Research Report

Biological Responses of the Marine Diatom Chaetoceros simplex Ostenfeld to Different Treatments of Phosphate and Nitrate  

Mohamed Zein Alabdein Nassar1 , Hamdy Ramadan Galal2 , Hanan Mohamed Khairy1 , Sara Hamdy Rashedy1
1. National Institute of Oceanography and Fisheries, Hydrobiology Lab, Egypt
2. Botany Department, Faculty of Science, South Valley University, Egypt
Author    Correspondence author
International Journal of Marine Science, 2016, Vol. 6, No. 1   doi: 10.5376/ijms.2016.06.0001
Received: 28 Sep., 2015    Accepted: 19 Nov., 2015    Published: 01 Jan., 2016
© 2016 BioPublisher Publishing Platform
This is an open access article published under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article:

 Nassar M. Z., Galal H. R. M., Khairy H. M., and Rashedy S. H., 2016, Biological Responses of the Marine Diatom Chaetoceros simplex Ostenfeld to Different Treatments of Phosphate and Nitrate, International Journal of Marine Science, Vol.6, No.01 1-8 (doi: 10.5376/ijms.2016.06.0001)

Abstract

Chaetoceros simplex Ostenfeld of marine diatoms was isolated and purified from the sub-surface waters of the north-western part of the Red Sea, Egypt (Hurgada). The isolated grown cells are exposed to different concentrations of nitrate and phosphate to estimate their effects on the growth and some metabolic activities of the organism. In general, the results indicated thatconcentrations of 100 μM nitrate and 5μM phosphate stimulated the algal growth by 60% and 32%, respectively in the 8th day of culture. While, the values of 300 μM nitrate and 50 μM phosphate exhibited a decrease in the algal growth by 44% and 49%, respectively as compared with the control culture. The photosynthetic pigments (Chl-a, Chl-c and Carotenoids), total soluble proteins and carbohydrates of Chaetoceros simplex followed a similar pattern of change to that of growth in response to different treatments of nitrate and phosphate. While, the addition of different treatments of nitrate and phosphate to F/2 medium decreased the total lipids of C. simplex.  Statistical analysis of the data indicated that 5 μM phosphates and 50 μM nitrates were the highly significant and most effective concentrations that positively affect most of different metabolic activities of Chaetoceros simplex. Thus, Chaetoceros simplex culture could be used as larval feed or other purposes cultured using 50 μM nitrate and 5μM phosphate.

Keywords
Chaetoceros simplex; Nitrate; Phosphate; Metabolic activities

Introduction
The physiology of microalgae is affected by physicochemical factors such as water temperature, salinity, light intensity, nutrient concentrations, and pH. It, however, is hard to identify the influence of environmental factors on the microalgae directly in nature. Thus, the physiological experiments are generally applied to understand the mechanism of environmental impacts on the different metabolic activities of microalgae through the laboratory culture studies (Cho et al., 1998).
 
Concentrations of cells in phytoplankton cultures are generally higher than those found in nature. Microalgae cultures must therefore be enriched with nutrients to make up for the deficiencies in the seawater. Macronutrients include nitrate, phosphate (in an approximate ratio of 6:1), and silicate. Silicate is specifically used for the growth of diatoms that utilize this compound for production of an external shell. Micronutrients consist of various trace metals and the vitamins like, thiamine (B1), cyanocobalamin (B12) and sometimes biotin.
 
Two enrichment media that have been used extensively and are suitable for the growth of most marine microalgae are Walne medium and Guillard’s F/2 medium. Generally, the most common nutrient medium used is F/2 standard described by Guillard and Ryther (1993). The complexity and cost of the above culture media often excludes their use for large scale culture operations. Alternative enrichment media that are suitable for mass production of microalgae in large-scale extensive systems contain only the most essential nutrients and are composed of agriculture grade rather than laboratory grade fertilizers (Lavens and Sorgeloos, 1996). Krichnavaruka et al. (2005) studied the optimal growth conditions and the cultivation of Chaetoceros calcitrans in airlift photobioreactor, and they found that a modified of standard F/2 medium with a two folds of silica and phosphorus concentrations was illustrated to result in a better growth of this diatom. Vitamin B12 in the range from 1 to 3 μg·l−1 did not significantly affect the growth.
 
