................................................................. 6
1.1.3. Microfluidics devices .................................................................................. 7
1.1.3.1. An introduction to soft lithography .......................................................... 8
1.1.3.2. Advantages of microfluidic for cell culture ............................................ 9
1.1.3.3. Microfluidic devices for cell biology ..................................................... 10
1.1.3.4. Microfluidic devices for single cell analysis .......................................... 11
1.2. Objectives ........................................................................................................................11
1.3. Scope of study ................................................................................................................12
PART 2: METHODS ......................................................................................... 13
2.1. Equipments and materials............................................................................................13
2.1.1. Equipments ................................................................................................ 13

2.1.2. Materials .................................................................................................... 13
2.1.2.1. Medium culture ...................................................................................... 13
2.1.2.2. Algal strains ............................................................................................ 14
2.1.2.3. Microfluidic devices design: an electrostatic using microwell based
microfluidic devices. ........................................................................................... 15
2.2. Methods ............................................................................................................................16
2.2.1. Algal strains cultivation ............................................................................. 16
iii


2.2.2. Make microfluidic devices ........................................................................ 17
2.2.3. Cell loading and cultivation in the microwell ........................................... 18
2.2.4. Imaging of cells and analysis methods ...................................................... 19
PART 3. RESULTS AND DISCUSSION ......................................................... 21
3.1. Cells/filaments in the standard Zarrouk’s medium ...............................................21
3.1.1. Comparison of growth length of single filament before fragmenting ....... 21
3.1.2. Compare fragmentation time of single filaments ...................................... 22
3.1.3. Comparison of growth rate of single filaments ......................................... 23
3.2. Filaments in the Zarrouk’s medium from stationary cell culture ......................25
3.2.1. Comparison of growth length of single filaments before fragmenting ..... 26
3.2.2. Compare fragmentation time of single filament........................................ 26
3.2.3. Comparison of growth rate of single filaments ......................................... 27
3.3. Compare growth rate of the same strain in the modified standard Zarrouk’s
media and Zarrouk’s medium from stationary cell culture.........................................28
3.4. Discussion: Life cycle of Arthrospira platensis. ...................................................29
PART 4. CONCLUSIONS AND SUGGESTIONS ........................................ 31
4.1. Conclusions .....................................................................................................................31
4.1.1. Cells/filaments were cultured in the standard Zarrouk’s medium ............ 31
4.1.2. Cells/ filaments in the Zarrouk’s medium from stationary cell culture .... 31
4.2. Suggestions ......................................................................................................................31

REFERENCE...................................................................................................... 33

iv


LIST OF FIGURES

Figure 1.1. Various applications of microalgae products for human, animals
and industries ......................................................................................................... 3
Figure 1.2. Helical trichomes and straight of Arthrospira platensis. The scale
bar (a)=40 µm, (b)=20 µm, respectively (Source: C.Sili, 2012). ........................... 5
Figure 1.3. The microfluidic devices: (a) including 3 layers: positively charged
glass slide, microwell layer, fluidic layer; (b) devices completed. ........................ 7
Figure 1.4. Overview of advantages of both macroscopic and microfluidic cell
culture (Halldorsson et al, 2015) .......................................................................... 10
Figure 2.1. Medium culture: (a) standard Zarrouk’s medium, (b) Zarrouk’s
medium from stationary cell culture..................................................................... 14
Figure 2.2. Morphology of Arthrospira platensis in the microwell. The scale
bar represents 100 µm, respectively. .................................................................... 15
Figure 2.3. The fabricated device. The device is composed 3 layers: microwell
layer (b), the positive charged glass slide (c) and fluidic layer (e); PDMS was
poured in the mold (a), glass slide and microwell were bonded by plasma
machine (d); then microfluidic were created by bonding between (d) and e.
Microfluidic devices were displayed in (f). .......................................................... 16
Figure 2.4. C005 str and Central Lab str were transferred new medium and
kept in the incubator from 1 to 5 days. ................................................................ 17
Figure 2.5. The process of making microfluidic devices .................................... 18
Figure 2.6. The process of cell loading and cultivation in the microwell-based
microfluidic devices. ............................................................................................ 18
Figure 2.7. Process set experiment: (a) sample was kept in a petri dish; (b)

