Currently, the commercial production of biodiesel is mainly from plant oil, which is far below the demand of transport fuels, because huge amounts of arable land are required for the cultivation of oil plants. Taking the United States as an example, if oil palm, a high-yielding oil crop, is used for biofuel production, 24% of the total national cropland would have to be devoted to meet only 50% of the transport fuel needs (Chisti, 2007; Mata et al., 2010). The requirement for huge amounts of arable land and the resultant conflicts between food and oil make biofuels from plant oil unrealistic to completely replace the petroleum-derived fuels in the foreseeable future. In this regard, microalgae, which are believed to possess various advantages over other candidates, have been envisioned as an emerging feedstock for biofuels. Microalgae, also known as microphytes, refer to algae in microscopic size ranging from a few micrometers to a few hundreds of micrometers. The research on algal oil for biodiesel production can be traced back to as early as 1970s, and the main superiorities of microalgae include (1) high oil contents, with some species exceeding 60–70% of the dry weight of biomass, much higher than oil crops; (2) rapid growth rates, much faster than higher plants, whilst the entire growth cycle can be completed within a few days; and (3) a better adaptability to distinctly different environmental conditions from water to land and even in unusual locations such as snow and desert soil, which causes no competition with food for arable lands. Much less land is needed if using microalgae as biodiesel feedstock. According to Chisti (2007), only 2.5% of the existing US cropping area would be sufficient for producing algal biomass to satisfy 50% of the transport fuel needs, which accounts for 0.7% of soybean. These unique properties, in combination with the CO₂ capture and recycling capacities, make microalgae a promising cell factory for biofuel production. The key properties of microalgae-derived biofuels, eg, energy density, viscosity, flash point, cold filter plugging point, and acid value, have been proved to comply with the specifications established by the American Society for Testing and Materials (Xu et al., 2006). In this chapter, an overview of the current research progress of microalgal biofuels is presented; obstacles and future research trends are also discussed.

Biomass harvest is a highly challenging process because of three reasons: (1) The nature of microalgae. Most algal cells are negatively charged so that the formation of aggregates is prevented. Besides, microalgae have tiny cell dimension and similar specific gravity to water, which leads to a very slow sinking rate. These properties make the algal suspension highly stable and difficult to harvest; (2) The biomass concentration. Culture broths are generally diluted and the concentration of biomass is low. Some commercial production systems only have less than 0.5 kg/m³ of dry biomass. Hence large volumes have to be handled; and (3) The cost. The capital requirement is high, especially when the microalgae are cultivated in saline systems (Molina Grima et al., 2003; Pahl et al., 2013).

The most common harvesting methods include flocculation, flotation, gravity sedimentation, centrifugation, and filtration. Flocculation is a preparatory step prior to others. As algal cells are negatively charged, the addition of flocculants, such as multivalent cations and cationic polymers, can neutralize the surface charge. As a result, the particle size can be effectively increased to disrupt the stability of the system and facilitate the aggregation. Ferric chloride (FeCl₃), aluminum sulfate [Al₂(SO₄)₃], and ferric sulfate [Fe₂(SO₄)₃] are commonly used multivalent metal salts (Brennan and Owende, 2010). Some researchers found that the effective flocculation may be simply achieved by adjusting the pH of the algal broth (McCausland et al., 1999; Knuckey et al., 2006). Poelman et al. (1997) reported that the efficiency of algal removal was 80–95% when electrolytic flocculation was applied. Flocculation technique can be used to handle large quantities of culture, and it is less energy-intensive than mechanical separation. However, this method alone is not sufficient, and other processes need to be combined.

Flotation is an air/gas-assisted separation process in which air or gas bubbles attach to algal cells, allowing the formation of float on the liquid surface for harvesting. This method can capture particles with a diameter of less than 500 μm (Chen et al., 2011). Dissolved air flotation and dispersed air flotation are the two commonly used flotation techniques. Usually, surfactants such as cationic N-Cetyl-N-N-N-trimethylammonium bromide (CTAB) and anionic sodium dodecylsulfate (SDS) are used to enhance the likelihood of microalgae–bubble attachment and thus the flotation efficiency.

Gravity sedimentation is a conventional method which has low efficiency and is time-consuming. It is considered to be only suitable for large particles and can be applied in the algal wastewater treatment system because of the large volumes treated and the low value of the biomass obtained (Nurdogan and Oswald, 1996). Similarly, the method of filtration is also only suitable for the recovering of large microalgae (>70 μm), such as Coelastrum and Spirulina. To recover the smaller microalgae (<30 μm), membrane-based microfiltration and ultrafiltration may be applied (Hung and Liu, 2006; Zhang et al., 2010), but they are more expensive. In short, the selection of a harvesting method depends on the microalgal properties and the final product desired.