Growth rate of microalgae is influenced by environmental conditions (Renaud et al., 2002). Microalgae growth and composition may be influenced by nutrients like nitrate, phosphate and silicate (Paerl, 2009). The influence of nitrate and phosphate concentrations on growth, extracellular polysaccharide production, and fatty acid profile of marine diatoms has been reported (Liang et al., 2006). Modifications in culture medium such as nitrogen, phosphorus and silicate concentrations affect the growth rate of microalgae, cellular composition and fatty acid profile of the lipid fraction (Sanchez et al., 2000). Fried et al. (2003) evaluated the effects of nine different combinations of nitrate and phosphate concentrations on the algal growth by measuring relative chlorophyll levels. They concluded that both nitrates and phosphates have positive effects on the algal growth. However, these variables affect the algal growth independently of each other and there is no interaction between the two. This implies that both nitrates and phosphates are effective limiting nutrients that can be reduced to control algal profile. Hemalatha et al. (2012) studied the combined effects of temperature, nitrate and silicate on growth of the marine diatom, Chaetoceros simplex. The growth rate was directly proportional to nutrient concentration and temperature, whereas chlorophyll-a and biochemical composition were directly proportional to the nutrient concentrations. Karthikeyan et al. (2013) investigated the suitability of the marine diatoms, Chaetoceros curvisetus and Chaetoceros simplex for the removal of macronutrients from different wastewater. The growth and nitrate-phosphate removal properties were studied with nitrate, ammonium and urea nitrogen sources. The results indicated the Chaetoceros simplex was more efficient than Chaetoceros curvisetus and suitable for the removal of macronutrients when cultured with urea and nitrate nitrogen sources. Recently, Chaetoceros simplex was examined for its growth and biochemical compositions in different nitrate, phosphate and silicate concentrations by Hemalatha et al. (2014).
 
Aim of the work
The aim of the present work is to study the response of growth, photosynthetic pigments, carbohydrates, total soluble proteins and total lipids of Chaetoceros simplex Ostenfeld to different treatments of nitrates and phosphates.
 
Materials and Methods      
Isolation and purification of the organism
One liter of sub-surface seawaters was collected from Hurgada in carefully cleaned polyethylene bottle and passed immediately through plankton net of 100 μm mesh size to eliminate the macrozooplankton. This water was supplemented in the laboratory with F/2 medium (Guillard and Ryther, 1993). This is a common and widely used general enriched seawater medium designed for growing coastal marine algae, especially diatoms. This culture was poured in several conical flasks of 500 ml capacity (250 ml were added in each one) and incubated for 12 days at 25±1 °C under day light fluorescent lamps of about 6400 LUX. The algal cultures were supplied with dry air to provide CO2 necessary for photosynthesis, to prevent the settling of the cells at the bottom of the containers and to maintain the algae in suspension without mechanical stress. The grown cells were examined under a binocular research microscope and identified as Chaetoceros simplex Ostenfeld. For accurate identification of the isolated cells; scanning electron microscope (SEM–JEOL-JSM5300) was used according to Ashour (2011) (Fig.1). Generally, the isolation and purification of C. simplex was carried out by the dilution method (Droop, 1954) and Picking up Capillary method (Stein, 1973).
 

 

Figure 1 Scanning electron micrograph of Chaetoceros simplex Ostenfeld (Ashour, 2011) 

 

Preparation of different treatments of nitrate and phosphate

In the collected water samples from Hurgada; the concentration of nitrate was 2.27μM and the phosphate value was 0.5 µM (Nassar et al., 2014). Thus, in this study, a six different treatments of nitrate were used as follows; 1 µM (below detected concentration), 5 µM, 50 µM, 100 µM, 200 µM and 300 µM (above detected concentration). Whereas, five different concentrations of phosphate were used as follows; 0.3 µM (below detected concentration), 1 µM, 5 µM, 25 µM and 50 µM (above detected concentration). Totally three triplicate experiments at the different treatments of phosphate and nitrate were carried out separately for 12 days under temperature 25±1 °C and day light fluorescent lamps of about 6400 LUX, pH 8.0–8.4 and salinity 35 ‰.

 

Determination of growth
Algal cells were counted every 48 hours intervals with haemocytometer (0.1 mm depth) under a binuclear research microscope with a magnification power of 480x and the counts were expressed as cells·ml-1.
 
Determination of different metabolic variables
The chlorophylls (a and c) were estimated according to the method recommended by Parsons and Strickland (1965). The total carbohydrate content was measured by the phenol-sulphuric acid method (Herbert et al., 1971). Total soluble protein was determined quantitatively using the method described by Lowry et al. (1951). The total lipids of algal cellswere extracted according to Bligh and Dyer (1959).
 