A.platensis cell was checked; (c) keep the sample and microscopy inside the
incubator (connect with computer) ....................................................................... 19
Figure 2.8. Measure the length of filaments of C005 strain (a) and Central
Lab strain (b) ........................................................................................................ 19
Figure 3.1. Growth length of single filaments of C005 str and Central Lab str
in modified Zarrouk’s medium. ............................................................................ 22
v


Figure 3.2. Fragmentation time of single filaments of C005 str and Central
Lab str in modified Zarrouk’s medium ................................................................ 22
Figure 3.3. Compare the growth rate of single filament of C005 and Central
Lab str in modified Zarrouk’s media. ................................................................... 23
Figure 3.4. The phenomenon color- changed filaments of C005 str (a) and
Central Lab str (b)................................................................................................. 25
Figure 3.5. Compare growth length of single filaments when they were
cultured in medium from stationary cell culture. ................................................. 26
Figure 3.6. Comparison of fragmentation time of single filaments when they
were cultured in medium from stationary cell culture ......................................... 27
Figure 3.7. Compare the growth rate of single filaments when they were
cultured in medium from stationary cell culture. ................................................. 27
Figure 3.8. Life cycle of Arthrospira by Ciferri and Tiboni, 1983 .................... 29
Figure 3.9. Life cycles of Arthrospira platensis from our experiment
(C005 str and Central Lab str). ............................................................................. 30

vi


LIST OF TABLES


Table 2.1. Equipments for studies ........................................................................ 13
Table 2.2. Constituents of Zarrouk’s medium ...................................................... 14
Table 3.1. Basic information of each Arthrospira platensis single filaments
in modified Zarrouk’s medium. ............................................................................ 24
Table 3.2. Basic information of each Arthrospira platensis single filaments
in the modified Zarrouk’s medium from stationary cell culture. ......................... 28
Table 3.3. Comparison of growth rate based on average and SD of each strain......... 29

vii


LIST OF ABBREVIATIONS
%

Percentage

µl

Microliter

µm

Micrometer

A.platensis

Arthrospira platensis

ACOI


Coimbra Collection of Algae

CL str

Central Lab strain

FAO

Food and Agriculture Organization

G

Gram

Min

Minutes

ºC

Degree centigrade or Celcius

Off

Offspring

PDMS

Polydimethysiloxane


Rpm

Revolutions per minute

SD

Standard Deviation

Str

Strain

DSLR

Digital single-lens reflexs

viii


PART 1. INTRODUCTION
At the present, we are standing the challenge of energy, food crisis due to
the population explosion of the world. One of factors to solve this difficult is
phytoplantonic. That is algae-Arthrospira platensis (A.platensis). A.platensis is a
filamentous cyanobacterium that has been studied and produced by large factories
in large-sized countries such as China, India and the United States (Pulz and Gross,
2004). A. platensis is an ideal food and dietary supplement for the 21st century by
Food and Agriculture Organization (FAO) of the United Nation (Pelizer, 2003).
The successful commercial exploitation of A.platensis because of its high
nutritional value, chemical composition such as protein, pigments, antioxidants,
fatty acids (γ-linoleic acid) (Pulz and Gross, 2004), pharmaceutical compounds

