4.2 CARBON DIOXIDE FIXATION BY MICROALGAE .1 Carbon Dioxide’s Role in Photobioreactors
4.2.3 Carbon Fixation of Industrially Important Microalgae
Carbon fixation by microalgae is in vogue. In the last decade, more than 4,000 papers were published globally on this subject.Table 4.1presents some rates of carbon dioxide described in the literature.
Among all species of microalgae, four are most common industrially:Spirulina,Chlorella, Dunaliella,andHaematococcus. Despite not being used industrially,Botryococcusis also largely studied due to its potential use as a source of hydrocarbons. These microalgae’s potential for carbon fixation is discussed next.
5 4.5 4 3.5 3 2.5 2 1.5 CO2 cons
1 0.5 0
1 25 49 73 97 121 145 169
Time (hours)
193 217 241 265 289 313 337 361
O2 cons
-3.5 -2.5 -1.5 -0.5 0.5 0 1 1.5 2
-3 -2 -1
CO2 consumed (g/h) CO2 base line (g/h) O2 Base Line (g/h) O2 consumed (g/h)
FIGURE 4.3 Gas phase analysis carried bySydney et al. (2011)showing the carbon consumption and oxygen production profiles.
75
4.2 CARBON DIOXIDE FIXATION BY MICROALGAE
4.2.3.1 Chlorella vulgaris
The first photosynthetic microbe to be isolated and grown in pure culture was the fresh- water microalga Chlorella vulgaris. It is a spherical unicellular eukaryotic green algae that presents a thick cell wall (100–200 nm) as its main characteristic. This cell wall provides mechanical and chemical protection, and its relation to heavy metals resistance is reported, which explains whyC. vulgarisis one of the most used microorganisms for waste treatment.
The uptake of carbon byC. vulgariscells is done through the enzyme carbonic anhydrase, which catalyzes the hydration of CO2to form HCO3 and a proton. Hirata and collaborators (1996) studied carbon dioxide fixation by this microalga, which showed important variations comparing cultivation under fluorescent lamps and sunlight. In the first case the estimated rate of carbon dioxide fixation was 865 mg CO2L1d1; in a sunlight regimen the estimated rate achieved 31.8 mg CO2L1d1. Winajarko et al. (2008) achieved a transferred rate of 441.6 g CO2L1d1under the same cultivation conditions asHirata et al. (1996). According toSydney et al. (2011), in experiments using classic synthetic media and a 12-h light/dark regimen,C. vulgarisbiofixation rate of carbon dioxide is near 250 mg L1day1.
Carbon fixation byChlorella vulgarisis variable and depends, among other factors, on the concentration of CO2in the gaseous source. Yun et al (1997) cultivated C. vulgarisin 15%
of carbon dioxide and achieved a fixation of 624 mg L1day1;Scragg et al. (2002)achieved a fixation of 75 mg L1day1under CO2concentration of 0.03%. In the same study, Scragg tested a medium with low nitrogen and the fixation rate was 45 mg L1day1, suggesting that nitrogen also influences carbon uptake rate.
Some studies (Chinassamy et al., 2009; Morais and Costa, 2007) indicate that the best concentration of CO2in the gas supplied toC. vulgarisgrowth is about 6%.
TABLE 4.1 Data of Biomass Productivity and CO2Fixation Rate from Microalgae.
