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Effects of salinity and nutrients on the growth and chlorophyll fluorescence of

Caulerpa lentillifera

Article  in  Chinese Journal of Oceanology and Limnology · March 2014

DOI: 10.1007/s00343-015-4105-y

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 Chinese Journal of Oceanology and Limnology
Vol. 33 No. 2, P. 410-418, 2015
http://dx.doi.org/10.1007/s00343-015-4105-y

Effects of salinity and nutrients on the growth and chlorophyll

fluorescence of Caulerpa lentillifera*

GUO Hui (郭辉)1, 2, YAO Jianting (姚建亭)1, SUN Zhongmin (孙忠民)1,

DUAN Delin (段德麟)1, **
1
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China
2
University of Chinese Academy of Sciences, Beijing 100049, China

Received Apr. 29, 2014; accepted in principle Jul. 22, 2014; accepted for publication Aug. 19, 2014
© Chinese Society for Oceanology and Limnology, Science Press, and Springer-Verlag Berlin Heidelberg 2015

Abstract Caulerpa lentillifera is a green algae that distributes worldwide and is cultivated for food.
We assessed vegetative propagation of C. lentillifera by measuring the specific growth rate (SGR) and
chlorophyll fluorescence of the green algae cultured at different salinities and nutrient levels. The results
indicated that C. lentillifera can survive in salinities ranging from 20 to 50, and can develop at salinities
of 30 to 40. The maximum SGR for C. lentillifera occurred at a salinity of 35. Both chlorophyll content
and the ratio of variable to maximum fluorescence (Fv/Fm) were also at a maximum at a salinity of 35.
Photosynthesis was inhibited in salinities greater than 45 and less than 25. Both the maximum SGR and
maximum chlorophyll content were found in algae treated with a concentration of 0.5 mmol/L of NO3-N
and 0.1 mmol/L of PO4-P. The photosynthetic capacity of photosystem II (PSII) was inhibited in cultures
of C. lentillifera at high nutrient levels. This occurred when NO3-N concentrations were greater than
1.0 mmol/L and when PO4-P concentrations were at 0.4 mmol/L. As there is strong need for large-scale
cultivation of C. lentillifera, these data contribute important information to ensure optimal results.

Keyword: Caulerpa lentillifera; salinity; nutrient level; specific growth rate (SGR); ratio of variable to
maximum fluorescence (Fv/Fm); non-photochemical quenching (NPQ)

