Joao C. M. Carvalho, Raquel P. Bezerra, Marcelo C. Matsudo, and Sunao Sato

Abstract This chapter comments on fed-batch cultivation of Arthrospiraplatensis under different carbon and nitrogen sources, pH, temperature, light intensity, type of photobioreactor and typical parameters of the fed-batch process, such as feeding time, addition protocol and flow rate. Inexpensive nitrogen sources, such as urea, ammonium salts and nitrogen-rich wastewaters can be used for A. platensis cultiva­tion, with results that can be comparable to those with classical nitrate sources. Closed photobioreactors are useful for preventing ammonia loss. The use of organic carbon sources needs to be carried out under aseptic conditions, and it is necessary to evaluate the best supplying conditions when using fed-batch process. The addi­tion of CO2 ensures the control of pH and, at the same time, supply of the carbon source into the culture medium. The fed-batch process can be useful for the produc­tion of A. platensis using CO2 from industrial plants, particularly from industrial alcoholic fermentation.

1 Introduction

The cultivation of microalgae and cyanobacteria is an important current issue because of the possibility of supplying human needs related to food production and removal of atmospheric or industrial carbon dioxide. Arthrospira (Spirulina) plat­ensis, Dunaliella salina and Chlorella vulgaris are among the most studied photo­synthetic microorganisms, but several other cyanobacteria and microalgae have been investigated lately, mainly for biodiesel production.

J. C.M. Carvalho (H) • R. P. Bezerra • M. C. Matsudo • S. Sato

Department of Biochemical and Pharmaceutical Technology, University of Sao Paulo, Av. Prof. Lineu Prestes 580, Bl. 16, Sao Paulo 05508-900, SP, Brazil e-mail: jcmdcarv@usp. br

J. W. Lee (ed.), Advanced Biofuels and Bioproducts, DOI 10.1007/978-1-4614-3348-4_33, 781

© Springer Science+Business Media New York 2013

The increasing demand for protein sources and other high biological value products, such as polyunsaturated fatty acids and pigments, associated with the need of the development of new technologies that contribute to the mitigation of environmental pollution indicates that the market for microorganisms such as A. platensis is going to increase in the coming years.

The previous uses of photosynthetic microorganisms as food are related to events in China 2,000 years ago, where Nostoc was used in periods of food shortage. Additionally, Spirulina sp. was consumed by the Aztecs in the Mexico Valley and by people living near Chad Lake in Central Africa [54]. They have been consumed by Africans, where French researchers first reported in 1940 the use of Spirulina platensis as food [54] .

Currently, the correct scientific designation for S. platensis is A. platensis [95] . Despite this, in this chapter, it was maintained the denomination given by the authors of the cited works. The genus Arthrospira (family Cyanophyceae) encompasses the photosynthetic cyanobacteria with helically coiled trichomes along the entire length of the multicellular filaments and visible septa (Fig. 1). The last characteristic dif­ferentiates this genus from true Spirulina which has invisible septa [16].

Arthrospira (Spirulina) platensis is one of the most promising microorganisms, among microalgae and cyanobacteria, not only to be used as food but also for other industrial applications because of its composition.

It contains a great amount of polyunsaturated fatty acids and pigments such as phycocyanin and zeaxantine [24]. Palmitic, linoleic, g-linolenic, and oleic acids are the predominant fatty acids in S. platensis. g-Linolenic is only found in significant amounts in breast milk, some fruits, species of fungi and cyanobacteria [25].

S. platensis is also an interesting source of chlorophyll, since this microorganism synthesizes only chlorophyll a, which is more stable than chlorophyll b, very com­mon in vegetables. Moreover the cell wall is composed of mucopolysaccharides and therefore easily digested [44], which is an advantage for the bioavailability of cell components.

This cyanobacterium shows low nucleic acid content in dry biomass (4-6%) in comparison with yeasts (8-12%) and other bacteria (20%) [3], so the daily intake of this biomass would not cause any damage to the human body [73].