Microalgae Strain Biomass
(mg L1d1)
CO2Fixation Rate
(mg L1d1) Reference
Spirulina platensis 145 318 Sydney et al., 2011
Chlorella vulgaris 129 251 Sydney et al., 2011
Synechocystis aquatilis 30 50 Zhang et al., 2001
Anabena sp. 310 1450 Lo´pez et al., 2009
Botryococcus braunii 207 500 Sydney et al., 2011
Dunaliella tertiolecta 143 272 Sydney et al., 2011
Chlorococcum littorale 530 900 Kurano et al., 1996
Aphanothece microscopica Nageli 301 562 Jacob-lopes et al., 2009
Chlorella,Oscillatoria,Oedogonium,Anabaena, Microspora andLyngbya(mixed culture)
131 161 Tsai et al., 2012
76 4. RESPIROMETRIC BALANCE AND CARBON FIXATION OF INDUSTRIALLY IMPORTANT ALGAE
4.2.3.2 Botryococcus braunii
Botryococcusis a colonial microalga that is widespread in fresh and brackish waters of all con- tinents. It is characterized by its slow growth and by containing up to 50% by weight of hydro- carbons.B. brauniiis classified into A, B, and L races, mainly based on the difference between the hydrocarbons produced (Metzger and Largeau, 2005).Banerjee et al. (2002)differentiate the races as follows: Race A produces C25to C31odd-numberedn-alkadienes and alkatrienes; B race pro- duces polymethylated unsaturated triterpenes, calledbotryococcenes(CnH2n–10,n¼30–37); and L race produces a single tetraterpene hydrocarbon C40H78known aslycopadiene.
The cells ofB. brauniiare embedded in a communal extracellular matrix (or “cup”), which is impregnated with oils and cellular exudates (Banerjee et al., 2002).B. brauniiis capable of synthesizing exopolyssaccharides, as reported byCasadevall et al. in 1985. Higher growth and production of EPS, which ranges from 250 g m–3for A and B races to 1 kg m–3 for the L race, occur when nitrate is the nitrogen source instead of urea or ammonium salts (Banerjee et al., 2002). Phosphorus and nitrogen are also important factors in accumulation of hydrocarbons by the microorganism (Jun et al., 2003).
The metabolic energy devoted to produce such large amounts of hydrocarbons makes this species noncompetitive in open mass cultures, since strains not so burdened can grow much faster and soon dominate an outdoor pond culture (Benemann et al., 2002).B. brauniihas been reported to convert 3% of the solar energy to hydrocarbons (Gudin and Chaumont, 1984).
Being synthesized by a photosynthetic organism, hydrocarbons from algae can be burned without contributing to the accumulation of CO2in the atmosphere.
Dayananda et al. (2007)cultivatedBotryococcus brauniistrain SAG 30.81 in shake flasks and obtained a maximum cell concentration of 0.65 g L1under 16:8 light:dark cycle. Experiments with different strains ofB. Braunii indicate that the biomass yield is inversely proportional to lipid accumulation. The maximum biomass yield achieved was 2 g L1(with 40% of lipids) and the lower was 0.2 g L1(with 60% of lipids). Outdoor experiments with this microalga achieved a high biomass yield of 1.8 g L1 but a very low lipid accumulation. It was also showed by Dayananda and collaborators that exopolyssaccharides production byBotryococcus brauniiSAG 30.81 is not affected by light regimen in MBM media, different from lipids and proteins pro- duction. Sydney et al. (2011) carried experiments with this same strain under 12 h light:
dark cycle in 5% CO2enriched air and achieved a high biomass production of 3.11 g L1with 33%
lipids in 15 days. Carbon dioxide fixation rate was calculated as near 500 mg L1day1.B. braunii biomass composition also included 39% proteins, 2.4% carbohydrates, 13% pigments, and 7.5% ash.
Marukami and Ikenouochi (1997)achieved a carbon dioxide fixation greater than 1 gram per liter byBotryococcus brauniicultivated for hydrocarbon accumulation.
4.2.3.3 Spirulina platensis
Spirulina are multicellular ilamentous cyanobacteria actually belonging to two separate genera:SpirulinaandArthrospira.These encompass about 15 species (Habib et al., 2008). This microorganism grows in water, reproduces by binary fission, and can be harvested and processed easily, having significantly high macro- and micronutrient contents. Their main photosynthetic pigments are chlorophyll and phycocyanin. The helical shape of the filaments (or trichomes) is characteristic of the genus and is maintained only in a liquid environment or culture medium.