(Kuntanawat, 2014). They are safety of the biomass has made it one of the most
important industrially cultivated microalgae. Knowledge of its biology, chemistry
and physiology, which is essential for understanding the growth kinetic,
morphology, have been used in the different conditioned medium.
Microfluidics, the study of fluid flow at microscale and its application in
biological, biomedical, biotechnology and chemical analyses, has been large
progress over the last two decades (Squires and Quake, 2005). Microfluidics
systems have many advantages over traditional technicals such as low cost, low
area, low reagent consumption, fast response time, flexibility of device design,
experimental flexibility and control, single cell handling, real-time on a chip
analysis. Some microfluidic systems created new functions based on the combine
physical, chemical and biological characteristics at microscale which are not
available for macro-systems (flask, petri dish, etc). One important class of
microfluidic systems are those for cell culture and the ability to control
parameters of the cell microenvironment at growth length, fragmentation time,
growth rate, cellular behaviors, growth kinetics in the specified physiological
microenvironment.
Studies of single-cell microalgae are grown interest, because these organisms
are being used as model systems for the studies of many fundamental biological
1


processes (Hoek et al, 1995), as well as in many commercial, industrial and
biological applications. Gaining a understanding of single-cell in Arthrospira
platensis that it will also be a great value to optimize the biotechnological
applications. As the reasons above, it inspires for us to study the properties of
Arthrospira platensis that nobody had ever known before. Fundamental
understanding of the cellular phenomena requires detailed investigations of the
growth, observe the activities of a single cell. The kinetic parameters have measured
the length, fragmentation time of each filament. Therefore, I was chosen the

topic:―Algal cell culture in microfluidic devices and microenvironment‖.
1.1 Background
1.1.1 Microalgae
Microalgae are sunlight-driven cell factories that convert dioxide to
potential biofuels, foods, feeds and high-value bioactives, agricultural, chemical
and pharmaceutical sectors (Yusuf Chisti, 2007) (Fig 1.1). Microalgae reproduce
themselves using photosynthesis to convert sun energy into chemical energy,
completing entire growth cycles every few days (Sheehan J et al, 1998).
Microalgae are prokaryotic or eukaryotic photosynthetic microorganisms that can
grow rapidly and live in harsh conditions due to their unicellular or simple
multicellular

structure

(Mata

et

al,

2010).

Examples

of

prokaryotic

microorganisms are Cyanobacteria (Cyanophyceae) and eukaryotic microalgae
are for example green algae (Chlorophyta) and diatoms (Bacillariophyta) (Mata

et al, 2010). Moreover, they can grow almost anywhere, requiring sunlight and
some simple nutrients, although the growth rates can be accelerated by the
addition of specific nutrients and sufficient aeration (Muhling et al, 2005).
Different microalgae species can be adapted to live in a variety of environmental
conditions. Microalgae are presented in all existing earth ecosystems, not just
aquatic but also terrestrial, representing a big variety of species living in a wide
range of environmental conditions. It is estimated that more than 50.000 species
exist, but only a limited number, of around 30.000, has been studied and
analyzed (Mata et al, 2010).
2


Figure 1.1. Various applications of microalgae products for human, animals
and industries
( />As studied before, some active organic compounds can be extracted from
algae. The antioxidant agents such as carotenoid and phycobiliprotein can be
extracted from green algae (Chlorella sp.) and blue-green algae (Spirulina sp.),
respectively. In addition, the Chlorella spp., Scenedesmus spp., Chlamydomonas
spp., Euglena viridis, Fragilari ambigua and Microcystis aeruginosa have been
reported as the main groups of microalgae to produce antimicrobial substances.
Those Antimicrobial substances are identified including of fatty acids,
glycolipids, acrylic acid phenolics, cyclic peptides, N-glycosides, sulphatepolysaccharides, ß-diketone, isonitriles-containing indole, alkaloids.
During the past decades, extensive collections of microalgae have been
created by researchers in different countries. An instance is the freshwater
microalgae collection of the University of Coimbra (ACOI) in Portugal
considered one of the world’s largest, having more than 4000 strains and 1000
species. This collection attests to the large variety of different microalgae
available to be selected for use in a broad diversity of applications, such as value
3