1 INTRODUCTION Island (Shi, 2008), little is known about how salinity

The siphonous green macroalgae Caulerpa spp.
distribute across a wide range of tropical and
subtropical seas throughout the world. Caulerpa
lentillifera J. Agardh was originally described along
the Red Sea coast (Agardh, 1837) and was reported to
occur at many locations in the tropical Indo-Pacific
(Hackett, 1977; Taylor, 1977; Menez and Calumpong,
1982; Coppejans and Beeckman, 1989; Phillips et al.,
1999; Schils and Coppejans, 2003; Titlyanov et al.,
2012). Morphologically, with terete stolons bearing
erect fronds of spherical ramuli and filiform rhizoids,
C. lentillifera is one of the edible algae known as ‘sea
grapes’. It contains high levels of polyunsaturated
fatty acids (PUFA) (Saito et al., 2010) and multiple
essential amino acids and low levels of lipids (Niwano
et al., 2009). Although this alga has been cultivated
widely in the Philippines (Zemke-White and Ohno,
1999), Okinawa (Kurashima et al., 2003) and Taiwan
and nutrient levels affect growth of C. lentillifera. It is
crucial to clarify the effects of these parameters before
implementing large-scale cultivation.
Salinity is one of the most important abiotic
environmental factors to influence algal growth and
distribution (Lobban and Harrison, 1994). Many
studies have documented the effects of salinity on
several Caulerpa species, including C. paspaloides
(O’Neal and Prince, 1988) and C. taxifolia (Theil et
al., 2007; West and West, 2007). Deraxbudsarakom et
al. (2003) suggested that a salinity range of 25–30 is
required for normal growth of C. lentillifera. In
addition, Wang (2011) showed that maximum growth
of C. lentillifera occurred at a salinity of 36. We
* Supported by the Technology Program of Basic Research of Qingdao
(No. 12-1-4-8-(2)-jch)
** Corresponding author: dlduan@qdio.ac.cn
No.2 GUO et al.: Salinity and nutrients effects on C. lentillifera
B
S 2000). Fluorescence values become stable when algae
R of salinity-stressed Sargassum thunbergii decreased
Fig.1 Diagram of Caulerpa lentillifera, showing a branch
(B), the stolon (S) and a rhizoid (R)
studied the C. lentillifera strain from Okinawa, which
is likely to differ from strains from Thailand
(Deraxbudsarakom et al., 2003) or Vietnam (Wang,
2011). High salinity may influence the photosynthesis
of macroalgae by inactivating the reaction centers of
photosystem II (PSII) and inhibiting electron transport
(Xia et al., 2004).
Nitrogen (N) and phosphorus (P) are two essential
nutrients for algal growth (Carpenter and Capone,
1983). Insufficient or excess nutrients may impede
growth and affect chlorophyll fluorescence (Yin et al.,
2007; Liang et al., 2008; Qin et al., 2010). However,
different algal species have different tolerances to
varying concentrations of nutrients. The growth of
Gracilaria was inhibited at high (60 μmol/L) and low
(10 μmol/L) concentrations of NO3-N (Huang et al.,
2006). Concentrations of over 50 μmol/L of NO3-N
inhibited the growth of Kappaphycus alvarezii (Liu et
al., 2008). The growth rate of Ulva pertusa was faster
at a low concentration (10 mg/L) of nitrate, compared
with a high concentration (30 mg/L) (Qin et al., 2010).
The growth rate of Enteromorpha prolifera reached a
maximum in high N and P concentrations (N:
0.5 mmol/L, P: 0.03 mol/L) (Li et al., 2010).
Deraxbudsarakom et al. (2003) studied C. lentillifera
in Thailand and concluded that a NO3-N concentration
of 0.6 mmol/L and a N:P ratio of 8:1 were optimal for
growth.
Chlorophyll fluorescence parameters are widely
applied to assess photosynthetic features in algae
(Lichtenthaler et al., 2005). Energy absorbed by
chlorophyll molecules is distributed in one of three
ways, it can be used in photosynthesis
(photochemistry), or re-emitted as heat or as
chlorophyll fluorescence (Lichtenthaler, 1996).
Measuring the proportion of energy that is re-emitted
411
gives a measure of the efficiency of photosynthesis,
which can be used as an indicator of the health of the
plant. Therefore, measuring the ratio of variable to
maximum fluorescence (Fv/Fm) and non-
photochemical quenching (NPQ) can indicate the
physiological status of algae (Maxwell and Johnson,
grew under favorable conditions. When plants are
exposed to abiotic or biotic stress, decreases in Fv/Fm
are frequently observed (Baker, 2008). For example,
the ratio of variable to maximum fluorescence (Fv/Fm)
from 0.62 to 0.55 (Liang et al., 2011). Rattan et al.
(2012) showed that a depressed Fv/Fm value can
indicate a nutrient deficiency of algae under natural
conditions. NPQ is the release of energy as heat.
Increased levels of NPQ are usually caused by
stressed photosynthetic apparatus (Baker, 2008).
The objective of this study was to determine the
optimal salinity and nutrient levels for the asexual
reproduction of Caulerpa lentillifera. Understanding
the optimal conditions for maximum growth is
necessary before implementing large-scale
commercial propagation of this species.
