**Abstract**

The search for a renewable source as an alternative to fossil fuels has driven the research on new sources of biomass for biofuels. An alternative source of biomass that has come to prominence is microalgae, photosynthetic micro-organisms capable of capturing atmospheric CO2 and accumulating high levels of lipids in their biomass, making them attractive as a raw material for biodiesel synthesis. Thus, various studies have been conducted in developing different types of photobioreactors for the cultivation of microalgae. Photobioreactors can be divided into two groups: open and closed. Open photobioreactors are more susceptible to contamination and bad weather, reducing biomass productivity. Closed photobioreactors allow greater control against contamination and bad weather and lead to higher rates of biomass production; they are widely used in research to improve new species and processes. Therefore, many configurations of closed photobioreactors have been developed over the years to increase productivity of microalgae biomass.

**Keywords:** raw material for biodiesel production, biomass, microalgae, photobioreactors, biodiesel synthesis

### **1. Introduction**

The continuous rise in global temperature caused by the emission of greenhouse effect gases, mainly carbon dioxide (CO2) mostly due to anthropogenic activities such as the burning of fossil fuels, has led the search for alternative and renewable fuels, such as biofuels. Biofuels present a balance between the CO2 emitted during burning and the subsequent absorption during the formation of biomass, a sustainable cycle.

Biofuels such as bioethanol and biodiesel are alternative substitutes to gasoline and diesel, respectively and have several conventional renewable sources of raw materials. The raw materials for the production of biodiesel are based on oils and fats, which may be of vegetable origin (soybean, peanut, cotton and sunflower oils), animal origin (fish oil, beef tallow, lard and fat from chicken) or residual frying oils and fats, all sources committed to the food chain.

However, another raw material, not competitive with the food chain, has received special attention for presenting many advantages when compared to conventional sources. These are microalgae which are photosynthetic microorganisms capable of accumulating high levels of oil in their cytoplasm which, in turn, can be extracted from biomass and converted into biodiesel.

Microalgae grow at a high speed (less than a week), requiring small territorial areas to develop, presenting high photosynthetic efficiency and, consequently, good absorption of atmospheric CO2. They may use domestic and industrial effluents to develop (not requiring clean water), thus standing out as a promising raw material for the production of biodiesel.

Microalgae cultivations generate high concentration of biomass and, therefore, can be used in the production of biodiesel. The cultivation is generally conducted in illuminated bioreactors, called photobioreactors, which can have different shapes and configurations. Some configurations include columns of bubbles, tubulars, flat plates and open tanks in the shape of "raceways", the most wellknown and widely spread configuration worldwide. However, the design and development of photobioreactors with new configurations and different lighting modes still play a crucial role in optimizing cell growth and increasing biomass formation.

In this context, this chapter will present the reasons for the use of microalgae as sources of biodiesel, the main photobioreactors used for the production of microalgae biomass and a brief description of the reaction synthesis of biodiesel.

### **2. Biodiesel sources**

The average temperature of the planet has been increasing year by year and causing serious climatic imbalances, such as changes in the rain pattern and melting of the glaciers, resulting in losses in local agriculture and rising sea levels. Much of this rise in global temperature is due to the anthropogenic emission of carbon dioxide (CO2) into the atmosphere, mainly from the burning of fossil fuels [1].

According to IPEA [2], the anthropogenic contribution to the emission of CO2 in the atmosphere is mainly related to energy purposes, in which transportation is responsible for 67% of the emissions as a result of carbon oxidation during the combustion of the most varied fuels.

For energy purposes, the contribution of fossil fuels is higher than 80% of the anthropogenic emission of CO2 into the Earth's atmosphere, corresponding to approximately 35 giga tons of CO2 emitted in 2015 [2, 3].

Although technically there is a diversified energy matrix, including hydroelectric, thermoelectric, wind, solar and nuclear energy sources, the planet is highly dependent on thermoelectric plants, which burn coal (mainly) and natural gas, fossil fuels that are of finite origin and unsustainable, basically transferring terrestrial carbon to the atmosphere in its majority in the form of CO2 [3].

In addition to the massive presence of fossil fuels in the generation of electric energy, they also dominate the transportation sector, overloaded with vehicles powered by combustion, with exponents busses and trucks fueled by diesel, a fossil fuel that highly contributes to emissions of CO2 [4].

Biodiesel is the biofuel that can replace diesel and can be produced from vegetable oils (soy, palm, sunflower, cotton, peanuts and others), oils from microorganisms (cyanobacteria and microalgae), animal fat and the frying oil reuse [5, 6]. The current biofuel projections are based on sources that are also food commodities and resources suitable for conventional agriculture. Thus, the biodiesel production involves the use of lands already used to food production [7].