added products for pharmaceutical purposes, food crops for human consumption
and an energy source (Singh el al, 2010).
1.1.2 General Arthrospira platensis
Arthrospira (Spirulina) is marketed widely under the name ―Spirulina” as
a food supplement for humans and animals. They are filamentous, nonheterocystous cyanobacteria that are generally found in tropical and subtropical
regions in warm bodies. They are one the most cultivated commercial
microalgae. Cyanobacteria constitute one of the largest groups of prokaryotes.
They are some of the simplest life forms on earth and the cellular structure is
simple prokaryote, which can perforgle filament of each
strain (Fig 3.1). We realize that filaments/cells are different growth length.
Furthermore, the Central Lab str is longer than C005 str. It was demonstrated by
average, SD, length of mother filaments and offspring filaments of each strain.
With total 10 mother filaments; in C005 str: date 31/1/2017 has the longest
length of 625.451 µm and date 11/1/2017 has the shortest length of 386.796 µm;
in Central Lab str: date 15/2/2017 has the longest length of 855.857 µm and date
3/2/2017 has the shortest length of 557.222 µm. Thus, compare growth length of
21


offspring; in C005 str: the length of offspring is 823.890 µm the longest (date
9/2-off 2) and 267.019 µm the shortest (date 9/2-off 1); the Central Lab str: the
length of offspring is 1.272.496 µm (date 15/2-off 1) the longest and 601.855 µm
(date 3/2-off 2) the shortest. It has contributed to prove that the length of the
Central Lab str is longer than C005 str.

Figure 3.1. Growth length of single filaments of C005 str and Central Lab str
in modified Zarrouk’s medium.
3.1.2. Compare fragmentation time of single filaments


Figure 3.2. Fragmentation time of single filaments of C005 str and Central
Lab str in modified Zarrouk’s medium
Fragmentation time of the Central Lab str is the same as C005 str.
Because, if we based on SD to compare, we realize that SD of C005 str is higher
than SD of Central Lab str. But average of C005 str is shorter than Central Lab
22


str. In C005 str, maximum time is 2916 minutes (date 11/1-off 2) and minimum
time is 225 minutes (date 9/2-off 1). In Central Lab str, maximum time is 2874
minutes (date 27/1-off 2) and minimum time is 555 min (date 19/1-off 1).
Fragmentation time of offspring 1 always is shorter than offspring 2 of Central
Lab str.
3.1.3. Comparison of growth rate of single filaments
The result of the experiment indicated that the single filaments of each
strain different growth rate even they were from the same batch (Fig 3.3). This
phenomena reflect the inhomogeneity among the Arthrospira platensis
population. Therefore, they cannot be studied from the descriptive data recording
and the conventional bulk scale measurement of growth based on such as
spectrophotometric methods or cell counting. Moreover, we found that the
growth rate of single filaments of every three minutes was not constant and
showed no specific manner. This result might reflect the different cellular
phenomena that occurred in multicellular filament at different time interval.
In C005 str, growth rate of filament is 210.950 µm/min (date 9/2-off 2) the
fastest and 42.264 µm/min (date 9/2-off 1) the slowest. In Central Lab str, the
growth rate of filament is 389.440 µm/min (date 15/2-off 1) the fastest and
144.090 µm/min (date 11/2-off 2) the slowest.

Figure 3.3. Compare the growth rate of single filament of C005 and Central
Lab str in modified Zarrouk’s media.

23


Therefore, we realize that the growth rate of Central Lab str is faster than
C005 str. Thus, the average of the Central Lab str is higher than C005 str and the
SD of Central Lab is same C005 str. To explain why two strains had the different
growth rate. We based on the structure of each strain. As Fig 2.2, C005 str is
helical morphology, which effect the fragmentation, movement of single
filaments. Central Lab str is straight which easy for fragmenting, moving,
absorbing nutrition.
Additionally, other basic information, including of the length before
fragmenting, length of offspring 1 and 2 from start to finish, fragmentation time
shows in table 3.1.
Table 3.1. Basic information of each Arthrospira platensis single
filaments in modified Zarrouk’s medium.

