Sewage sludge consists principally of microorganisms and organic materials and is a by­product of the aerobic treatment of sewage. Sewage sludge can be disposed of on land, dried, put in landfill, composted, or incinerated, but one of the most efficient methods of disposal is to anaerobically digest the sewage sludge. The advantage of anaerobic digestion is that it reduces the volume of sludge, produces biogas and as it is enclosed less smell is associated with the process. There are a number of anaerobic digester designs but the simplest design is a large, sealed vessel of 200,000-400,000 l which is maintained at 30-37°C for 30 to 60 days (Fig. 5.5). The sewage sludge is broken down by anaerobic microorganisms producing a gas which is predominantly methane (biogas) and also contains carbon dioxide and hydrogen. A digester can produce gas at a rate of

1.0 m3/m3/day that represents between 4.2 and 10.4 GJ. The biogas collected has a reduced calorific value compared with 100% methane as it contains carbon dioxide.

One of the problems of using enzymes to degrade lignocellulose is the cost of the enzymes. Recent reduction in the cost of cellulose enzymes (Greer, 2005) has allowed lignocellulose to be considered as a viable alternative to sugar and starch as a substrate

for ethanol production. A number of countries have pilot plants in operation process­ing lignocellulose, although most are in the USA (Table 6.10) and larger commercial plants are being developed. Figure 6.13 shows the process developed by Iogen, a Canadian enzyme manufacturer, for the use of lignocellulose to produce ethanol. Wood chips or straw are treated with dilute acid steam explosion and the cellulose and hemi — cellulose hydrolysed with enzymes produced by Trichoderma sp. Initially the fermenta­tion used S. cerevisiae but later a recombinant Z. mobilis has been introduced which could ferment the pentoses produced from the hemicellulose. The lignin is separated and burnt in a combined heat and power unit to provide energy for the process.

Another process is that designed by the National Renewable Energy Laboratory (NREL) (Fig. 6.14). The biomass, maize stover, is pretreated with dilute acid followed by simultaneous saccharification and co-fermentation (SSCF) with a recombinant Z. mobilis capable of fermenting glucose, xylose and T. reesei. The cellulose hydroly­sis is carried out with T. reesei cellulases and the hemicellulase hydrolysate is detoxi­fied before adding to the fermenter.

One of the advantages of biodiesel and some of the other biofuels is that they are non-toxic and degrade more rapidly than fossil fuels. This is an important feature in the case of accidents and spillages. Marine environments, freshwater, soil and various

184

Fig. 8.21. Carbon dioxide produced per unit of energy for gaseous fuels compared with some fossil fuels. CNG, compressed natural gas; LPG, liquid petroleum gas; DME, dimethyl ether; hydrogen produced from electrolysis, natural gas and coal. (From Gustavsson et al, 1995; Matthews, 2001; Gielen and Unander, 2005; IEA, 2005a.)

sediments have been contaminated with oil components throughout the world as a result of accidents, leaks, spills and disposal. Oil components can cause considerable environmental disruption and the most spectacular are those accidents involving oil tankers.

One definition of biodegradability is where a fuel is 90% or more degraded within 21 days under fixed conditions (Sendzikene et al., 2007). The determination of degradation can be followed by the reduction in the concentration of the fuel as estimated by analysis such as gas liquid chromatography or the release of breakdown products such as carbon dioxide (Zhang et al., 1998). The two methods give some­what different results. Using carbon dioxide emissions rapeseed methyl esters degrade by 77-89% in 28 days compared with 18% for mineral diesel. Using gas liquid chromatography (GLC) the values were 88% for rapeseed methyl esters and 26% for mineral diesel (Zhang et al., 1998). This is not surprising as there should be a delay for the fuels to be fully mineralized to carbon dioxide compared with the metabolism of the methyl esters.

More recently the concentrations of oils and fatty acid methyl esters have been estimated using infrared spectroscopy and Fourier-transformed infrared spectroscopy (FTIR). In Europe, the biodegradability of fuel has been measured using infrared with the absorption of the C-M stretch of CH2—CH3 at 2930 cm-1 (Sendzikene et al., 2007). In other determinations the band at 1573 cm-1 (COO-) was used (Al-Alawi et al., 2006).

