Effect of nitrogen and phosphorus amendment on the yield of a Chlorella sp. strain isolated off the Lebanese coast.

Microalgae are prokaryotic or eukaryotic microorganisms characterized by their efficient photosynthetic activity, capacity to survive in different environments and high rate of reproduction (Mata, 2010). The most common habitat of many microalgae is open waters, thus constituting the phytoplankton. The large biodiversity of microalgae manifested in many marine studies (Norton et al., 1996; Cappo et al., 2003; Guiry, 2012), coupled with their ability to accumulate high quantities of biomass within a relatively short time, has attracted the attention of researchers since several decades as potential source of food, feed and feedstock for renewable bioenergy (Sheehan et al., 1998; Chisti, 2007; Chisti, 2008). 

Given their simple structures and photosynthetic ability, microalgae are capable of rapidly generating important primary and secondary metabolites such as lipids, proteins, carbohydrates and antioxidants, from which high value products including food and feed supplements, industrial chemicals, para-pharmaceutical, pharmaceuticals (Borowitzka, 2013) and biofertilizers (Rani et al., 2008). 

Microalgae are often used as food and feed due to their nutritional value. Many species are known to have double the protein content (up to 60%) of the traditional protein supplements like meat or eggs, and contain essential amino acids that are responsible of the major metabolic processes such as energy and enzyme production, high amounts of simple and complex carbohydrates which provide the body with a source of additional energy, an extensive fatty acid profile, including Omega 3 and Omega 6, as well as an abundance of vitamins, minerals, and trace elements; hence, they are being cultivated to be used as food or food supplements (Kay & Barton, 1991). For instance, Spirulina (Arthrospira) is often used commercially as a nutritional supplement to treat for malnutrition because it has high protein content and other important nutrients (Habib et al., 2008). Microalgae are also used as animal feed or feed supplements. Improvement in growth rates, carcass quality and coloration, increase in survival rates, reduction in the requirement for medication and higher immunity are the main benefits associated with the use of feed containing microalgae biomass (Belay et al., 1996). However, the high cost of most of these algae may limit their commercial uses to few applications. 

Microalgae are exposed to a variety of environmental factors and nutrient availability that influence the growth rate and cellular composition in both natural and engineered systems. Hence, to develop a suitable high productivity bio-algal system, understanding synergistic interactions between multiple nutritional factors and environmental variables is essential (Radzun et al., 2015). 

Microalgae require inorganic nutrients, light, and favorable temperatures to grow; the primary inorganic nutrients are nitrogen and phosphorus (Fogg, 1973; Bold and Wynne, 1978). In laboratory studies, Guillard’s F/2 medium and Walne medium are the two enriched media extensively used for the growth of most marine microalgae except for cyanobacteria (Lavens and Sorgeloos, 1996; Bartley et al., 2013; Roleda et al., 2013). These media contain a mixture of macro- and micromineral elements and some vitamins required for microalgae growth. However, for large-scale production, agricultural grade fertilizers such as urea, ammonium sulfate and calcium superphosphate may be used as cost effective alternatives for the macronutrients (Lavens and Sorgeloos, 1996). Microalgae growth, lipid and protein content or composition can be altered by the composition of culturing media. The form and concentration of nitrogen in the medium has been found to affect growth significantly. When both ammonium and nitrate are available in a culture, ammonium is often preferred over nitrate as a nitrogen source since ammonium does not need to be reduced prior to amino acid synthesis (Grobbelaar, 2004). However, ammonium concentrations greater than 25μM are reported to be toxic to phytoplankton (Grobbelaar, 2004). Nitrogen starvation has been extensively studied as a means of increasing total lipid production for using microalgae biomass as feedstock for biodiesel production (Huang et al., 2012). However, relatively fewer studies were conducted on the effect of nitrogen supplementation on the production of primary or secondary metabolites, and considerable variations in the response of various isolates were reported (Ratha et al., 2013; Aremu et al., 2016). 

Temperature is a major influencing factor for optimal growth of microalgae as temperature can go below or above the optimal for each species. Microalgae can tolerate a temperature ranging from 18-24°C depending on the species and culture media. However, some microalgae can survive beyond that range. Temperatures lower than 16°C will slow down growth, while those higher than 35°C become lethal to many species. The effect temperature exerts on biochemical reactions and how it affects the biochemical composition of algae makes temperature one of the most important environmental factors (Hu, 2004; Wei et al., 2015).

The aim of this study was to select, identify and culture a local strain of microalgae promising for potential biotechnological exploitation in the MENA (Middle East and North Africa) region. The isolate was cultured under optimal laboratory and greenhouse conditions experimenting on the effect of two major nutrient requirements (nitrogen and phosphorus) on the total biomass production, and on its protein, lipids and chlorophyll content. The isolated strain was identified as a taxon of the green alga Chlorella, a genus already known for its commercial by-products.