24


3.2. Filaments in the Zarrouk’s medium from stationary cell culture
We conducted to with total 10 single filaments, including 5 filaments
C005 str and 5 filaments Central Lab str. In C005 str, it has 2 filaments that
fragment (rate 40%) and 3 filaments change the color (Fig 3.4) when we collect
picture from our experiment. In Central Lab str, it has 3 filaments that fragment
(rate 60%) and 2 filaments change the color (Fig 3.4). So, based this rate, we can
easily see, the strain Central Lab is still able to grow faster. To prove in detail,
we continually compare the growth length, fragmentation time, the growth rate of
single filament of each strain. Additionally, to find the cause that the
phenomenon of color change of these two strains. We think that the main reason
which the environment lacked nutrients. Because, the medium was filted when

cell cultured in the station phase.

Figure 3.4. The phenomenon color- changed filaments of C005 str (a) and
Central Lab str (b).

25


3.2.1. Comparison of growth length of single filaments before fragmenting
The growth length of single filaments of each strain which they are
different. We realize that the length of C005 str is longer than Central Lab str. It
is demonstrated based on graph, average and SD of each strain (Fig 3.5). The
both mother filaments of C005 str are longer than mother filaments of the Central
Lab star. The mother filament is 920.160 µm (date 3/3-C005) the longest and
476.117 µm the shortest (date 19/3-Central Lab). Following, we compared the
growth length of offspring of each strain. In C005 str, the length is 527.470 µm
(date 13/3-off 2) the longest and 360.160 µm (date 13/3-off 1) the shortest. In
Central Lab str, the length is 546.214 µm (date 16/3-off 1) the longest and
385.715 µm (date 19/3-off 1) the shortest. So, we found that the growth length of
filaments of the Central Lab str is equivalent.

Figure 3.5. Compare growth length of single filaments when they were
cultured in medium from stationary cell culture.
3.2.2. Compare fragmentation time of single filament
Fragmentation time of C005 str is shorter than Central Lab str. It was
demonstrated by average, SD, time that showed Fig 3.5.
We can easily realize that fragmentation time of the Central Lab str is five
times longer than C005 str. In C005 str, maximum time is 390 minutes (date 3/3off 1&2) and minimum time is 120 minute (date 6/3-off 1). In Central Lab str,
maximum time is 2070 minute (date 19/3-off 2) and minimum time is 150
minutes (date 16/3-off 1). Furthermore, growth length and fermentation time of

the two strains are inverse proportion together.
26


Figure 3.6. Comparison of fragmentation time of single filaments when they
were cultured in medium from stationary cell culture
3.2.3. Comparison of growth rate of single filaments
We observed the development of two filaments of C005 str and three
filaments of the Central Lab str. As mentioned the rate above, we realize that the
growth rate of Central Lab is faster than C005 str, to make sure for this
hypothesis. We compared the growth rate of offspring of the Central Lab str and
C005 str that showed Fig 3.6.

Figure 3.7. Compare the growth rate of single filaments when they were
cultured in medium from stationary cell culture.
27


In C005 str, growth rate of filaments is 7.940 µm/min (date 3/3-off 2) the
fastest and 3.441 µm/min (date 3/3-off 1) the slowest. In Central Lab str, the
growth rate of filaments is 150.080 µm/min (date 31/3-off 1) the fastest and
18.612 µm/min (date 16/3-off 2) the slowest. Additionally, we can based on
average and SD of each strain. The growth rate of Central Lab str is nearly 20
times higher than C005 str. The basic information of each strain in Zarrouk’s
medium from stationary cell culture and showed table 3.2.
Table 3.2. Basic information of each Arthrospira platensis single filaments in
the modified Zarrouk’s medium from stationary cell culture.