2 MATERIAL AND METHOD
2.1 Algal pre-culture
Fresh Caulerpa lentillifera were transported from
Kumejima, Okinawa of Japan. The algae was washed
with sterile seawater several times, then placed in a
flask in an illuminating incubator (GXZ series)
(Jiangnan, Ningbo, China) and held at 25–26°C. Light
was provided using fluorescent tubes with an
irradiance of 10–20 μmol photons/(m2∙s) and a photo
period of 12 h:12 h (light:dark). Light irradiance was
measured with a LI-250 light meter (LI-COR, USA).
The algal thalli of Caulerpa lentillifera consists of
creeping stolons, erect branches and rhizoids (Fig.1).
Green, healthy thalli were selected and the erect
branches were cut into 5 cm sections with a scalpel.
To test the effects of salinity, algae were exposed to
one of nine saline concentrations (15, 20, 25, 30, 35,
40, 45, 50, and 55). Three replicate algal samples
were tested at each salinity. Saline solutions were
prepared using sea salt (Haidatongyong, Qingdao,
China) and distilled water. Salinities were measured
using a salinity meter (ATC, China). Algae were
cultured in flasks containing 100 mL of solution.
Algae were exposed to the temperature 25.0°C and a
light of 40 μmol photons/(m2∙s).
412 CHIN. J. OCEANOL. LIMNOL., 33(2), 2015
3 (Beckman, USA). The concentration of total
2.5
2 Total chlorophyll (mg/L)=8.02A663+20.21A645
SGR (%/d)
1.5
1
0.5
0
15 20 25 30 35 40 45 50 55
Salinity
Fig.2 SGR of Caulerpa lentillifera at different salinities
The water temperature was 25.0°C and algae were exposed to a light of
40 μmol photons/(m2∙s) (n=3). Error bar shows std. deviation.
2.2 Preparation of PO4-P and NO3-N solutions
Sodium nitrate (NaNO3) salt was weighed into
batches of 0.043 g, 0.085 g, 0.425 g, 0.850 g, 1.100 g,
and 3.400 g and then dissolved in distilled water and
made up to a final volume of 10 mL. Monopotassium
phosphate (KH2PO4) salt was weighed into batches of
0.014 g, 0.136 g, and 0.544 g and dissolved in distilled
water to a final volume of 10 mL. For each treatment,
300 μL of each NaNO3 and KH2PO4 solution was
added into 300 mL of autoclaved seawater (salinity
31). The final concentrations of NaNO3 solutions
were 0.05, 0.1, 0.5, 1.0, 2.0, and 4.0 mmol/L and the
final concentrations of KH2PO4 were 0.01, 0.1, and
0.4 mmol/L.
2.3 Specific growth rate measurements
At the beginning of culture, the fresh weight of
algae (W0) was measured using an electronic balance
(Mettler Toledo, Shanghai, China). Seven days later,
algae were observed and the algal fresh weight (W7)
was measured. Specific growth rate (SGR) for each
algal sample was calculated as follows:
SGR=ln(W7/W0)/t×100, where t is time elapsed.
2.4 Extraction and measurements of chlorophyll
Fresh branches of Caulerpa lentillifera were
ground in 80% (V/V) acetone using a mortar and
pestle, then transferred to a 2-mL centrifuge tube. The
tube was placed in the dark for 5 min, and then
centrifuged at 4 000×g for 15 min. The supernatant
was then transferred into a flask and acetone added to
make a total volume of 10 mL. The concentration of
chlorophyll was measured in the solution at
wavelengths of 630, 645, 647, 663, and 664 nm using
a UV-visible light spectrophotometer (DU650)
Vol.33
chlorophyll and chlorophyll a was calculated as
follows:
(Arnon, 1949);
Chlorophyll a (mg/L)=11.85A664–1.54A647–0.08A630
(Jeffrey and Humphrey, 1975).
Mass fraction was calculated using to the following
formula:
Mass fraction of pigment (mg/g)=(ρV)/m
(Ma, 2013),
in which ρ=mass concentration, V=total volume
(10 mL) and m=sample weight.
2.5 Chlorophyll fluorescence parameters
After seven days’ treatment, all the cultured algal
samples were held in the dark for 10 min. Chlorophyll
fluorescence parameters including the ratio of variable
to maximum fluorescence (Fv/Fm) and NPQ were
measured using a fluorescence image system
(FC1000-H) (PSI, Czech).
2.6 Statistical analysis
All data were tested with analysis of variance
(ANOVA) using the software package PASW 18.0
(SPSS). Tukey’s multiple range tests were applied to
evaluate differences between significant means (Sokal
and Rohlf, 1969). Analyses had a critical probability
level of 0.05.
3 RESULT
3.1 Effect of salinity on the growth rate of
C. lentillifera
Caulerpa lentillifera did not survive at salinities of
15 and 55, decaying within three days, and gradually
turning white and becoming soft (Fig.3). There was a
significant difference among the SGR of C. lentillifera
at salinities of 20–50 (ANOVA: F=6.084, P<0.05).
The maximum SGR of 2.038%±0.465% per day
occurred at a salinity of 35 (Fig.2).
At salinities of 20 and 45, only stolons regenerated
from branches. However, new branches with ramuli
grew from stolons at salinities of 30–40. The results
suggest that the optimum salinity for growth of
C. lentillifera was 35.
3.2 Effect of salinity on photosynthetic
characteristics
There was a significant difference in the chlorophyll
No.2 GUO et al.: Salinity and nutrients effects on C. lentillifera 413