Statistical Analysis
The ANOVA analysis was carried out for the obtained data on the computer using the program of STATISTICA at p < 0.01.
 
Results and Discussion
The chemical well as to the experimental conditions applied like temperature, light content of microalgae can vary with culture age and with changes in culture conditions (Araújo and Garcia, 2005). The effect of variation of these parameters on many algal species has been studied by several workers (Uriarte et al., 1993; El-Sherif, 1993; Hemalatha et al., 2012, 2014; Abdel-Hamid et al., 2015). Data on the chemical composition of microalgae may also vary widely due to differences of the methods of measurement used (Barbarino and Lourenço, 2005); the physiological state of the microalgae, as intensity, medium cultivation or in outdoors conditions (Banerjee et al., 2011). In addition, due to the interaction of the organisms with the culture medium, a batch culture is under a continuous chemical change. These variations reflect on the cell metabolism and consequently on their chemical composition (Lourenço et al., 2002).
 
Growth of Chaetoceros simplex
Nutrients are among the most important factors controlling phytoplankton growth. However, nutrient contents in marine environments greatly change over space and time (Eker-Develi et al., 2006).In the present study, the concentrations of 1, 5, 50 and 100 μM of nitrate stimulated the algal growth of Chaetoceros simplex by 7, 14, 26 and 60%, respectively over the control culture in the 8th day of culture. While, the values of 200 and 300 μM nitrate caused about 13 and 44 % growth reduction, respectively below the control culture. On the other hand, phosphate concentrations of 0.3, 1 and 5μM stimulated the growth of Chaetoceros simplex by 11, 23 and 32 %, respectively, while, the values of 25 and 50 μM exhibited a decrease in the algal growth by 19 and 49 %, respectively (Tables 1, 2 and Figs.2, 3). In general, the maximum growth of Chaetoceros simplex 330×104 cell·ml-1 and 310×104 cell·ml-1 were achieved at the value of 100 μM nitrate and 5 μM phosphates. While, the concentrations of 200 and 300 μM nitrate and 25 and 50 μM phosphate showed lower growth than the other treatments. These nutrients are usually involved in many biochemical reactions including proteins, nucleic acids, chlorophylls and amino acids biosynthesis in the photosynthetic organisms (Geider and La Roche, 2002; Van Mooy et al., 2009 and Van Mooy et al., 2009). However, the growth of Chaetoceros simplex decreased gradually after eight days of incubation may due to the expected gradual decrease of nitrate and phosphate concentrations with time as well as excretions of unfavorable metabolites in the culture media. Padhi et al. (2009)reported that the maximum growth of Chaetoceros curvisetus was foundat 3 mM NaNO3 and slightly declined at 6 mM NaNO3. Hemalatha et al. (2012) stated that the combination effect of temperature and nutrients on Chaetoceros simplex, recording the maximum cell density of 18.1×105 cells·ml-1 at 20 ºC under 1764 μM nitrate in the 10th day of culture, whereas at 25ºC and 29 ºC, the maximum cell density reached 18.89×105 cells·ml-1 and 23.5 ×105 cells·ml-1, respectively. Also, Hemalatha et al.(2014) found that the maximum cell density of Chaetoceros simplex was reached in the culture treated with 1764 μM nitrate (18.23×105 cells·ml-1) and 72.4 μM phosphate (18.21×105 cells·ml-1). The growth rate was enhanced with increasing of nutrients up to the concentration 2646 μM nitrate and 90.5 μM phosphates. However, the concentrations of 1764 μM nitrate and 72.4 μM phosphate that persistent the highest cell densities of Chaetoceros simplex by Hemalatha et al. (2012, 2014) are extremely higher than recorded in the present study. This may be due to the combination effects of temperature and the nutrients; nitrate, phosphate and silicate in the culture F/2 media. This result may be also due to the differences in the environmental conditions, including salinity of seawater, pH value, dissolved oxygen concentrations and the nutrient salts of the filtered seawater that used in the culture F/2 media in the present study.