3.3. Compare growth rate of the same strain in the modified standard
Zarrouk’s media and Zarrouk’s medium from stationary cell culture

Through table 3.3, we realize that the growth rate of two strains in the
standard Zarrouk’s medium had significantly faster than two strains in the
Zarrouk’s medium from stationary cell culture. So, to explain why this growth
rate is different, we give a reason: When the cell was cultured in stationary phase,
the cellular was able to the growth very bad. The medium had been filtered from
stationary Arthrospira platensis cell culture and this medium was used to kept
filaments in the microwell. Additionally, the nutrient was contented to reduce in
this medium. Therefore, the cell growth is normal very low. Therefore, the cell in
28


this environment could dead, changed colors, slow movement, fragmentation
time from mother filaments until offspring filaments were very long.
Table 3.3. Comparison of growth rate based on average and SD of each strain.
Standard
Strains

Average
(µm/min)

Stationary
SD

Average
(µm/min)

SD

C005


148.116

52.83

5.990

1.65

Central Lab

194.670

52.59

81.24

44.37

3.4. Discussion: Life cycle of Arthrospira platensis.
From the experiment, we found that the growth manner of our Arthrospira
platensis was not in the same old manner that was described by Ciferri and
Tiboni, 1983 (Fig 3.7).

Figure 3.8. Life cycle of Arthrospira by Ciferri and Tiboni, 1983
Our Arthrospira platensis didn’t separate into small fragment and grown to
the hormogonia, necridia. A.platensis fragmented into two long filaments and
further elongated (Fig 3.8). They will continually elongate, fragment and didn’t
stop. They will only stop when the nutrients decrease in the environment.
29



Figure 3.9. Life cycles of Arthrospira platensis from our experiment (C005 str
and Central Lab str).
3.8.a: Mother filaments and continually elongate.
3.8.b: Mother filaments fragment into two offspring (off 1&2)
3.8.c: One offspring fragment and create three filaments
3.8.d: Four filaments

30


PART 4
CONCLUSIONS AND SUGGESTIONS

4.1. Conclusions
4.1.1. Cells/filaments were cultured in the standard Zarrouk’s medium
The average length of C005 str is about 540 µm; Central Lab str is about
815 µm. Therefore, The length of Central Lab str is longer than C005 str. The
fragmentation time of C005 str is the same Central Lab str, around 1500 minutes.
Two strains is different growth rate even they were from the same batch. So, this
phenomena reflect the inhomogeneity among the Arthrospira platensis
population. The average growth rate of C005 str is about 148 µm/min and
Central Lab str is 194 µm/min. Thus, Central Lab str grew faster than C005 str.
4.1.2. Cells/ filaments in the Zarrouk’s medium from stationary cell culture
The length of the two strains is shorter than filaments was kept in the
standard Zarrouk’s medium. Furthermore, the average length of Central Lab str is
about 508 µm and C005 str is 583 µm. So, the length of C005 str is longer than
Central Lab str. The average for fragmentation time of C005 str is 285 minutes
and Central Lab str is 1435 minutes. Regarding, fragmentation time of Central
Lab is the five times higher than C005 str. Beside that, the average growth rate of

C005 is 6 µm/min and Central Lab str is 80 µm/min. The growth rate of Central
Lab str is approximately twenty times faster than C005 str.
The growth rate of Arthrospira platensis is not constant. Two strains were
cultured in standard Zarrouk’s medium, it had grown faster than two strains in
Zarrouk’s medium from stationary cell culture. Thus, Central Lab str has always
grown faster than C005 str. The maximum length at fragmentation time of
Central Lab str is longer than C005 str.

4.2. Suggestions
- Continually experiment about 10 filaments of each strain for clearly and
convince conclusion

31


- Continually study, observe activation the growth rate of two strains (
C005 str and Central Lab str) in the Zarrouk’s medium with high salinity
(NaCl=0.5M and NaCl=0.75M) using microwell based microfluidic devices.
Comparison the growth rate of the same strain or different strain with the same
medium or different medium. (growth length, fragmentation time, growth rate) in
the three different mediums.
- The ability to survey the growth rate of two strains when the algal cells
were cultured in the glass slide and observe activation under the

confocal

microscopy.

32