15 20 25

30 35 40

45 50 55

Fig.3 Caulerpa lentillifera cultured in different salinities after 7 days

The numbers represent salinity. Scale bar: 1 cm.

0.35 data were generated for algae cultured at a salinity of

Chlorophyll a 50 as regenerated stolons did not form. Chlorophyll a
0.30
Total chlorophyll values were highest in algae cultured at a salinity of
0.25 35. Total chlorophyll increased in algae samples at
Content (mg/g)

30, reaching a maximum at 35 (Fig.4). This suggests

0.20 that a salinity of 35 is optimal for photosynthesis in C.
0.15
lentillifera.
There were significant differences among the
0.10 values for Fv/Fm (ANOVA: F=69.904, P<0.001) and
NPQ (F=14.349, P<0.001) in Caulerpa lentillifera
0.05
cultured at different salinities (Fig.5). Initial Fv/Fm
0
values at different salinities were around 0.85. After
20 25 30 35 40 45 seven days the Fv/Fm value decreased to 0.623 at a
Salinity salinity of 15, and the algae died or became completely
Fig.4 Chlorophyll content of regenerated stolons at white. At a salinity of 55, the Fv/Fm value decreased to
different salinities 0.710. The Fv/Fm values for algae cultured in salinities
Algae were cultured with a water temperature of 25.0°C and light of 25–45 were above 0.80, indicating that
of 40 μmol photons/(m2∙s) (n=3). Error bar shows std. deviation.
C. lentillifera was healthy (Fig.5a). The NPQ value
was highest (0.56) in algae cultured at a salinity of 15.
a content (F=12.198, P<0.05) and total chlorophyll It was also elevated to 0.12 in algae cultured at a
content (F=80.569, P<0.05) of Caulerpa lentillifera salinity of 55, but relatively low (0.10) in algae
cultured at different salinities (Fig.4). No chlorophyll cultured at salinities of 25–45 (Fig.5b).
414 CHIN. J. OCEANOL. LIMNOL., 33(2), 2015 Vol.33

0.90 a 6
PO4-P: 0.01 mmol/L
0.85 PO4-P: 0.1 mmol/L
5
PO4-P: 0.4 mmol/L
0.80
4
0.75

SGR (%/d)
Fv/Fm

0.70 3
0.65
0 day 2
0.60
7 day
0.55 1

0.50
0
15 20 25 30 35 40 45 50 55
0.02 0.1 0.5 1 2 4
Salinity NO3-N (mmol/L)

0.90 b Fig.6 SGR of the erect branches of C. lentillifera cultured at

a range of nitrogen and phosphorus concentrations
0.80 0 day
Error bar shows std. deviation.
0.70 7 day
3.4 Effect of NO3-N on photosynthetic characters
0.60