77
4.2 CARBON DIOXIDE FIXATION BY MICROALGAE
Spirulina is found in soil, marshes, freshwater, brackish water, seawater, and thermal springs. Alkaline, saline water (>30 g/L) with high pH (8.5–11.0) favors good production ofSpirulina, especially where there is a high level of solar radiation. It predominates in higher pH and water conductivity. Like most cyanobacteria,Spirulinais an obligate photoautotroph, i.e., it cannot grow in the dark on media containing only organic carbon compounds.
It reduces carbon dioxide in the light and assimilates mainly nitrates.
Spirulina contains unusually high amounts of protein, between 55% and 70% by dry weight, depending on the source. It has a high amount of polyunsaturated fatty acids (PUFAs), 30% of its 5–6% total lipids, and is a good source of vitamins (B1, B2, B3, B6, B9, B12, C, D, E).Spirulina is a rich source of potassium and also contains calcium, chromium, copper, iron, magnesium, manganese, phosphorus, selenium, sodium, and zinc. These bacteria also contain chlorophyll a and carotenoids.
The optimum pH of theSpirulina sp.culture is between 8.5 and 9.5 (Watanabe et al., 1995).
Cyanobacteria possess a CO2-concentating mechanism that involves active CO2uptake and HCO3 transport. In experiments conducted byMorais and Costa (2007), carbon fixation in terms of biomass bySpirulina platensiswas estimated in 413 mg L1d1, near those achieved bySydney et al. (2011).
4.2.3.4 Dunaliella sp.
Dunaliellais a biflagellate unicellular green alga. Cells are round-shaped and found in brackish environments; it is a motile species and has a high tolerance for salt, temperature, and light.
Motion of cells is important since it facilitates nutrient transport, especially in poor-nutrient waters.Dunaliellaspecies are relatively easy to culture. The cell divides by simple binary fission, and no evidence of cell lysis, encystment, or spore formation is observed (Segovia et al., 2003).
Dunaliellathrives over a wide pH range and expresses a capacity for extremely efficient DIC accumulation, incorporating a capacity to use HCO3 in addition to CO2(Aizawa and Miyachi, 1986;Young et al., 2001).Kishimoto et al. (1994)cultivated aDunaliellastrain for pigment pro- duction with 3% of CO2and achieved a carbon uptake of 313 mg L1day1.Sydney et al. (2011) cultivated aD. tertiolectastrain and achieved a CO2fixation rate of 272 mg L1day1.
Dunaliellais an important microalgae for industrial processes since it produces a wide variety of commercial products (mainly pigments) and the rupture of the cells is very easy.
b-carotene large-scale production facilities are in operation around the world (Hawaii, United States, Australia, Japan).
4.2.3.5 Haematococcus sp.
Haematococcusis a green algae (Chlorophyta), mobile, single-celled, and capable of synthe- sizing and accumulating the pigment astaxanthin in response to environmental conditions, reaching from 1.5% up to 6% by weight astaxanthin (VanessaGhiggi, 2007). The astaxanthin produced by Haematococcus pluvialis is about 70% monoester, 25% diesters, and 5% free (Lorenz and Cysewski, 2000).
These algae, however, have some undesirable characteristics compared to other microalgae grown successfully on a commercial scale. The biggest concern is mainly related to a relatively slow growth rate, allowing easy contamination. Therefore, many studies have sought to improve the low rate of growth of vegetative cells, which is, exceptionally, 1.20 div/day (Gonza´les et al., 2009).
78 4. RESPIROMETRIC BALANCE AND CARBON FIXATION OF INDUSTRIALLY IMPORTANT ALGAE
Alternatively, its mixotrophic (Guerin et al., 2003; Gonza´les et al., 2009) and heterotrophic (Hata et al., 2001) metabolism, using acetate as carbon source, has also been studied and documented; however, these conditions have not been applied to commercial-scale cultures and are not interesting in terms of carbon fixation.