to external photic zone at higher gas flow rate (Janssen et al., 2003). Photosynthetic efficiency can be increased by increasing the gas flow rate (0.05 m/s), leading to shorter light and dark cycles.Degen et al., 2001used a bubble column photobioreactor to improve light utilization efficiency of the strainChlorella vulgaristhrough a flashing-light effect in batch mode opera- tion and achieved 1.7 times higher productivity of biomass (Degen et al., 2001).
8.5.2.4 Flat Panel Photobioreactors
It has been reported that with flat panel/plate photobioreactors, high photosynthetic effi- ciencies can be achieved (Hu et al., 1996;Richmond, 2000). Accumulation of dissolved oxygen concentrations in flat plate photobioreactors is relatively low compared to horizontal tubular photobioreactors.Milner’s (1953) work paved the way to the use of flat culture vessels for cultivation of algae. Flat panel photo bioreactors were used extensively for mass cultivation of different algae (Tredici and Materassi, 1992; Hu et al., 1996;Zhang et al., 2002;Hoekema., 2002). Lack of temperature control and gas engagement zones are some of the inherent dis- advantages observed with this type of photobioreactor.
8.5.2.5 Helical-Type Photobioreactors
A coiled transparent and flexible tube of small diameter with separate or attached degassing unit is the basis for the helical type of bioreactor. A centrifugal pump is used to drive the culture through a long tube to the degassing unit. CO2gas mixture and feed can be circulated from either direction, but injection from the bottom gives better photosynthetic efficiency (Morita et al., 2001). A degasser facilitates removal of photosynthetically produced oxygen and residual gas of the injected gas stream. This system facilitates better CO2transfer from gas phase to liquid phase due to a large CO2absorbing pathway (Watanabe et al., 1995).
The energy required by the centrifugal pump in recirculating the culture and associated shear stress limits this reactor’s commercial use (Briassoulis et al., 2010). Fouling on the inside of the reactor is another disadvantage of this system.
8.5.2.6 Stirred-Tank Photobioreactors
Stirred-tank photobioreactors are the conventional reactor setup in which agitation is provided mechanically with the help of impellers or baffles by providing illumination exter- nally. CO2-enriched air is bubbled at the bottom to provide a carbon source for algae growth (Petkov, 2000; Demessie and Bekele, 2003).Protoceratium reticulatumgrowth studied in 2 L and 15 L stirred photobioreactors equipped with internal spin filters showed average biomass cell productivity 3.7 times higher than that of the static cultures (Camacho et al., 2011). Low surface-area-to-volume ratio, which in turn decreases light-harvesting efficiency, is the inher- ent disadvantage of this system. Low surface-area-to-volume ratio and high shear stress imposed due to mechanical agitation limits this reactor’s use in CO2sequestration (Demessie and Bekele, 2003).
processes for converting the algae biomass to biodiesel are crucial and involve a series of se- quentially integrated post-harvesting steps: harvesting, drying, cell disruption, extraction, and transesterification, followed by the characterization of the fuel (Figure 8.9). These post-harvesting steps can be performed in different ways depending on the strain, substrate, and extraction method employed. Harvesting algal biomass could be the most energy- demanding process due to its concentration, smaller size, and surface charge, especially when the cultures are operated in open pond systems (Singh et al., 2011). Flocculation, sedimenta- tion, and filtration are the common harvesting techniques that are widely used (Harun et al., 2010). Drying the biomass prior to extraction is a prerequisite so as to avoid moisture inter- ference with the solvents. Drying can be performed using dryers or by exposing the biomass to diffused solar drying. Exposure to solar drying minimizes the production cost as well as power consumption. Subsequent to drying, cell disruption, oil extraction, transesterification of oil to fuel, and characterization of the fuel are explained in the following sections.
8.6.1 Cell Disruption
The disruption of algae cells prior to extraction is of particular importance because the con- tents of the extracted lipids are determined according to the disruption method and device employed. The selection of appropriate device for disruption is the key factor for enhancing the lipid extraction efficiency (Lee et al., 2010). The following are the methods commonly used for the disruption of algae cells.
