Volume 2, No.

9 September 2023 - (1036-1045)

p-ISSN 2980-4868 | e-ISSN 2980-4841
https://ajesh.ph/index.php/gp

EFFECT NUTRIENTS FOR GROWTH CHLORELLA VULGARIS AND

SPIRULINA PLATENSIS
1*R Mohammad Alghaf Dienullah, 2Harmin Sulistiyaning Titah
Faculty of Civil, Planning and Geo Engineering, Institut Tekonologi Sepuluh November
Email: [email protected]

ABSTRACT:
Microalgae are versatile organisms with the ability to adapt to various environments and exhibit
rapid growth. They play a significant role in photosynthesis by converting carbon dioxide into
oxygen and utilizing nutrients in their surroundings. This study aims to investigate the impact of
nutrient availability on the growth of Chlorella vulgaris and Spirulina platensis. The experiment
involves culturing the microalgae in mineral water and distilled water mediums supplemented
with Walne medium nutrients. The growth of microalgae is monitored daily by calculating cell
density using a counting chamber method. The pH and temperature of the culture are also
measured. The results show that both microalgae species can grow in both mediums, but the
usagee of mineral water medium promotes higher cell density and growth rates compared to
distilled water medium. The pH values of the cultures vary between mediums, with mineral water
medium generally exhibiting higher of pH. The temperature remains relatively stable throughout
the experiment. These findings indicate that nutrient availability has a significant influence on
microalgae growth, and the use of mineral water medium enhances their growth potential.

Keywords: Nutrients, Growth Rate, Microalgae, Chlorella vulgaris, Spirulina platensis

INTRODUCTION microalgae is one of most significantly

In world nature, microalgae is one contributed photosynthetic activity. It can
living things that have the potential to be convert carbon dioxide to oxygen by
studied due to their adaptability to metabolism process itself for cell growth
anywhere environment (cosmopolite) and (Thakur et al., 2020). While producing
has fast growing ability. It can adapt until oxygen, microalgae using carbon dioxide by
extreme environment such as pH, salt levels, absorption process and assimilating some
and extreme climate. Besides that,

Journal of Engineering, Social and Health - Vol 2 (9) September 2023 1036
R Mohammad Alghaf Dienullah, Harmin Sulistiyaning Titah

nutrients in their life medium (Gani et al., carbon, phosphorus, silicon, vitamins, and
2016). some metals such as zinc, copper, iron can
Some researchers using many species affect for growth microalgae. Among them,
of microalgae in nature which commonly carbon is most important which has more
used for experiment, such as Chlorella effect on microalgae metabolism. It being
vulgaris, and Spirulina platensis. Chlorella energy source and the main structural
vulgaris is categorized as green microalgae component of microalgae cell. If microalgae
from genus Chlorella, has 2-10 micrometer categorized as autotropic culture,
spherical microscopic cell and it growth to bicarbonate component or carbon dioxide is
maturity range is range of 17-21 nanometer. used as main carbon source. But if
It is also identified as eukaryotic and microalgae categorized as heterotrophic
photosynthetic microorganism frrom family culture, glucose, sucrose, acetate, glycerol
chlorellaceae (Daliry et al., 2017). Chlorella and other similar things are organic
vulgaris reproduces asexually (Okoro et al., materials that used for source carbon (Daliry
2019). When reproduction, a mother cell of et al., 2017).
Chlorella vulgaris can reproduce 4 cell in This study was focused to know the
range times about 19 hours (Daliry et al., impact of the availability of nutrients for
2017). While Spirulina platensis or also called growth rate culture microalgae. By using
Arthospira platensis is categorized as Chlorella vulgaris and Spirulina platensis for
Oscillatoria, Cyanophycean and well known object experiment, the results are expected
as blue-green microalgae (Cyanophyta). It to describe effect composition nutrient for
mostly found in alkaline salt lake. It is also Chlorella vulgaris and Spirulina platensis
identified as ancient lower prokaryotic plant growth.
that composed of 5-10 micrometer wide,
200-500 micrometer long, a multicellular or RESEARCH METHODS
single filamentous body like loose or tight Preparation Microalgae Culture
regular spiral bending (Ai et al., 2023). The microalgae species used for the
The best condition for grown experimental is the starter culture of
microalgae is mostly depends on some Chlorella vulgaris and Spirulina platensis. The
factors, such as environment condition (pH, medium used is distilled water and mineral
salinity, temperature, light intensity and its water. The nutrient in mineral water was
period), culturing technique, and availability contained: Calcium 9,14 mg/L; Magnesium
of nutrients in their medium (Gani et al., 5,87 mg/L; Natrium 35 mg/L; Potassium 3
2016). Environment condition is categorized mg/L; Bicarbonate 118 mg/L; Sulfate 3,06
as physical factor, and availability of mg/L; Chloride <0,01 mg/L; Total Dissolved
nutrients is categorized as chemical factor. Solid (TDS) 177 mg/L; and pH 7,7. Both
Availability of nutrients is about amount and reactors containing distilled water and
composition in culture medium. Nitrogen, mineral water will be added with additional
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 Effect Nutrients for Growth Chlorella Vulgaris and Spirulina Platensis