**385**

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production*

In addition, the use of microalgae biomass for the production of biodiesel has been gaining prominence, as this type of microorganism has high growth rates even on smaller areas of cultivation, microalgae biomass can generate high amounts of biomass and oils extracted from its dry biomass compared to vegetable oilseeds as seen in **Figure 1**. The cultivation of microalgae promotes the biofixation of CO2, allows the use of wastewater to treat industrial and domestic effluents and does not

*Different sources of biodiesel and their respective production efficiencies and territorial demand.*

Microalgae are single-celled or multicellular organisms with microscopic dimensions, capable of photosynthesis due to the presence of chlorophyll [10]. Microalgae mainly inhabit aquatic environments such as: lakes, rivers and oceans, although they can live in solid humid environments such as wet soil and rocks. They have about 50% carbon in their composition, based on their dry biomass, as a result of

In the sea, microalgae constitute phytoplankton and are responsible for approximately 90% of photosynthetic activity in the oceans which can lead to an annual fixation of 45–50 Gton of carbon in the oceans [13]. It is estimated that 1.8 kg of absorbed CO2, during photosynthesis, generates 1 kg of microalgal biomass [14, 15]. The photosynthesis reaction that represents this conversion of CO2 into biomass can

The biological composition of microalgae varies widely between species and is strongly influenced by environmental factors such as temperature, lighting, photoperiod, pH of the culture medium, mineral nutrients, CO2 supplement, etc. [15]. It is known that the concentration of nitrate in the microalgae culture medium significantly influences the production of lipids by the cell, especially when available in

low concentrations inducing its accumulation inside the cell [16].

6 6 *CO H O C H O O* 2 2 6 12 6 2 +→ + 6 (1)

*DOI: http://dx.doi.org/10.5772/intechopen.93547*

compete with agricultural food production [8, 9].

be represented in a simplified way by Eq. (1).

**3. Microalgae**

**Figure 1.**

photosynthesis [11, 12].

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production DOI: http://dx.doi.org/10.5772/intechopen.93547*

#### **Figure 1.**

*Biotechnological Applications of Biomass*

for the production of biodiesel.

formation.

**2. Biodiesel sources**

combustion of the most varied fuels.

approximately 35 giga tons of CO2 emitted in 2015 [2, 3].

fuel that highly contributes to emissions of CO2 [4].

carbon to the atmosphere in its majority in the form of CO2 [3].

extracted from biomass and converted into biodiesel.

However, another raw material, not competitive with the food chain, has received special attention for presenting many advantages when compared to conventional sources. These are microalgae which are photosynthetic microorganisms capable of accumulating high levels of oil in their cytoplasm which, in turn, can be

Microalgae grow at a high speed (less than a week), requiring small territorial areas to develop, presenting high photosynthetic efficiency and, consequently, good absorption of atmospheric CO2. They may use domestic and industrial effluents to develop (not requiring clean water), thus standing out as a promising raw material

Microalgae cultivations generate high concentration of biomass and, therefore, can be used in the production of biodiesel. The cultivation is generally conducted in illuminated bioreactors, called photobioreactors, which can have different shapes and configurations. Some configurations include columns of bubbles, tubulars, flat plates and open tanks in the shape of "raceways", the most wellknown and widely spread configuration worldwide. However, the design and development of photobioreactors with new configurations and different lighting modes still play a crucial role in optimizing cell growth and increasing biomass

In this context, this chapter will present the reasons for the use of microalgae as sources of biodiesel, the main photobioreactors used for the production of microal-

The average temperature of the planet has been increasing year by year and causing serious climatic imbalances, such as changes in the rain pattern and melting of the glaciers, resulting in losses in local agriculture and rising sea levels. Much of this rise in global temperature is due to the anthropogenic emission of carbon dioxide (CO2) into the atmosphere, mainly from the burning of fossil fuels [1]. According to IPEA [2], the anthropogenic contribution to the emission of CO2 in the atmosphere is mainly related to energy purposes, in which transportation is responsible for 67% of the emissions as a result of carbon oxidation during the

For energy purposes, the contribution of fossil fuels is higher than 80% of the anthropogenic emission of CO2 into the Earth's atmosphere, corresponding to

Although technically there is a diversified energy matrix, including hydroelectric, thermoelectric, wind, solar and nuclear energy sources, the planet is highly dependent on thermoelectric plants, which burn coal (mainly) and natural gas, fossil fuels that are of finite origin and unsustainable, basically transferring terrestrial

In addition to the massive presence of fossil fuels in the generation of electric energy, they also dominate the transportation sector, overloaded with vehicles powered by combustion, with exponents busses and trucks fueled by diesel, a fossil

Biodiesel is the biofuel that can replace diesel and can be produced from vegetable oils (soy, palm, sunflower, cotton, peanuts and others), oils from microorganisms (cyanobacteria and microalgae), animal fat and the frying oil reuse [5, 6]. The current biofuel projections are based on sources that are also food commodities and resources suitable for conventional agriculture. Thus, the biodiesel production involves the use of lands already used to food production [7].

gae biomass and a brief description of the reaction synthesis of biodiesel.