 

 

Figure 2 Effect of different concentrations of nitrate (μM) on the growth of Chaetoceros simplex (Cell×104·ml-1) (Each value is the mean ± SD)

 

 

Figure 3 Effect of different concentrations of phosphate (μM) on the growth of Chaetoceros simplex (Cell×104·ml-1) (Each value is the mean ± SD)

 

 

Table 1 Effect of nitrate (µM) on the growth of Chaetoceros simplex (Cell×104·ml-1) (Each value is the mean ± SD)

 

 

Table 2  Effect of phosphate (μM) on the growth of Chaetoceros simplex (Cell×104·ml-1) (Each value is the mean ± SD)

 

Photosynthetic pigments

Chlorophylls are key compounds in plants for trapping light energy for photosynthesis, thus their quantitative determination is of great importance in the studies of photosynthesis, primary production and related subjects. Chlorophyll-a, chlorophyll-c and the accessory photosynthetic pigment Carotenoids were measured in this study.
 
The results in Fig. 4, 5, 6, 7, 8 and 9 indicated that, chl-a, chl-c and carotenoids content of Chaetoceros simplex followed a similar pattern of change to that of growth in response to different treatments of nitrate and phosphate. The addition of 100 μM nitrate and 5 μM phosphate to the culture medium recorded a maximum Chl-a of 0.72 ±0.02 μg·ml-1 and 0.69±0.05 μg·ml-1, respectively as compared with the control culture. While, the concentrations of 300 μM nitrate and 50 μM phosphate sustained the lowest chlorophyll-a of 0.35±0.02 μg·ml-1 and 0.36±0.02 μg·ml-1, respectively. This result is in accordance with that obtained by De la Curz et al. (2006) who reported that chlorophyll–a content was directly related to cellular density and indirectly to concentration of nutrients. In the present study, the chlorophyll-a content sustained its highest value at the early exponential growth phase, which coincided with the data reported by Hemalatha et al. (2014). The chl-c and carotenoid contents showed an increasing trend up to 100 μM nitrate and 5 μM phosphates. The addition of 100 μM nitrate to the culture medium recorded a maximum chlorophyll-c of 0.52±0.02 μg·ml-1 and carotenoids value of 4.1±0.03 μg·ml-1, while additions of 5 μM phosphate sustained chl-c of 0.49±0.03 μg·ml-1 and carotenoids of 3.4±0.02 μg·ml-1).

 

 

Figure 4 Effect of different concentrations of nitrate (μM) on the chlorophyll-a (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

  

 

Figure 5 Effect of different concentrations of phosphate (μM) on the chlorophyll-a (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

  

 

Figure 6 Effect of different concentrations of nitrate (μM) on the chlorophyll-c (μg·ml-1) of Chaetoceros simple (Each value is the mean ± SD)

 

 

Figure 7 Effect of different concentrations of phosphate (μM) on the chlorophyll-c (μg·ml-1) of Chaetoceros simplex. (Each value is the mean ± SD)

 

 

Figure 8 Effect of different concentrations of nitrate (μM) on the carotenoids (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

  

 

Figure 9 Effect of different concentrations of phosphate (μM) on the carotenoids (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

 

Total soluble proteins

The total soluble proteins of Chaetoceros simplex in the present study were increased in the culture treated with 100 μM nitrate and 5 μM phosphates by about 37 and 32 %, respectively over the control culture after eight days of incubation. On the other hand, the concentrations of 300 μM nitrate and 50 μM phosphate inhibited the total soluble proteins of the tested organism by about 33 and 38%, respectively, below the control culture (Tables 3 and 4). Hemalatha et al. (2014) indicated that protein content of Chaetoceros simplex showed the maximum values at 2205 μM nitrates and 90.5 μM phosphates. It is generally recognized that the protein content of microalgae depends on the amount of nitrogen available in the culture medium; an increase in nitrogen concentration in the medium may increase the cellular protein content of the organism (Li et al., 2005).

 

 

Table 3 Effect of different concentrations of nitrate (μM) on the total carbohydrates, total soluble proteins and total lipids (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

 

 

Table 4 Effect of different concentration of phosphate (μM) on the total carbohydrates, total soluble proteins and total lipids (μg·ml-1) of Chaetoceros simplex (Each value is the mean ± SD)

 

Carbohydrates
The main storage compounds of diatoms are lipids (TAGs) and a β-1, 3-linked carbohydrate known as chrysolaminarin, which are sun light driven cell factories that convert carbon dioxide to potential biofuels, foods and feeds (Walter et al., 2005). In the present study, the carbohydrates showed increasing trends with increase of nitrate and phosphate in the culture media up to 100 μM nitrate and 5 μM phosphates and then it gradually decreased. The addition of 5 μM phosphate to the culture medium of Chaetoceros simplex recorded a maximum carbohydrate of 54.6±3.5 μg·ml-1 and the addition of 100 μM nitrate sustained the highest value of 50±5.3μg·ml-1. Whereas, the minimum carbohydrate content was observed with the concentrations of 300 μM nitrate and 50 μM phosphate (Tables 3 and 4).