nutrients in the form of Walne medium at a were capable of developing in medium as

dose of 1 mL/L. It addition to culture only long as they had a certain content that can
after inoculation. Walne medium are promoted metabolic development (Sánchez-
contained: NaNO3 100 g/L; H2BO3 33,6 g/L; Bayo et al., 2020). The growth of Chlorella
Na2EDTA 45 g/L; NaH2PO4H2O 20 g/L; vulgaris are shown in Figure 1 and the
FeCl3.6H2O 1,3 g/L; MnCl2.4H2O 0,36 g/L; growth of Spirulina platensis are shown in
Vitamins solution 1 mL; and trace element Figure 2.
0,1 mL.
Experimental set-up
The experiment was carried out in a
high growth rate microalgae reactor scale
batch consisting of several components such
as a 1.2 L tube, an aerator with an aeration
rate of 2 liters O2/minute, and a 6000 lux
cooldaylight LED lamp. The total volume of
the reactor is 1 liter with 10% of which
Figure 1. Cell Density Chlorella vulgaris at
contains microalgae inoculum. aeration will
The Medium Mineral Water and Distillated
be turned on for 24 hours, while the lighting
Water
time period is planned 12/12 (12 hours light,
Cell density of Chlorella vulgaris
12 hours dark), and the experiment will run
showed increase from the first day to the
for 7 days.
fifth day. After that it began decrease to the
Determination of Microalgae Growth
seventh day. Both medium variations show
Every day, the density of microalgae
the same trend of cell density changes. Until
cells in each reactor will be calculated using
day fifth observation, increased cell density
the counting chamber method with the
continuously caused by Chlorella vulgaris
Haemocytometer Improved Neubauer on a
utilized the nutrients to reproduce and
microscope with a magnification of 100X
diffusion that can be affected on growth
(10X ocular lens and 10X objective lens).
rates. It was also suspected that Walne had
Besides that, every day the parameters of pH
given more nutrients for cell proliferation
and temperature in all reactors are also
(Hismayasari et al., 2021). Based on the
observed.
growth curve on Figure 1, the largest cell
density was shown on day 5, with a cell
RESULTS AND DISCUSSION
density of 6.08 x 108 cells/mL by using
Effect Nutrients for Cell Growth mineral water medium. (Wardani et al.,
Chlorella vulgaris and Spirulina 2022) refers that the increase in cell density
platensis studied able to grow in medium day also showed that microalgae can utilize
mineral water and distillated water during 7 the nutrients in the medium for cell growth
days on batch reactor. The two microalgae
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R Mohammad Alghaf Dienullah, Harmin Sulistiyaning Titah