**384**

*Different sources of biodiesel and their respective production efficiencies and territorial demand.*

In addition, the use of microalgae biomass for the production of biodiesel has been gaining prominence, as this type of microorganism has high growth rates even on smaller areas of cultivation, microalgae biomass can generate high amounts of biomass and oils extracted from its dry biomass compared to vegetable oilseeds as seen in **Figure 1**. The cultivation of microalgae promotes the biofixation of CO2, allows the use of wastewater to treat industrial and domestic effluents and does not compete with agricultural food production [8, 9].

#### **3. Microalgae**

Microalgae are single-celled or multicellular organisms with microscopic dimensions, capable of photosynthesis due to the presence of chlorophyll [10]. Microalgae mainly inhabit aquatic environments such as: lakes, rivers and oceans, although they can live in solid humid environments such as wet soil and rocks. They have about 50% carbon in their composition, based on their dry biomass, as a result of photosynthesis [11, 12].

In the sea, microalgae constitute phytoplankton and are responsible for approximately 90% of photosynthetic activity in the oceans which can lead to an annual fixation of 45–50 Gton of carbon in the oceans [13]. It is estimated that 1.8 kg of absorbed CO2, during photosynthesis, generates 1 kg of microalgal biomass [14, 15]. The photosynthesis reaction that represents this conversion of CO2 into biomass can be represented in a simplified way by Eq. (1).

$$6\,\text{CO}\_2 + 6\,H\_2\text{O} \to \text{C}\_6\text{H}\_{12}\text{O}\_6 + 6\,\text{O}\_2\tag{1}$$

The biological composition of microalgae varies widely between species and is strongly influenced by environmental factors such as temperature, lighting, photoperiod, pH of the culture medium, mineral nutrients, CO2 supplement, etc. [15]. It is known that the concentration of nitrate in the microalgae culture medium significantly influences the production of lipids by the cell, especially when available in low concentrations inducing its accumulation inside the cell [16].

Microalgae synthesize lipids from the carbon source, whether inorganic such as CO2 or organic (glucose, acetate, etc.). The components and levels of lipids in microalgal cells vary from species to species, being basically divided into neutral lipids (triglycerides and cholesterol) and polar lipids, such as phospholipids. Neutral lipids, such as triglycerides, are considered as the main material to produce biodiesel [17].

The oil content commonly found in microalgae is in the range of 15 to 50%, but these levels can be increased according to the manipulation of the crops aiming at the accumulation of lipids. In addition, microalgae of the Chlorella genus require less cultivation time and because of its high cell productivity and potential lipid extraction, microalgae can be widely used as a raw material in the production of biodiesel. The conversion to biodiesel occurs by the transesterification reaction using alcohol in the presence of acid or base as catalysts [12, 18].

Another group of substances abundant in the composition of most microalgae species are proteins. The high amount of proteins present in the cell composition in the most varied species of microalgae gives these microorganisms their recognition as an unconventional source of proteins. Proteins extracted from microalgal biomass for human nutrition are currently commercialized in various forms, such as tablets, capsules and liquids, which can be incorporated into pasta, cakes, sweets and drinks [19].

Along with proteins, the considerable composition of numerous essential vitamins in microalgae stands out: vitamin A, B1, B2, B6, B12, C, E, biotin, folic acid, among others, in levels that vary between species and according to cultivation techniques, as well as nucleic acids, which comprise from 1 to 6% of the cellular


**387**

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production*

composition of microalgae and play a vital role in cell growth and repair. Due to the presence of phosphorus and nitrogen in large amounts in nucleic acids, microalgal

Carbohydrates, on the other hand, comprise the final product of photosynthesis in microalgae and can vary from 4 to 64% of their cell composition (**Table 1**) [10]. These are synthesized by fixing CO2 in the form of glucose, disaccharides and starch

In terms of microalgal biomass, carbohydrates can be extracted and converted into ethanol by the anaerobic fermentation process using yeasts (*Saccharomyces cerevisiae*, for example), or undergo degradation of organic matter in anaerobic conditions leading to the formation of methane and carbon dioxide [10].

All microalgae have one or more types of chlorophyll, among the four existing types, chlorophyll a, b, c and d, with chlorophyll being the most important pigment for photosynthesis playing a major role in the arrangement of photosystems to capture light energy. The other chlorophylls play secondary roles, contributing to increasing the total light absorbed by the microalgae. Commercially, chlorophylls

Photobioreactors are equipment designed to allow the use of light energy, whether natural or artificial, by the cells of the micro-organism present in it, with the purpose of providing the desired bioproduct. In the case of microalgae, it is desired that the photobioreactor helps promoting cell growth, and for this purpose several types of photobioreactors have been designed, such as open tanks, bubble

According to Huang et al. [22], photobioreactors can be confined and the protective covered structures if built with transparent materials allow natural li lighting if the artificial lighting is not an option. They may also be available outdoors, subjected to weather conditions which randomly interfere in the development of the crop.