There was a general trend for chlorophyll content

Fv/Fm

0.50

0.40 (chlorophyll a and total chlorophyll) to increase with

0.30
0.20
0.10
0 concentration of 0.4 mmol/L (Fig.7). The maximum
15 20 25 30 35 40 45 50 55
Salinity
Fig.5 a. Ratio of variable to maximum fluorescence (Fv/Fm)
of the erect branches of C. lentillifera cultured at a
range of salinities (n=3); b. NPQ of the erect branches
of C. lentillifera cultured at a range of salinities (n=3)
Measurements were taken at the beginning of the test period and
again after seven days. Error bar shows std. deviation.
3.3 Effect of PO4-P on growth
There were significant differences in SGR
(F=3.374, P<0.05) among algae cultured in different
concentrations of NO3-N, when the PO4-P
concentration was 0.1 mmol/L. However there were
no significant differences in SGR among algae
cultured in different NO3-N concentrations (ANOVA:
F=2.244, P>0.05) when the PO4-P concentration was
0.01 mmol/L, or 0.4 mmol/L (F=0.782, P>0.05). This
suggests that the optimal concentration of PO4-P is
0.1 mmol/L. At a PO4-P concentration of 0.1 mmol/L,
the SGR of C. lentillifera increased with NO3-N
concentration, from 0.05 to 0.5 mmol/L, then leveled
out (Fig.6). SGR increased linearly with NO3-N
concentration in the 0.05–0.5 mmol/L range
(R2=0.999 3). This implies that growth rate may be
inhibited by high NO3-N concentrations.
rising NO3-N concentration at low concentrations of
PO4-P (0.01 mmol/L). In contrast, there was a trend
for levels of chlorophyll to gradually decrease at
increasing concentrations of NO3-N, at a PO4-P
chlorophyll content (0.150 1±0.037 7 mg/g) occurred
in algae cultured at a concentration of 0.1 mmol/L of
PO4-P. At a concentration of 0.1 mmol/L of PO4-P,
chlorophyll content decreased at NO3-N concentrations
of 1 mmol/L or greater (Fig.7), suggesting that
photosynthesis of new branches of C. lentillifera were
inhibited at high NO3-N concentrations.
There were no significant differences among Fv/Fm
values (P>0.05) at PO4-P concentrations of 0.01 and
0.1 mmol/L. At a PO4-P concentration of 0.4 mmol/L
the Fv/Fm value decreased rapidly at NO3-N
concentrations above 1 mmol/L (Fig.8a). The NPQ
value increased at high concentrations of NO3-N,
when the PO4-P concentration was 0.4 mmol/L
(Fig.8b). These results indicate that high PO4-P and
NO3-N concentrations have a negative effect on
photosynthesis in C. lentillifera.
4 DISCUSSION
Our experiments indicate that growth of Caulerpa
lentillifera is optimal at a salinity of 35, and although
the algae can survive salinities of 20–50, growth only
occurs in salinities of 20–45. Chlorophyll content and
Fv/Fm were also highest at a salinity of 35, indicating
optimum photosynthetic capacity was at salinity 35.
Our results are not consistent with those of
No.2 GUO et al.: Salinity and nutrients effects on C. lentillifera 415

0.2 0.35
a b
0.18
0.3
0.16

Total chlorophyll (mg/g)

Chlorophyll a (mg/g)

0.14 0.25
0.12 0.2
0.1
0.15
0.08
0.06 0.1
0.04
0.05
0.02
0 0
0.01 0.1 0.4 0.01 0.1 0.4
PO4-P (mmol/L) PO4-P (mmol/L)
NO3-N : 0.05 mmol/L NO3-N : 1 mmol/L
NO3-N : 0.1 mmol/L NO3-N : 2 mmol/L
NO3-N : 0.5 mmol/L NO3-N : 4 mmol/L

Fig.7 Ch a content (a) and total chlorophyll content (b) of regenerated stolons of C. lentillifera cultured at a range of
phosphorus and nitrogen concentrations
Water temperature was maintained at 25.0°C and light was 40 μmol photons/(m2∙s) (n=3). Error bar shows std. deviation.

0.90 a Deraxbudsarakom et al. (2003). These authors

0.85 examined the growth of C. lentillifera in Thailand,
and found that salinities of 25–30 were optimal for
0.80
fast growth. Caulerpa letillifera has been recorded to
0.75 occur in the Red Sea at salinities of 38–40 (Friedman
1968), indicating a tolerance to high salinities. In
Fv/Fm

0.70

0.65 PO4-P: 0.01 mmol/L addition, there are some similarities with
PO4-P: 0.1 mmol/L C. paspaloides (O’Neal and Prince, 1988) and
0.60 PO4-P: 0.4 mmol/L C. taxifolia (Theil et al., 2007; West and West, 2007).
0.55 Kirst et al. (1989) suggested that Caulerpa can resist
0.50 osmotic stress using a giant vacuolar system. Here we
0.05 0.1 0.5 1 2 4 deduced that C. lentillifera may possess a complicated
NO3-N (mmol/L) osmotic mechanism, however, further work is
required to examine the vacuolar system of this
1.20
b
PO4-P: 0.01 mmol/L
species.
1.00 PO4-P: 0.1 mmol/L The ratio of variable to maximum fluorescence
PO4-P: 0.4 mmol/L (Fv/Fm) is a sensitive parameter to indicate the degree
0.80 of stress in plants (Chen and Liu, 2007). Values of less
than the optimal value of 0.83 indicate that the plant
NPQ