and division. However, until the day there However, giving excess nutrients will result
was a decrease in the number of cell in stress conditions on cultivated microalgae
densities indicating that the cells had and lead to a decrease in biomass (Pratiwi et
reached their saturation point and the rate al., 2019).
of cell death would increase which was not Based on experiments, both species of
matched by the rate of cell growth. microalgae showed that the use of mineral
water containing nutrients as a medium is
better than using distilled water. Besides
from the growth curve on Figure 1 and Figure
2, the results of calculating the growth rate
of microalgae also showed the same thing.
Refers on (Andersen, 2005; Adriyanti et al.,
2021), the growth rate is the rate of increase
or increase in the number of microalgae cells
in the culture at a certain time. Chlorella
Figure 2. Cell Density Spirulina platensis at
vulgaris which cultured in mineral water
The Medium Mineral Water and Distillated
medium showed growth rate results 0,14
Water
cell/mL/day. Whereas if it using the medium
Cell density of Spirulina platensis
of distilled water, the growth rate decreased
showed increase from the first day to the
to 0.07 cell/mL/day. Spirulina platensis
third day. After that, it began decrease to the
which cultured in mineral water medium
seventh day. Both medium variations
showed growth rate results -0,13
showed the same trend of cell density
cell/mL/day. Whereas if it using the medium
changes. The increased density cell showed
of distilled water, the growth rate decreased
that under these conditions the microalgae
to -0.32 cell/mL/day. These results bear out
cells have utilized the nutrients present in
that the availability of nutrients has an
the media (Caturwati & Setyati, 2020; Hasim
influence on the growth rate of microalgae
et al., 2022). Based on the growth curve on
cells.
Figure 2, the largest cell density was shown
Effect Nutrients for pH
on day 5, with a cell density of 8 x 106
Abiotic factor can affect the
cells/mL by using mineral water medium.
microalgae growth. During the period of
But the cell density of Spirulina platensis
cultivation of microalgae, pH value indicated
when it is at its peak growth is not as much
the photosynthetic assimilations of the
as Chlorella vulgaris when it is at its peak
carbon dioxide. It inferred another factor
growth. The growth of microalgae is indeed
responsible microalgae for availability of
influenced by the availability of nutrients in
carbon (Azov, 1982; Thakur et al., 2020). pH
the growing media which are divided into
also has great significance in microalgal
macronutrients and micronutrients.
cultures, it determines the solubility of
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 Effect Nutrients for Growth Chlorella Vulgaris and Spirulina Platensis

minerals and CO2 in the medium (Chowdury

et al., 2020).

Figure 4. pH Spirulina platensis at The

Medium Mineral Water and Distillated
Figure 3. pH Chlorella vulgaris at The Water
Medium Mineral Water and Distillated Based on Figure 4, The pH of Spirulina
Water platensis in distilled water and mineral water
Based on Figure 3, The pH of Chlorella showed different results. Where the pH of
vulgaris in distilled water and mineral water microalgae in mineral water medium is
showed different results. Where the pH of higher than in distilled water. The use of
microalgae in mineral water medium is distilled water will reduce the pH of the
higher than in distilled water. The use of medium from 7,9 to 7,1 on the seventh day.
distilled water will reduce the pH of the The higher pH was 7,9 only until first day.
medium from 8 to 7,4 on the second day. But After that, the pH values is decreased until
until the seventh day, the pH gradually day of 7. Whereas if used mineral water
increased to 7,7. Whereas if used mineral medium, the pH of microalgae was in the
water medium, the pH of microalgae is in the range of 7.6 to 8.4. The microalgae pH was
range of 7.8 to 8.2. The microalgae pH was initially 7,9 after inoculation (at zero day). It
initially 8 after inoculation (at zero day), but increased until 8,4 on fourth day until day of
increased to 8.2 on the second day, before 5. After that, it decreased on until 7.6 at the
gradually decreasing to 7.8 until the seventh day of 7. In the last day, microalgae using
day. In the last day, microalgae using mineral mineral water medium had a higher pH value
water medium had a higher pH value compared to using distilled water.
compared to using distilled water. Chlorella vulgaris reported can grow
pH range 4 until 10, but most biomass
productivity can be achieved if it lives in an
environment with a pH 9 until 10 (Khalil et al,
2010; Daliry et al., 2017). Meanwhile,
spirulina platensis can survive up to pH 11
and will grow well if it is at a pH in the range
of 7,2 to 9,5 (Isnansetyo & Kurniastuty, 1995;

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R Mohammad Alghaf Dienullah, Harmin Sulistiyaning Titah