 

Total Lipids
Regarding to the lipid content of C. simplex, the addition of different treatments of nitrate and phosphate to F/2 medium decreased the total lipids of C. simplex as compared with the control culture, which exhibited the highest value of 86±5μg·ml-1 in the 8th day of culture. While, the lowest contents of 40±3 and 46.3±15 μg·ml-1 were observed in the culture treated with 50 μM phosphate and 300 μM nitrate, respectively (Tables 3 and 4). Generally, Praveenkumar et al. (2012) reported that nitrogen is the most critical nutrient affecting the lipid metabolism in the algae and the accumulation of lipids in response to nitrogen deficiency has been observed in numerous species and strains of various microalgae. Also, the effects of sodium nitrate as a nitrogen source on the cell growth and lipid content had been studied by Li et al. (2008) in the green alga, Neochloris oleoabanddans (one of the most promising oil–rich microalgal species). They found that the highest lipid content was obtained at the sodium nitrate concentration of 3 mM, whereas the lowest lipid content was observed at 5 mM. Furthermore, Kumari et al. (2013) concluded that decreasing in nitrate and phosphate concentrations caused reduction in the cell division rate of the algae, surprisingly activates the biosynthesis of storage lipids.
 
Statistical Analysis
The ANOVA analysis of the data in Tables 1-4, indicated that 5 μM phosphate was the highly significant and most effective concentration that positively affects the cell density and the photosynthetic pigments (Chl-a, Chl-c and Carotenoids), total soluble protein, carbohydrates  of Chaetoceros simplex as compared with the control culture (F = 38.241, p < 0.01). Whereas, the value of 50 μM nitrate was the most effective value that positively affects the above metabolic activities of the organism (F = 1312, p < 0.07). But the statistical analysis didn't appear any significance for the total lipids.
 
Conclusion
The growth and biochemical composition of Chaetoceros simplex changed significantly than the control F/2 medium. As a general trend, the concentrations of 100 μM nitrate and 5μM phosphate stimulated the algal cell density, photosynthetic pigments (Chl-a, Chl-c and Carotenoids), total soluble proteins and carbohydrates of the marine diatom C. simplex in the 8th day of culture as compared with the control culture. While, the addition of different treatments of nitrate and phosphate to F/2 medium decreased the total lipids of C. simplex. Based on the analysis of ANOVA; theconcentrationsof 5 μM phosphates and 50 μM nitrates were the most suitable treatments in the present study, that positively affect most of different metabolic activities of Chaetoceros simplex. Thus, Chaetoceros simplex culture could be used as larval feed or other purposes cultured using 50 μM nitrate and 5μM phosphate.
 
References
Abdel-Hamid M. I., Belal S. A., Azab Y. A., Abdel-Mogib M., and Abdel-Aal E. I., 2015, Nutritional value of some selected green microalgae, Mansoura Journal of Environmental Sciences - Egypt JOESE. 44(3): In Press. 
 
Araújo S.C., and Garcia V.T., 2005, Growth and biochemical composition of the diatom Chaetoceros cf. wighamii Brightwell under different temperature, salinity and carbon dioxide levels, I. Protein, carbohydrates and lipids. Aquaculture. 246: 405-412.
    http://dx.doi.org/10.1016/j.aquaculture.2005.02.051
 
Ashour M. A., 2011, Studies on culture of some marine microalgae and its utilization as feed for some invertebrates and fish larvae, M.Sc. thesis, Department of Animal Production, Faculty of Agriculture, Tanta University
 
Banerjee S., Hew W. E., Khatoon H., Shariff M., and Yusoff F. M., 2011, Growth and proximate composition of tropical marine Chaetoceros calcitrans and Nannochloropsis oculata cultured outdoors and under laboratory conditions, African Journal of Biotechnology, 10: 1375-1383.
 