Besides the high protein content in dry biomass, S. platensis shows a satisfacto­rily balanced amino acid content, possessing even methionine, which is absent in most microalgae [35]. About 20% of the cellular protein is represented by the main pigments in this microorganism, called phycobilins [82] .

This biomass also contains important vitamins such as cyanocobalamin (B12), pyridoxine (B6), thiamin (B1), tocopherol (E), and phylloquinone or phytonadione (K) [12]. Moreover, recent studies indicate that some trace elements such as chro­mium III [57] and selenium [19] can be accumulated in S. platensis biomass depend­ing on the cultivation conditions.

In fact, Spirulina spp. are noted in the literature as an alternative protein source, due to the high protein content in dry biomass (reaching as high as 70%), good digestibility, low nucleic acid content, and presence of vitamins, polyunsaturated fatty acids, immunomodulatory polysaccharides, pigments, and antioxidants [24] .

S. platensis is mainly used as a food supplement. One of the applications is the use of this microorganism as a source of pigments for food industries [33, 69]. S. platensis was also shown to act as a prebiotic, improving the growth in vitro of lactic acid bacteria such as Lactobacillus lactis, Lactobacillus delbrueckii and Lactobacillus bulgaricus [73]. For application in animal feed, some researchers have studied the use of S. platensis in aquaculture, to feed shrimp larvae, for instance [46].

Another important aspect of this microorganism is the possibility of obtaining bioactive compounds [21]. Since the 1980s, several studies have evaluated the use of S. platensis as a dietary supplement for intestinal disorders [39] , diabetes melli — tus, hyperglycemia [74], hyperlipidemia [67, 85], anemia [17], and hypertension [102). Moreover, it can act as an anti-inflammatory )101]. Recent studies also focused on the isolation of fatty acids, particularly the polyunsaturated ones, and pigments from photosynthetic microorganisms. Such characteristics indicate that this microorganism can be a source of molecules with potential use in pharmaceuti­cal and cosmetic industries as well. Besides, Arthrospira (Spirulina) spp. have been used for the removal of heavy metals from wastewater [27, 45], and it is important to emphasize its potential for CO2 biofixation [114], including CO2 from ethanol production plants [15, 40, 63].

In the large-scale production process, S. platensis can be easily cultivated due to the fact that it grows at high alkalinity and high salinity inorganic medium, with high content of carbonate and bicarbonate. These characteristics make it possible to inhibit or prevent contamination. Large-scale cultivation can thus be carried out in open ponds, which is very common in algae cultivation farms, where 5,000 m2 ponds are employed [93], even though there are several studies about their cultiva­tion in closed bioreactors. A. platensis biomass recovery is also facilitated due to its fi lamentous morphology.

Tacon and Jackson [104] list the following advantages for cultivation of S. platensis: they are able to use both organic and inorganic carbon sources; they exhibit a short generation time under optimum growth conditions; and they are easily cultivated in small areas. Cyanobacterial strains are carefully selected among collections around the world, which are periodically sub-cultured in the laboratory in order to maintain actively growing cells. Major criteria in the selection of strains are growth rate, biochemical composition, and resistance to environmental stress at each production site. It must be emphasized, however, that a strain that shows a good performance in the laboratory does not always display the same behavior in an outdoor open pond operation [93] .

Even though microalgae and cyanobacteria have been used by humans for a long time, microalgal biotechnology has only begun in the middle of the last century. Currently, 5,000 tons of dry microalgal biomass is marketed per year, representing up to US$1.25B [99]. The production of photosynthetic microorganisms consider­ably increased in the world due to the possibility of using this kind of culture for oxygen production and as a source of protein for food in space travels [9].

At the end of the 1970s, Sosa Texcoco Co., in Mexico, was the first responsible for large-scale Spirulina production [23, 93]. Afterward, several countries such as Taiwan, India, USA and Japan also started producing this cyanobacterium in open ponds [93]. Among the different cyanobacterial species, A. platensis stands out due to its characteristics related to cell composition, cell growth and cell recovery.