The harvesting process generally produces a biomass slurry with a solid content of 20–30%. The concentrated biomass, however, still contains substantial moisture, which can spoil the biomass within several hours if exposed to a hot environment. Therefore, dewatering is an important step to keep the quality of oil. The harvesting and drying processes contribute 20–30% of the total cost of microalgal biomass production (Molina Grima et al., 2003). Sun drying is the oldest method that works well in low-humidity climates. However, due to the high water content of algal biomass, this method is not very effective for algal powder production. Also, it always introduces unpleasant odors and potential bacterium contamination. Other commonly used drying methods include freeze-drying, spray drying, drum drying, and so on (Lin, 2005). Compared with sun drying, they are more efficient, but also more energy-intensive. It is estimated that the removal of 1 kg of water requires over 800 kcal of energy. Thus, dewatering is considered as one of the main economical bottlenecks in the entire process.

Mechanical disruption includes pressing, bead milling, and homogenization. When 100–200 g/L of biomass concentrations are used, bead milling is the most effective and economical way (Greenwell et al., 2010). Osmotic shock treatment can also be used to break algal cells for oil extraction, especially suitable for cell-wall-lacking microalgae. It is a sudden reduction in osmotic pressure causing cell rupture and release of cellular components including oil. Ultrasonic-assisted extraction is another approach, where the collapsing cavitation bubbles help cause cell walls to break and release the oil into the solvent. After being mechanically disrupted, the algal cells are exposed to solvents. Bligh and Dyer (1959) procedure, originally designed to extract lipids from fish tissue, has been used as a benchmark for comparison of solvent extraction techniques. A wide range of organic solvents such as benzene, cyclohexane, hexane, acetone, and chloroform have been proved to be effective in treating algal cells, of which hexane is the major solvent used. In some cases, the solvent extraction can be enhanced by using organic solvents at temperature and pressures above the boiling point, which is known as the accelerated solvent extraction (Cooney et al., 2009). Supercritical CO₂ is another way for efficient extraction of microalgal oil, but the high energy demand is a limitation for commercialization of this technology (Herrero et al., 2010). The effectiveness between these methods is shown in Table 22.3.

The oil obtained from microalgae usually has a higher viscosity than diesel oil, which cannot be directly applied to engines. Transesterification is needed to reduce the viscosity and increase the fluidity. Transesterification occurs stepwise with the first conversion of TAG to DAG and then to MAG and finally to glycerol. In this process, large, branched triglycerides are transformed into smaller, straight-chain molecules, which are similar in size to the molecules of the species present in diesel fuel (Sinha et al., 2008).

The complete transesterification of 1 mol of TAG requires 3 mol of alcohol, producing 1 mol of glycerol and 3 mol of fatty esters (Fig. 22.8). As the reaction is reversible, alcohol is commonly added in excess in industrial processes to ensure the direction of fatty acyl esters. A number of alcohols can be used as the substrates, eg, methanol, ethanol, propanol, butanol, and amyl alcohol. Methanol is the most preferable one because of its low cost. The most abundant composition of microalgal oil transesterified with methanol is C₁₉H₃₆O₂, which has been demonstrated to meet the standard of biodiesel (Xu et al., 2006).

A catalyst is needed to facilitate the transesterification. The catalysts can be acids, alkalis, or enzymes. Acid-mediated transesterification (such as sulfuric, sulfonic, phosphoric, and hydrochloric acids) is suitable for the conversion of feedstock with high free fatty acids. However the reaction rate is usually slow, and the equipment could be damaged because of the corrosion by acids. On the contrary, alkali-catalyzed transesterification has a much higher reaction rate, approximately 4000 times faster than the acid-catalyzed one (Fukuda et al., 2001). Therefore alkali is preferred as the catalyst for industrial production of biodiesel. The commonly used alkali catalysts include sodium hydroxide (NaOH), potassium hydroxide (KOH), and sodium methoxide (CH₃ONa). For those free fatty acids-containing samples, an acid–alkali two-step transesterification could be the best choice. In addition to acid–base catalysis, enzymes are also widely used for transesterification. For example, the use of lipase has attracted much attention as it produces high-purity product and enables easy separation from the byproduct glycerol (Ranganathan et al., 2008). However, the cost of enzyme is still relatively high and remains a barrier for its industrial implementation. In addition to catalytic transesterification, some noncatalytic transesterification techniques are also available, such as supercritical fluid. Their comparison is given in Table 22.4.