0.60
has been exposed to stress (Maxwell and Johnson,
0.40 2000). In our study C. lentillifera with (Fv/Fm) values
of less than 0.80 showed relatively slow growth rates,
0.20 and plants with (Fv/Fm) values of less than 0.70 died.
SGR of C. lentillifera were highest at a PO4-P
0
0.05 0.1 0.5 1 2 4
concentration of 0.1 mmol/L and a NO3-N
NO3-N (mmol/L) concentration of 0.5 mmol/L. Deraxbudsarkom et al.
(2003) suggested that a NO3-N concentration of
Fig.8 Ratio of variable to maximum fluorescence (Fv/Fm)
(a) and NPQ (b)of the erect branches of C. lentillifera
0.6 mmol/L and N:P ratio of 8:1 were optimal for the
cultured in a range of phosphorus and nitrogen growth of C. lentillifera, and that nitrate is the
concentrations (n=3) preferable nitrogen source compared with nitrite.
Water temperature was maintained at 25.0°C and light was 40 μmol Huang (2012) found that the wet weight of
photons/(m2∙s). Error bar shows std. deviation. C. lentillifera was the highest when it was cultured in
416 CHIN. J. OCEANOL. LIMNOL., 33(2), 2015
nitrogen concentrations of 15 mg/kg (ca.
0.16 mmol/L). Our results indicated that a N:P ratio
of 5:1 was optimal for growth, differing slightly from
the 8:1 ratio described by Deraxbudsarkom et al.
(2003). These differences may be due to different
environmental conditions.
The SGR of C. lentillifera was strongly correlated
with NO3-N concentrations in the range of 0.05 to
0.5 mmol/L (R2=0.999 3), but growth rate was
relatively stable at NO3-N concentrations of greater
than 0.5 mmol/L. These results suggest that a NO3-N
concentration of 0.5 mmol/L is the upper threshold
for nutritional assimilation and absorption, due to
complicated enzyme behaviors for nitrate nitrogen
conversion and the reduction of ammonium nitrogen
in algae (Hockin et al., 2012). Chlorophyll is a key
compound for restoring nitrogen (Xu et al., 2006), it
will be degraded and release nitrogen primarily in
macroalgae when there is lack of nitrogen supply
(Bjomsater and Wheeler, 1990). This may explain
why the chlorophyll content of C. lentillifera was
lower at NO3-N concentrations of 0.05 mmol/L
compared with higher NO3-N concentrations when
the PO4-P concentration was low in our study.
At low concentrations of NO3-N (0.05 mmol/L)
and PO4-P (0.01 mmol/L), the Fv/Fm value for
C. lentillifera was also relatively low, suggesting that
photosynthetic capacity was impeded. Low nitrate
concentrations can cause a decrease in the number of
photosystem II (PSII) reaction center proteins (Kolber
et al., 1988; Berges et al., 1996). Phosphorus
deficiency decreases photochemical quantum yield by
reducing ATP and NADPH in plants (Rao et al., 1989;
Freden et al., 1990). The photosynthetic capacity of
Sargassum thunbergii seedlings decreased
significantly when cultured in low concentrations of
nitrogen (0.2 mg/L) and phosphorus (0.02 mg/L)
(Liang et al., 2011). The Fv/Fm value for the red alga
Gracilaria lemaneiformis, was 0.46 (Peng et al.,
2007). The Fv/Fm value for C. lentillifera reached
0.85, and this high value was attibuted to its close
relationship with higher plants (Björkman and
Demmig, 1987; Johnson et al., 1993).
In C. lentillifera the Fv/Fm value decreased rapidly
when the concentrations of NO3-N and PO4-P were
greater than 0.5 mmol/L and 0.4 mmol/L, respectively,
suggesting that consumption of nutrients resulted in a
decrease in the availability of electron transport and
assimilation of CO2. Consequently, more energy was
dissipated as heat from the PSII reaction center,
resulting in an increase in NPQ values for the alga.
Vol.33
During our indoor culture experiment, the culture
duration was limited to seven days, and only one
nitrogen source was applied. Further experiments
over a longer duration would help to optimize the
parameters for algal growth at a large-scale.
5 CONCLUSION
In conclusion, optimal conditions for vegetative
reproduction of C. lentillifera occurred at a salinity of
35, with a concentration of 0.1 mmol/L of PO4-P and
0.5 mmol/L of NO3-N. Further studies are required to
understand the optimal conditions to culture this algae
outdoors and at a large-scale.
6 ACKNOWLEDGEMENT
Thanks to the anonymous reviewers for their
critical comments and suggestions.
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