To minimize the interference associated with external factors, such as variations

in the climate, impaired light by cloudy days or long rainy periods and possible contamination by insects, the photobioreactors that allow this direct contact of the culture medium with the external environment (called "open" photobioreactors), can be replaced by "closed" photobioreactors, in which the culture medium has no contact with the external environment, and if provided by artificial lighting they do not suffer from changes in radiance, ensuring greater control and predictability [23]. Among the open photobioreactors, we highlight the open ponds or ponds, known by the English term "Open ponds", which are cultivation containers made of plastic materials (PVC, for example), fiberglass or cement, and should contain smooth internal surfaces in order to reduce likely damages from friction and guarantee ease cleaning. In the construction of the tanks, the depth can vary from 10 to 50 cm approximately, so that it allows the diffusion of carbon dioxide from the atmosphere and the penetration of sunlight as well as allowing aeration by bubbling

The circular ponds, or circular tanks, reach an average of 1000 m<sup>2</sup>

provided by large rotational arms that can extend up to 45 m in diameter, reaching yields that can vary from 1.5 to 16.5 g m−2 d−1 of dry biomass, while the "raceway pond" type gets its name from the format that resembles a racing circuit. It is the most used system at both pilot and commercial scale due to its ease of operation [24]. Productivities can vary from 0.19 to 23.5 g m−2 d−1 of dry biomass [10].

, 30cm deep,

columns, flat and tubular plates (conical or helical), among others [21].

*DOI: http://dx.doi.org/10.5772/intechopen.93547*

biomass can also be used as a fertilizer [10].

are used mainly in the dye industry [20].

air in the culture medium [12, 20].

**Figure 2** shows open ponds photobioreactors.

**4. Photobioreactors**

as the primary source of energy for the cell [20].

#### **Table 1.** *Composition of microalgae.*

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production DOI: http://dx.doi.org/10.5772/intechopen.93547*

composition of microalgae and play a vital role in cell growth and repair. Due to the presence of phosphorus and nitrogen in large amounts in nucleic acids, microalgal biomass can also be used as a fertilizer [10].

Carbohydrates, on the other hand, comprise the final product of photosynthesis in microalgae and can vary from 4 to 64% of their cell composition (**Table 1**) [10]. These are synthesized by fixing CO2 in the form of glucose, disaccharides and starch as the primary source of energy for the cell [20].

In terms of microalgal biomass, carbohydrates can be extracted and converted into ethanol by the anaerobic fermentation process using yeasts (*Saccharomyces cerevisiae*, for example), or undergo degradation of organic matter in anaerobic conditions leading to the formation of methane and carbon dioxide [10].

All microalgae have one or more types of chlorophyll, among the four existing types, chlorophyll a, b, c and d, with chlorophyll being the most important pigment for photosynthesis playing a major role in the arrangement of photosystems to capture light energy. The other chlorophylls play secondary roles, contributing to increasing the total light absorbed by the microalgae. Commercially, chlorophylls are used mainly in the dye industry [20].

## **4. Photobioreactors**

*Biotechnological Applications of Biomass*

biodiesel [17].

and drinks [19].

Microalgae synthesize lipids from the carbon source, whether inorganic such as CO2 or organic (glucose, acetate, etc.). The components and levels of lipids in microalgal cells vary from species to species, being basically divided into neutral lipids (triglycerides and cholesterol) and polar lipids, such as phospholipids. Neutral lipids, such as triglycerides, are considered as the main material to produce

The oil content commonly found in microalgae is in the range of 15 to 50%, but these levels can be increased according to the manipulation of the crops aiming at the accumulation of lipids. In addition, microalgae of the Chlorella genus require less cultivation time and because of its high cell productivity and potential lipid extraction, microalgae can be widely used as a raw material in the production of biodiesel. The conversion to biodiesel occurs by the transesterification reaction

Another group of substances abundant in the composition of most microalgae species are proteins. The high amount of proteins present in the cell composition in the most varied species of microalgae gives these microorganisms their recognition as an unconventional source of proteins. Proteins extracted from microalgal biomass for human nutrition are currently commercialized in various forms, such as tablets, capsules and liquids, which can be incorporated into pasta, cakes, sweets

Along with proteins, the considerable composition of numerous essential vitamins in microalgae stands out: vitamin A, B1, B2, B6, B12, C, E, biotin, folic acid, among others, in levels that vary between species and according to cultivation techniques, as well as nucleic acids, which comprise from 1 to 6% of the cellular