8.6.1.1 Expeller Press Method
Expeller pressing (also calledoil pressing) is a mechanical method applied for the disrup- tion of algae cell membranes by squeezing the cells under high pressure (Mercer and Armenta, 2011). Expeller pressing can also be used as an extraction technique because it
Pre-harvesting
Post-harvesting
Wastewater CO2
Cultivation of microalgae Biomass harvesting
Drying of biomass Cell disruption Extraction of lipids Transesterification
Glycerol Fatty Acid Methyl Esters
(FAME) - Biodiesel
Extraction
FIGURE 8.9 Schematic view of the processes involved in microalgae processing, from algae biomass cultiva- tion to biodiesel production
172 8. ALGAE OILS AS FUELS
can recover nearly 75% of the oil from algae cells in a single step. The advantages of this method include elimination of a solvent requirement and easy operation, the drawback associated with it is the requirement of a large amount of biomass.
8.6.1.2 Bead-Beating Method
The bead-beating method involves the application of beads for the disruption of the algal cell wall. Continuous exposure of biomass to beads leads to cell-wall rupture, resulting in the release of intracellular contents into the solvent medium. Similar to expeller pressing, this method can also be applied for both disruption and extraction. The influence of bead beating on cell-wall disruption was evaluated for the strainsBotrycoccus braunii,Chlorella vulgaris,and Scenedesmussp. using a bead beater (bead diameter of 0.1 mm) (Lee et al., 2010). The method showed a lipid productivity of 28.1%.
Though the disruption of algae cell walls prior to extraction requires an additional step, which is the selection of a cost-effective method, it helps to enhance lipid production efficien- cies. The methods discussed here are economical and applicable to mass cultures compared to few other techniques, such as microwaves, sonication, and autoclaving.
8.6.2 Extraction of Algae Oil
Microalgae are composed of single cells surrounded by an individual cell wall, which in- cludes “unusual” lipid classes and fatty acids that differ from those in higher animals and plants (Guschina and Harwood, 2006). For extraction of lipids from microalgae, regular extraction methods may not be applicable (Eline et al., 2012). Extracting and purifying oil from algae is considered challenging due to its energy- and economically intensive nature (Fajardo et al., 2007; Lee et al., 2010; Mercer and Armenta, 2011).
8.6.2.1 Solvent Extraction
The existing procedures for the extraction of lipids from source material usually involve selective solvent extraction, and the starting material may be subjected to drying prior to ex- traction (Lee et al., 2010). Lipids are soluble in organic solvents but sparingly soluble or in- soluble in water. Solubility of lipids is an important criterion for their extraction and typically depends on the type of lipid present and the proportion of nonpolar lipids (princi- pally triacylglycerols) and polar lipids (mainly phospholipids and glycolipids) in the sample (Huang et al., 2010). Several solvent systems are used, depending on the type of sample and its components. The solvents of choice are usually hexane in the case of Soxhlet and Goldfish methods (Additions and Revisions, 2002); chloroform/methanol or chloroform/methanol/
water in the case of the Folch Method (Folch and Sloane-Stanley, 1957); or modified Bligh and Dyer Procedure (Bligh and Dyer, 1959). This method is best suited to extract nonpolar lipids because polar lipids are scarcely soluble in nonpolar solvents.
8.6.2.2 Soxhlet Extraction
The Soxhlet extraction procedure is also used commonly for oil extraction. The goldfish extraction procedure may also be employed for this purpose. The Soxhlet extraction proce- dure is a semicontinuous process that allows the buildup of a solvent in the extraction 173
8.6 PREPARATION OF ALGAL FUEL/BIODIESEL
chamber for 5 to 20 minutes (Additions and Revisions, 2002). The solvent surrounding the sample is siphoned back into the boiling flask. The procedure provides a soaking effect and does not permit channeling. Polar and bound lipids are not recovered from this method.
8.6.2.3 Wet Lipid Extraction
The wet lipid extraction process uses wet algae biomass by using solvent proportionately (Sathish and Sims, 2012). This method resembles the solvent extraction process but varies with the nature of biomass (wet). The advantage of the process includes the elimination of a drying step, the interference of moisture content with the extraction solvents and lack of wide applicability to all kinds of solvents are the major limitations of this extraction procedure.
8.6.2.4 Hydrothermal Liquefaction
Hydrothermal liquefaction is a process in which biomass is converted in hot compressed water to a liquid biocrude (Brown et al., 2010; Biller et al., 2012). Processing temperatures range from 200–350C with pressures of around 15–20 MPa, depending on the temperature, because the water has to remain in the subcritical region to avoid the latent heat of vaporiza- tion (Biller et al., 2012). At these conditions, complex molecules are broken down and repolymerized to oily compounds (Peterson et al., 2008). This procedure is ideal for the con- version of high-moisture-content biomass such as microalgae because the drying step of the feedstock is not necessary.