Barbarino E. and Lourenço S. O., 2005, An evaluation of methodologies for extraction and quantification of protein of marine macro- and microalgae, Journal of Applied Phycology, 17: 447-460.
    http://dx.doi.org/10.1007/s10811-005-1641-4
 
Bligh E. G. and Dyer W.J., 1959, A rapid method for total lipid extraction and purification, Canadian Journal of Biochemistry and Physiology, 37: 911-917.
    http://dx.doi.org/10.1139/o59-099 PMid: 13671378
 
Cho J.H., Lee T.K., Shim K., Lee W.S., and Chang M., 1998, Changes of biochemical composition of Prorocentrum minimum causing red tide in different light intensities, Korean Journal of Environmental Biology, 16: 391-396.
 
De La Cruze F. L., Espinoza E. V., Trees R. M., Del-Angel S., and Nunez-Cebrer F., 2006, Nutrient uptake, Chlorophyll a and carbon fixation by Rhodomonas sp. Cultured at different irradiance and nutrient concentrations, Aquacultural Engineering, 1:51-60.
    http://dx.doi.org/10.1016/j.aquaeng.2005.08.004
 
Droop M.R., 1954, A note on the isolation of small marine algae and flagellates for pure cultures, Journal of Marine Biological Association of the United Kingdome, 3: 511-514.
    http://dx.doi.org/10.1017/S002531540000850X
 
Eker-Develi E., Erkan Kideys A. E., and Tugrul S., 2006, Effect of nutrients on culture dynamics of marine phytoplankton, Aquat. Sci., 68(1): 28-39.
    http://dx.doi.org/10.1007/s00027-005-0810-5
 
El-Sherif Z. M., 1993,Effect of nitrate, phosphate and salinity on the growth of the unicellular algae Nannochloropsis salina and Isochrysis galbana, Bull. High Inst. of Public Health, Alexandria, XXIII (4): 777-787.
Fried S., Mackie B., and Thwehr E., 2003, Nitrate and phosphate levels positively affect the growth of algae species found in Perry Pond, Tillers, 4: 21-24.
 
Guillard R. R. L., and Ryther J.H., 1993, Culture Methods. In: Manual on Harmful Marine Microalgae (eds. G.M. Hellegraeff, D.M. Anderson and A.D. Cembella), pp. 45-62. IOC Manuals and Guides No. 33. UNESCO.
 
Hemalatha A., Karthikeyan P., Anantharaman P., Sampathkumar P., and Manimaran K., 2012, Effect of Temperature on the Growth of Marine Diatom, Chaetoceros simplex (Ostenfeld, 1901) with different nitrate: silicate concentrations, Asian Pacific Journal of Tropical Biomedicine, S1817-S1821.
    http://dx.doi.org/10.1016/S2221-1691(12)60501-2
 
Hemalatha A., Karthikeyan P., Girija K., Saranya C., Anantharaman P., and Sampathkumar P., 2014, Effect of nutrients on the growth and biochemical composition of the marine diatom, Chaetoceros simplex (Ostenfeld, 1901), International Journal of phytopharmacy research, 5(1):30-35.
 
Herbert D., Phipps P.J., Strange P. E., 1971, Chemical analysis of microbial cells, Methods Microbiol. VB: 249-344.
 
Karthikeyan P., Manimaran K., Sampathkumar P., and Rameshkumar L., 2013, Growth and nutrient removal properties of the diatoms, Chaetoceros curvisetus and C. simplex under different nitrogen sources. Applied Water Science. 3:49–55.
    http://dx.doi.org/10.1007/s13201-012-0056-z
 
Krichnavaruka S., Worapannee L., Sorawit P., and Prasert P., 2005, Optimal growth conditions and the cultivation of Chaetoceros calcitrans in airlift hotobioreactor. Chemical Engineering Journal. 105: 91–98.
    http://dx.doi.org/10.1016/j.cej.2004.10.002
 
Kumari R., Kumar M., Reddy C. R. K., and Jha B., 2013, Algal lipids, fatty acids and sterols. Woodhead publishing limited. 3:87-134.
 
Lavens P., and Sorgeloos P., 1996, Manual on the Production and Use of Live Food for Aquaculture, FAO, Laboratory of Aquaculture and Artemia Reference Centre, University of Ghent, Ghent, Belgium.
 
Li M., Gong R., Rao X., Liu Z. and Wang X., 2005, Effect of nitrate concentration on growth and fatty acid composition of marine microalgae Paviova viridis, Annals of microbiology. 55: 51-55.
 