**Species Proteins Carbohydrates Lipids Nucleic acids** *Scenedesmus obliquus* 50–56 10–17 12–14 3–6 *Scenedesmus quadricauda* 47 — 1.9 — *Scenedesmus dimorphus* 8–18 21–52 16–40 — *Chlamydomonas rheinhardii* 48 17 21 — *Chlorella vulgaris* 51–58 12–17 14–22 4–5 *Chlorella pyrenoidosa* 57 26 2 — *Chlorella minutissima* 35.5 — 31 — *Spirogyra sp.* 6–20 33–64 11–21 — *Dunaliella bioculata* 49 4 8 — *Dunaliella salina* 57 32 6 — *Euglena gracilis* 39–61 14–18 14–20 — *Prymnesium parvum* 28–45 25–33 22–38 1–2 *Tetraselmis maculata* 52 15 3 — *Porphyridium cruentum* 28–39 40–57 9–14 — *Spirulina platensis* 46–63 8–14 4–9 2–5 *Spirulina máxima* 60–71 13–16 6–7 3–4.5 *Synechoccus sp.* 63 15 11 5 *Anabaena cylindrica* 43–56 25–30 4–7 —

using alcohol in the presence of acid or base as catalysts [12, 18].

**386**

*Fonte: [10].*

*Composition of microalgae.*

**Table 1.**

Photobioreactors are equipment designed to allow the use of light energy, whether natural or artificial, by the cells of the micro-organism present in it, with the purpose of providing the desired bioproduct. In the case of microalgae, it is desired that the photobioreactor helps promoting cell growth, and for this purpose several types of photobioreactors have been designed, such as open tanks, bubble columns, flat and tubular plates (conical or helical), among others [21].

According to Huang et al. [22], photobioreactors can be confined and the protective covered structures if built with transparent materials allow natural li lighting if the artificial lighting is not an option. They may also be available outdoors, subjected to weather conditions which randomly interfere in the development of the crop.

To minimize the interference associated with external factors, such as variations in the climate, impaired light by cloudy days or long rainy periods and possible contamination by insects, the photobioreactors that allow this direct contact of the culture medium with the external environment (called "open" photobioreactors), can be replaced by "closed" photobioreactors, in which the culture medium has no contact with the external environment, and if provided by artificial lighting they do not suffer from changes in radiance, ensuring greater control and predictability [23].

Among the open photobioreactors, we highlight the open ponds or ponds, known by the English term "Open ponds", which are cultivation containers made of plastic materials (PVC, for example), fiberglass or cement, and should contain smooth internal surfaces in order to reduce likely damages from friction and guarantee ease cleaning. In the construction of the tanks, the depth can vary from 10 to 50 cm approximately, so that it allows the diffusion of carbon dioxide from the atmosphere and the penetration of sunlight as well as allowing aeration by bubbling air in the culture medium [12, 20].

The circular ponds, or circular tanks, reach an average of 1000 m<sup>2</sup> , 30cm deep, provided by large rotational arms that can extend up to 45 m in diameter, reaching yields that can vary from 1.5 to 16.5 g m−2 d−1 of dry biomass, while the "raceway pond" type gets its name from the format that resembles a racing circuit. It is the most used system at both pilot and commercial scale due to its ease of operation [24]. Productivities can vary from 0.19 to 23.5 g m−2 d−1 of dry biomass [10]. **Figure 2** shows open ponds photobioreactors.

**Figure 2.** *Open pounds photobioreactors. Raceway pond (A); circular pond (B). Source: authors.*

According to Huang [22], in the class of "closed" photobioreactors, the bubble column type is widely used by both researchers on a laboratory scale and commercial purposes due to its versatility of operation, handling and construction, as they are basically cylinders with a high height/diameter ratio and made of transparent materials such as glass and mainly polymers (acrylics and polyvinyl chlorides for example).

According to Carvalho [21], the bubble column photobioreactor has a unique configuration with an air dispersion system in its internal structure, which allows controlling the size and release of air bubbles promoting a random pneumatic agitation of the medium of cultivation.

In order to promote an oriented movement of the liquid inside the cylinder, photobioreactors of "air-lift" models can be used, which are equipped with devices called "draft tube". These devices that allow the distinction of two regions: one that it presents an upward flow of medium where air bubbles are released (Riser), and another of downward liquid flow (Downcomer) [25].

Flat plate photobioreactors are characterized by cobblestones of glass, plastic or other material that allows the passage of solar radiation. The air, enriched with carbon dioxide necessary for the growth of biomass is injected by the base, and promotes turbulence so that all cells are affected by solar radiation [26].

The arrangement of flat plate photobioreactors can vary vertically, horizontally or at an angle. They have the advantage of having a large surface area, which allows better use of the received radiation favoring the photosynthetic activity of microalgae [10]. **Figure 3** shows bubbles column, air-lift and flat plate photobioreactors.

Tubular photobioreactors are the most popular; they have a high ratio of exposed surface to volume, high efficiency of CO2 usage and sunlight. They consist of sets of transparent tubes, usually glass or plastic, whose diameters are less than 0.1 m, since light is required in the photosynthesis process. Such photobioreactors can be designed in several formats (**Figure 3**): serpentine, inclined, spiral, coils and in parallel [10, 21, 24, 27].