8.6.2.5 Ultrasonic Extraction
Ultrasonic-assisted extractions can recover oils from microalgae cells through cavitation (Harun et al., 2010). During the low-pressure cycle, high-intensity small vacuum bubbles are created in the liquid. When the bubbles attain a certain size, they collapse violently during a high-pressure cycle. During the implosion very high pressures and high-speed liquid jets are produced locally, and the resulting shear forces break the cell structure mechanically.
This effect supports the extraction of lipids from algae (Wei et al., 2008). The high-pressure cycles of the ultrasonic waves support the diffusion of solvents, such as hexane, into the cell structure. As ultrasound breaks the cell wall mechanically by the cavitation shear forces, it facilitates the transfer of lipids from the cell into the solvent (Cravotto et al., 2008).
8.6.2.6 Supercritical Carbon Dioxide Extraction (SC-CO2)
Carbon dioxide usually behaves as a gas in air at standard temperature and pressure (STP) or as a solid calleddry icewhen frozen (Sahena et al., 2009; Mendiola et al., 2007). If the tem- perature and pressure are both increased from STP to at or above the critical point for carbon dioxide, CO2can adopt properties midway between a gas and a liquid and behave as a su- percritical fluid, expanding like a gas but with a density like that of a liquid. Supercritical CO2
is becoming an important commercial and industrial solvent due to its role in chemical extraction in addition to its low toxicity and environmental impact (Cooney et al., 2009).
The relatively low temperature of the process and the stability of CO2also allow most com- pounds to be extracted with little damage or denaturing. The main drawbacks of this method include high power consumption and expense and difficulty involved in scaling up at this time (Eller, 1999).
174 8. ALGAE OILS AS FUELS
8.6.2.7 Pulse Electric Field Technologies
Pulsed electric field (PEF) processing is a method for processing cells by means of brief pulses of a strong electric field (Guderjan et al., 2007). Algal biomass is placed between two electrodes and the pulsed electric field is applied. The electric field enlarges the pores of the cell membranes and expels its contents (Guderjan et al., 2004).
8.6.2.8 Enzymatic Treatment
Enzymatic extraction uses enzymes to degrade the cell walls, with water acting as the solvent (Mercer and Armenta, 2011). This makes the fraction of oil much easier. The combi- nation of “sono-enzymatic treatment” causes faster extraction and higher oil yields compared to individual ultrasonication and enzymatic extractions alone (Fajardo et al., 2007). The draw- backs associated with the process are lack of commercial feasibility and inapplicability for mass cultures (Halim et al., 2011).
8.6.2.9 Osmotic Shock
Osmotic shock or osmotic stress is a sudden change in the solute concentration around a cell, causing a rapid change in the movement of water across its cell membrane (Fajardo et al., 2007). This shock causes a release in the cellular contents of microalgae. The method is more applicable for the strains cultivated in marine environments (eg.Nannochloropsissp.). Os- motic shock is also induced to release cellular components for biochemical analysis (Mario, 2010). This method is also applied forHalorubrumsp. isolated from saltern ponds. The results showed increased lipid productivities and variations in lipid compositions (Lopalco et al., 2003).
Extraction of lipids is a key aspect involved in biomass-to-biodiesel production, the method directly influences the lipid productivity potential of the process. So far, several methods have been employed for extracting the cellular contents (lipids) of microalgae. Each method has its own advantages and disadvantages for practical applicability. Among the pro- cesses described, solvent extraction is suitable for extracting lipids from mass cultures but requires large volumes of solvent. The Soxhlet extraction method is applicable only when a single solvent is used and is not suitable for binary solvent applications. However, recovery and reusability of the solvent are possible with this method. The ultrasonic extraction method can perform well when coupled with the enzymatic treatment, but both methods lack cost effectiveness and feasibility for large-scale applications. Supercritical carbon dioxide extrac- tion (SC-CO2), pulse electric field procedure, osmotic shock, hydrothermal liquefaction, and wet lipid extraction require more optimization efforts for large-scale applications. A suitable method operatable with both binary and single solvents, applicable at large scales and yield- ing higher lipid productivities, is yet to be optimized for achieving enhanced microalgae lipid yields.