Li Y., Horsman M., Wang B., Wu N., and Lan C.Q., 2008, Effect of nitrogen sources on cell growth and lipid accumulation of green alga Neochloris oleoabanddans. Applied Microbiology and Biotechnology. 81: 629 – 636.
    http://dx.doi.org/10.1007/s00253-008-1681-1 PMid: 18795284
 
Liang Y., Beardall J., and Heraud P., 2006, Effects of nitrogen source and UV radiation on the growth, chlorophyll fluorescence and fatty acid composition of Phaeodactylum tricornutum and Chaetoceros muelleri (Bacillariophyceae). Journal of Photochemistry and Photobiology B: Biology. 82: 161–172.
    http://dx.doi.org/10.1016/j.jphotobiol.2005.11.002 PMid:16388965
 
Lourenço S. O., Barbarino E., Mancini-Filho J., and Schinke K. P., 2002, Effects of different nitrogen sources on growth and biochemical profile of ten marine microalgae under batch cultures. Phycologia. 41:158-168.
    http://dx.doi.org/10.2216/i0031-8884-41-2-158.1
 
Lowry O.H., Rosebrough N.N., Farr A.L. and Randall R.Y., 1951, Protein measurement with the Folin Phenol reagent. Journal of Biological Chemistry. 193:265- 275. PMid: 14907713
 
Nassar M.Z., Hamdy R.M., Khairy H.M., and Rashedy S.H., 2014, Seasonal fluctuations of phytoplankton community and physico-chemical parameters of the north western part of the Red Sea, Egypt. Egyptian Journal of Aquatic Research, 40 (4): 395-403.
    http://dx.doi.org/10.1016/j.ejar.2014.11.002
 
Padhi S.B., Behera G., Swain P.K., Behura S.K., and Dash P.K., 2009, Effect of environmental factors on growth and biochemical composition of Chaetoceros curvisetus for use in mariculture. Indian Hydrobiology. 12(1): 58-64.
 
Paerl H.W., 2009, Controlling eutrophication along the freshwater-marine continuum: dual nutrient (N and P) reductions are essential. Estuaries and Coasts. 32: 593-601.
    http://dx.doi.org/10.1007/s12237-009-9158-8
 
Parsons T.R., and Strickland J.D.H., 1965, Particulate organic matter III. I. Pigment analysis III, I.I. Determinations of phytoplankton pigments. J. Fish. Res. 18:117-127.
 
Praveenkumar R., Shameera K., Mahalakshmi G., Akbarsha M.A., and Thajuddin N., 2012, Influence of nutrient deprivations on lipid accumulation in a dominant indigenous microalga Chlorella sp., BUM11008: Evaluation for biodiesel production. Biomass Bioenergy.  37: 60-66.
    http://dx.doi.org/10.1016/j.biombioe.2011.12.035
 
Renaud S. M., Luong-Van T., George L., and David L. P., 2002, Effect of temperature on growth, chemical composition and fatty acid composition of tropical Australian microalgae grown in batch cultures. Aquaculture 211: 195–214.
    http://dx.doi.org/10.1016/S0044-8486(01)00875-4
 
Sanchez S, Martınez ME, Espinola F., 2000, Biomass production and biochemical variability of the marine microalga Isochrysis galbana in relation to culture medium, Journal of Biochemical Engineering.6: 13–18.
    http://dx.doi.org/10.1016/S1369-703X(00)00071-1
 
Stein J. R., 1973, Handbook of phycological methods. Cambridge: Cambridge University Press.
Uriarte I., Farías A., Hawkins A. J. S., and Bayne B. L., 1993, Cell characteristics and biochemical composition of Dunaliella primolecta Butcher conditioned at different concentrations of dissolved nitrogen. Journal of Applied Phycology. 5:447-453.
    http://dx.doi.org/10.1007/BF02182737
 
Van Mooy B.A.S., Fredricks H.F., Pedler B.E., Dyhr- man S.T., Karl D.M., and Koblížek M., 2009, Phytoplankton in the ocean use non-phosphorus lipids in response to phosphorus scarcity. Nature. 458: 69-72.
    http://dx.doi.org/10.1038/nature07659 PMid: 19182781
 
Walter T.L., Purton S., Becker D.K., and Collet C., 2005, Microalgae as bioreactor, Plant Cell Reports. 24: 629-641.
    http://dx.doi.org/10.1007/s00299-005-0004-6 PMid: 16136314

 

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