The photobioreactors of the agitated tank type, on the other hand, have a structure composed of cylindrical tubes of glass or material that allows the passage of natural or artificial light and occupy small areas. The reactor may have mechanical or pneumatic agitation systems (air distributors), or even both. They are known as mixing bioreactors, responsible for more than 90% of applications in the fermentation industry [10, 21]. **Figure 4** shows tubular and agitated tank type photobioreactors.

**389**

**Figure 4.**

*Source: authors.*

**Figure 3.**

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production*

*Bubbles column (A), air-lift (B and C) and flat plate (D) photobioreactors. Source: authors.*

In the municipality of Almeria, Spain, the microalgae *Scenedesmus almeriensis* is grown commercially in a 3000 L vertical tubular photobioreactor, owned by the CAJAMAR Foundation. The tubes are made of transparent polymeric material 90 mm in diameter and 400 m long, connected to a 3.5 m high bubble column [28].

*Tubular photobioreactors: serpentine (A), inclined (B) and coil (D). Agitated tank photobioreactor (C).* 

*DOI: http://dx.doi.org/10.5772/intechopen.93547*

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production DOI: http://dx.doi.org/10.5772/intechopen.93547*

#### **Figure 4.**

*Biotechnological Applications of Biomass*

According to Huang [22], in the class of "closed" photobioreactors, the bubble column type is widely used by both researchers on a laboratory scale and commercial purposes due to its versatility of operation, handling and construction, as they are basically cylinders with a high height/diameter ratio and made of transparent materials such as glass and mainly polymers (acrylics and polyvinyl chlorides for

*Open pounds photobioreactors. Raceway pond (A); circular pond (B). Source: authors.*

According to Carvalho [21], the bubble column photobioreactor has a unique configuration with an air dispersion system in its internal structure, which allows controlling the size and release of air bubbles promoting a random pneumatic

In order to promote an oriented movement of the liquid inside the cylinder, photobioreactors of "air-lift" models can be used, which are equipped with devices called "draft tube". These devices that allow the distinction of two regions: one that it presents an upward flow of medium where air bubbles are released (Riser), and

Flat plate photobioreactors are characterized by cobblestones of glass, plastic or other material that allows the passage of solar radiation. The air, enriched with carbon dioxide necessary for the growth of biomass is injected by the base, and

The arrangement of flat plate photobioreactors can vary vertically, horizontally

promotes turbulence so that all cells are affected by solar radiation [26].

or at an angle. They have the advantage of having a large surface area, which allows better use of the received radiation favoring the photosynthetic activity of microalgae [10]. **Figure 3** shows bubbles column, air-lift and flat plate

Tubular photobioreactors are the most popular; they have a high ratio of exposed surface to volume, high efficiency of CO2 usage and sunlight. They consist of sets of transparent tubes, usually glass or plastic, whose diameters are less than 0.1 m, since light is required in the photosynthesis process. Such photobioreactors can be designed in several formats (**Figure 3**): serpentine, inclined, spiral, coils and

The photobioreactors of the agitated tank type, on the other hand, have a structure composed of cylindrical tubes of glass or material that allows the passage of natural or artificial light and occupy small areas. The reactor may have mechanical or pneumatic agitation systems (air distributors), or even both. They are known as mixing bioreactors, responsible for more than 90% of applications in the fermentation industry [10, 21]. **Figure 4** shows tubular and agitated tank type

**388**

example).

**Figure 2.**

photobioreactors.

photobioreactors.

in parallel [10, 21, 24, 27].

agitation of the medium of cultivation.

another of downward liquid flow (Downcomer) [25].

*Tubular photobioreactors: serpentine (A), inclined (B) and coil (D). Agitated tank photobioreactor (C). Source: authors.*

In the municipality of Almeria, Spain, the microalgae *Scenedesmus almeriensis* is grown commercially in a 3000 L vertical tubular photobioreactor, owned by the CAJAMAR Foundation. The tubes are made of transparent polymeric material 90 mm in diameter and 400 m long, connected to a 3.5 m high bubble column [28].

**Figure 5.**

*Tubular arrangements of photobioreactors: (A) serpentine tubular reactor; (B) serpentine tubular reactor with mirror at its base to increase the efficiency of light capture by reflecting the incident light. Source: authors.*

Many researchers have been designing and developing new configurations or strategies for the arrangement of photobioreactors to stimulate cell growth and the accumulation of metabolites in microalgae. As an example, the photobioreactor developed by Liao et al. [29] (**Figure 5A**), who horizontally disposed a serpentine tubular reactor, with regions of the tube that do not allow the passage of light, creating light and dark regions in photobioreactor to induce the accumulation of lipids during cell growth. A similar strategy was devised and developed by Ilus and Abu-Goshi [30] (**Figure 5B**), with a similar photobioreactor, plus a mirror at its base to increase the efficiency of light capture by reflecting the incident light.