Ningsih et al., 2019). The changes in pH will

cause produce changes in metabolism in
microalgae cultures because they affect the
balance of inorganic carbon. In a liquid
medium, most of the carbon is in the form of
bicarbonate in the pH range of 7 to 9.
Microalgae cells can process bicarbonate
into a carbon source due to the presence of
the rubisco enzyme (Okoro et al., 2019;
Figure 5. Temperature Chlorella vulgaris at
Meier et al., 2022). Then its converted to the
The Medium Mineral Water and Distillated
enzyme carbonic anhydrase (Zhu et al.,
Water
2020). The decomposition of dissolved
Based on Figure 5, temperature of
carbon causes a decrease in pH. However,
Chlorella vulgaris in distilled water and
the activity of the carbonic anhydrase
mineral water showed similar results. The
enzyme will cause an increase in pH outside
use of mineral water medium produces a
the cell due to the hydroxide ion transport
slightly higher final temperature than the
which is closely related to the capture of H+
use of distilled water medium. In day zero
ions to the inside of the thylakoid membrane
(after inoculation), the temperature in both
(Kumar & Das, 2012). Referring to the
mineral water and medium distilled water
experimental results, an increase in pH
was 27.9oC, then increased until the second
indicates that the microalgae cells carry out
day to a temperature of 28.9oC for mineral
metabolic activities, while a decrease in pH
water medium and 28.8oC for distilled water
may indicate a decrease in the metabolic
medium. After the second day, the
activity of microalgae cells
temperature in both mediums decreased
Effect Nutrients for Temperature
slightly which lasted until the seventh day of
Temperature is indicator that can
the experiment.
significantly limit and affect growth of
microalgae. It has varies value the optimum
temperature for growh in the range 15oC
until 30oC, but it depends on the microalgae
species (Kaloudas et al., 2021).

Figure 6. Temperature Spirulina platensis at

The Medium Mineral Water and Distillated
Water
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 Effect Nutrients for Growth Chlorella Vulgaris and Spirulina Platensis

Based on Figure 6, temperature of temperatures to growth about 30oC. But when

Spirulina platensis in distilled water and it has an excessive temperatures up to 38oC,
mineral water showed similar results. The it will make Chlorella vulgaris cell die and an
use of mineral water medium produces a abrupt halt in growth (Daliry et al., 2017).
slightly higher final temperature than the Referring to the experimental results, both
use of distilled water medium. In day zero types of microalgae showed that in seven
days there are not microalgae in various
(after inoculation), the temperature in both
media reached the optimum temperature for
mineral water and medium distilled water
growth. It is known that the highest
was 27.9oC, then increased until the second
temperature reached by Chlorella vulgaris
day to a temperature of mineral water
and Spirulina platensis is almost at the
medium increased to 28,9oC and optimum temperature and does not reach the
temperature of distillated water medium temperature that inhibits growth. In addition,
increased to 28,7oC. After the second day, experimental results prove that the highest
the temperature in both mediums decreased temperature for both types of microalgae is
until the seventh day of the experiment obtained when using mineral water medium,
which mineral water medium reach so that nutrient availability can affect the
temperature 28oC and distillated water temperature in the high growth rates reactor.
medium reach temperature 27,8 oC.
An increased temperature will have a CONCLUSION
positive effect on photosynthesis and cell The effect of availability nutrients on
division if it below on optimal growth the growth of Chlorella vulgaris and Spirulina
temperatures. This trend is explained by the platensis has evaluated. Nutrient availability
enhancement of enzymatic activities that
was known to affect the growth rate of
related to the Calvin cycle. If growth
microalgae. Chlorella vulgaris and Spirulina
temperature exceeding the optimal
platensis can grow well in a medium
temperature, it can make growth rate of
containing more nutrients. This was
microalgae sharply decrease because the heat
stress can make enzymes inactivated reinforced by several other parameters
,denaturation, and modify proteins which are besides growth rate, namely pH and
involved in photosynthetic processes (Ras et temperature parameters. The pH and
al., 2013). Spirulina platensis has the temperature that were in the range close to
optimum temperature for growth, the optimum indicate that microalgae carry
biochemical, and antioxidants at 30oC. At out more metabolism, thus accelerating the
these temperatures, some antioxidant growth of microalgae cells.
enzymes activity increased. Antioxidant
affect to prevent the damage of cellular
membranes by lipid preoxidation (Ismaiel &
Piercey-Normore, 2020). Meanwhile,
Chlorella vulgaris has the optimum
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R Mohammad Alghaf Dienullah, Harmin Sulistiyaning Titah

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FROM MgSO 4 . 7H 2 O AND IRON

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