An unconventional photobioreactor was developed by Huang et al. [31], named as a rotating float consisting of bottles in hexagonal arrangement and used as paddles of a raceway pond for the cultivation of microalgae *Dunaliella tertiolecta*. Another interesting photobioreactor has been proposed by Pruvost et al. [32], in which a flat plate photobioreactor is integrated into a building, with the purpose of producing microalgae biomass. This photobioreactor takes advantage of the exhaust gases from the chimney of a commercial building, thereby increasing the production of biomass during cultivation. In addition, it provides thermal comfort to the building in which it is integrated, as it reduces the internal temperature of the place due to the absorption of sunlight by microalgae while growing in the photobioreactor, which is located next to the building. **Figure 6** shows flat plate photobioreactor integrated into a building and rotating float photobioreactor.

It seems that microalgal cultures can be conducted under the most diverse configurations of photobioreactors from open to closed tanks, bubble columns, airlift, tubular

#### **Figure 6.**

*Flat plate photobioreactor integrated into a building (A) and rotating float photobioreactor (B). Source: authors.*

**391**

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production*

temperature, CO2 and nutrients

Bubble column The marine microalgae *Chlorella minutissima* was grown in a 20 L

Raceway The cultivation of *Nannochloropsis* microalgae under nutritional stress

Tubular The microalgae *Chlorella vulgaris* was cultivated in two stages as a

Placas planas The cultivation of the microalgae *Chlorella pyrenoidosa* was evaluated

(vertical, horizontal, inclined or spiral), combinations of more than one photobioreactor or even allied to architectural projects like the plate-type photobioreactor built next to an industrial building that rejects CO2 in its chimney. **Table 2** summarizes some works carried out with several microalgae in the most common photobioreactors.

strategy to accumulate lipids for the biodiesel production

**Configuration Summary Reference**

photobioreactor for the study of several controlled factors, such as

conditions in an open tank in order to accumulate lipids to produce

under different lighting gradients throughout the photobioreactor.

Loures et al. [33]

Perrier et al. [34]

Chen et al. [35]

Huang et al. [36]

After the microalgae cultivation period in the photobioreactors, the microalgal biomass must be harvested. This step is considered by many to be the key stage for the industrial production of biofuels from microalgae oils because they grow in very diluted cultures. The energy consumption is high due to the processing of large

The main methods of harvesting microalgae are: flocculation, electroflocculation, biofloculation, gravimetric sedimentation, flotation, filtration and centrifugation [38]. The flocculation process involves the addition of chemicals capable of inducing the aggregation of microalgae cells either by neutralization, inversion of the electrical charges on the cell walls, or by the formation of bonds between the microalgae. It allows biomass recovery with lower economic costs when compared to the

The main flocculating agents used for harvesting microalgae are salts such as FeCl3, Al2(SO4)3 and Fe2(SO4)3, polymers (chitosan) and bases such as NaOH or

The electrofloculation process, not widely disseminated, is based on the characteristic of microalgae to be electronegatively charged. Thus, by adapting a culture medium with the insertion of electrodes and allowing the passage of electrical current, it is possible to separate the microalgae cells of culture medium [38]. According to Barros et al. [38], in the biofloculation process, the flocculating agent is a microorganism, such as bacteria of the genus *Flavobacterium*, *Terrimonas* and *Sphingobacterium*, or fungi (*Rhizopus oryzae*, *Penicillium expansum*, *Aspergilus* and *Mucor circinelloides*), forming aggregates with the microalgal cells and favoring

Biofloculation can eliminate the need for chemical flocculating agents, although for some cases it is necessary to add CaCl2 to guarantee the efficiency of the process [38]. The flotation process can be defined as the "reverse" of sedimentation, where air bubbles promote the ascension of the cells of the culture by separating it from the liquid medium. However, cultures of marine microalgae are unlikely to undergo

quantities of liquid which are needed to guarantee a feasible harvesting [37].

*DOI: http://dx.doi.org/10.5772/intechopen.93547*

biodiesel.

*Use of traditional photobioreactors in research on microalgae cultivations.*

**5. Harvesting biomass**

**Table 2.**

centrifugation process [37].

KOH, due to the change in pH [38].

the microalgal cells gravimetric settling.

flotation, due to the high salinity of the medium [38].

*Microalgae Cultivation in Photobioreactors Aiming at Biodiesel Production DOI: http://dx.doi.org/10.5772/intechopen.93547*


**Table 2.**

*Biotechnological Applications of Biomass*

**Figure 5.**

Many researchers have been designing and developing new configurations or strategies for the arrangement of photobioreactors to stimulate cell growth and the accumulation of metabolites in microalgae. As an example, the photobioreactor developed by Liao et al. [29] (**Figure 5A**), who horizontally disposed a serpentine tubular reactor, with regions of the tube that do not allow the passage of light, creating light and dark regions in photobioreactor to induce the accumulation of lipids during cell growth. A similar strategy was devised and developed by Ilus and Abu-Goshi [30] (**Figure 5B**), with a similar photobioreactor, plus a mirror at its base to increase the efficiency of light capture by reflecting the incident light.

*Tubular arrangements of photobioreactors: (A) serpentine tubular reactor; (B) serpentine tubular reactor with mirror at its base to increase the efficiency of light capture by reflecting the incident light. Source: authors.*

An unconventional photobioreactor was developed by Huang et al. [31], named

It seems that microalgal cultures can be conducted under the most diverse configurations of photobioreactors from open to closed tanks, bubble columns, airlift, tubular

*Flat plate photobioreactor integrated into a building (A) and rotating float photobioreactor (B). Source:* 

as a rotating float consisting of bottles in hexagonal arrangement and used as paddles of a raceway pond for the cultivation of microalgae *Dunaliella tertiolecta*. Another interesting photobioreactor has been proposed by Pruvost et al. [32], in which a flat plate photobioreactor is integrated into a building, with the purpose of producing microalgae biomass. This photobioreactor takes advantage of the exhaust gases from the chimney of a commercial building, thereby increasing the production of biomass during cultivation. In addition, it provides thermal comfort to the building in which it is integrated, as it reduces the internal temperature of the place due to the absorption of sunlight by microalgae while growing in the photobioreactor, which is located next to the building. **Figure 6** shows flat plate photobioreactor

integrated into a building and rotating float photobioreactor.

**390**

**Figure 6.**

*authors.*

*Use of traditional photobioreactors in research on microalgae cultivations.*

(vertical, horizontal, inclined or spiral), combinations of more than one photobioreactor or even allied to architectural projects like the plate-type photobioreactor built next to an industrial building that rejects CO2 in its chimney. **Table 2** summarizes some works carried out with several microalgae in the most common photobioreactors.

#### **5. Harvesting biomass**

After the microalgae cultivation period in the photobioreactors, the microalgal biomass must be harvested. This step is considered by many to be the key stage for the industrial production of biofuels from microalgae oils because they grow in very diluted cultures. The energy consumption is high due to the processing of large quantities of liquid which are needed to guarantee a feasible harvesting [37].

The main methods of harvesting microalgae are: flocculation, electroflocculation, biofloculation, gravimetric sedimentation, flotation, filtration and centrifugation [38].

The flocculation process involves the addition of chemicals capable of inducing the aggregation of microalgae cells either by neutralization, inversion of the electrical charges on the cell walls, or by the formation of bonds between the microalgae. It allows biomass recovery with lower economic costs when compared to the centrifugation process [37].

The main flocculating agents used for harvesting microalgae are salts such as FeCl3, Al2(SO4)3 and Fe2(SO4)3, polymers (chitosan) and bases such as NaOH or KOH, due to the change in pH [38].

The electrofloculation process, not widely disseminated, is based on the characteristic of microalgae to be electronegatively charged. Thus, by adapting a culture medium with the insertion of electrodes and allowing the passage of electrical current, it is possible to separate the microalgae cells of culture medium [38].

According to Barros et al. [38], in the biofloculation process, the flocculating agent is a microorganism, such as bacteria of the genus *Flavobacterium*, *Terrimonas* and *Sphingobacterium*, or fungi (*Rhizopus oryzae*, *Penicillium expansum*, *Aspergilus* and *Mucor circinelloides*), forming aggregates with the microalgal cells and favoring the microalgal cells gravimetric settling.

Biofloculation can eliminate the need for chemical flocculating agents, although for some cases it is necessary to add CaCl2 to guarantee the efficiency of the process [38].

The flotation process can be defined as the "reverse" of sedimentation, where air bubbles promote the ascension of the cells of the culture by separating it from the liquid medium. However, cultures of marine microalgae are unlikely to undergo flotation, due to the high salinity of the medium [38].

The filtration method consists of passing the culture medium through a small pore membrane that retains the microalgal cells and allows only the passage of the liquid medium [37].

According to Lourenço [20], in the filtration of small volumes, a filtration apparatus such as a Kitasato flask or similar is applicable, however, on a large-scale cultivation, the separation of huge volumes of microalgae and biomass processing is only feasible if the species have large or filamentous cells, which are easily retained in the filter; a short process time.

Regarding the harvest time, gravimetric sedimentation is a slow and simple process which involves the separation of microalgae cells from the culture medium by the action of gravitational force. The downside of this process is the low efficiency [20].

On the other hand, centrifugation, cell sedimentation equipment that acts by the action of centrifugal force, is a fast process of biomass recovery, however, it requires high energy consumption. By this technique, biomass is concentrated without the addition of chemicals, preserving its original characteristics [20].