The biodegradation of a number of plant oil — and animal fat-derived biodiesel has been determined using a microbial mixture obtained from a wastewater system. The results are shown in Fig. 8.23. Diesel was about 60% degraded by day 21 whereas linseed-, tallow-, lard — and rapeseed-derived biodiesel were over 90% degraded in the same time. The rate of degradation was greatest with tallow — and lard-derived biodiesel.

Mixing biodiesel with mineral diesel increases the rate of degradation of mineral diesel and as the concentration of biodiesel increases so does the rate of degradation (Table 8.3). The reason for the increase in degradation is not known but may be due

20 0

Fig. 8.23. Biodegradation of various biodiesel preparations over 21 days (Redrawn from Sendzikiene et al., 2007.)

either to the provision of a more accessible substrate biodiesel or the solubilization of mineral diesel. Biodiesel has been shown to solubilize mineral diesel and has been used to remove crude oil from contaminated sand (Pereira and Mudge, 2004).

Harnessing the power of the wind is one of the most promising alternative methods of electricity generation as it has the potential to generate substantial amounts of energy without pollution. Wind can also be used to drive water pumps in order to store energy, to charge batteries in remote regions, or as off-grid power sources. The potential for wind power has been recognized and wind farms have been installed in at least 15 countries including Brazil, China, Denmark, Spain, USA, India and the UK (Herbert et al., 2007).

Geothermal

The centre of the Earth is very hot at about 4000°C, and most of the heat which reaches the surface cannot be utilized, but in areas of volcanic activity high-grade heat is retained in molten or hot rocks at a depth of 2-10 km. The heat from these hot or molten rocks can be extracted from hot springs and used to run steam turbines directly for the generation of electricity. If the water is below 150°C, it can be used as a supply of hot water for industrial or domestic heating.

Solar energy

Sunlight can be used either directly or indirectly for solar panels for hot water gener­ation, solar collectors for steam generation, solar architecture for heating buildings, solar thermal-electric, and steam linked to electricity generation, photovoltaic, direct generation of electricity and solar hydrogen generation. A recent review of photo — voltaics was published in 2007 (Jager-Waldau, 2007).

The advantage of hydrogen as a fuel in the internal combustion and gas turbine engine is the product of combustion is only water and the exhaust contains no carbon dioxide, sulfur dioxide or carbon monoxide. Burning hydrogen does however pro­duce NOx which is a function of the temperature where hydrogen burns at a little higher temperature than petrol. This is not a problem encountered with fuel cells. Research is under way using recirculation of exhaust gas to reduce NOx emissions (Heffel, 2003). Both liquid hydrogen and fuel cell vehicles have been developed by a number of automotive companies. The problems with hydrogen are its supply to vehicles and storage on board.

If we consider that hydrogen may be used as a universal transport fuel either in an internal combustion engine or in a fuel cell, then a new infrastructure will be required. The main question is what needs to be developed first, the hydrogen power vehicles or the infrastructure? The widespread use of hydrogen-powered vehicles will need a network of filling stations but these will probably not be built until there is a number of vehicles to justify the cost of construction. The most likely sequence will be the construction of the infrastructure prior to the widespread introduction of hydrogen-powered vehicles. A study of the early development of the fossil fuel automotive industry illustrates the same problems (Melaina, 2007). The car was introduced via mass production but in the beginning there were few petrol stations, as is the case for hydrogen at present. However, motorists could obtain fuel in cans from shops and repair garages. This supply system had developed for the sales of paraffin (kerosene) for lighting and heating before the introduction of cars. As the car ownership increased there was a demand for filling stations which were built in increasing numbers. Such is the nature of compressed or liquid nitro­gen that small-scale supplies cannot be obtained without the development of a widespread infrastructure.

Some of the options for the supply of hydrogen to filling stations are shown in Fig. 5.14. The first option is to produce hydrogen, in this case from natural gas, at a central production unit. The hydrogen is liquefied and transported to the filling sta­tion by tanker where it is stored until required. This is very much the system that is

Reforming Compressor

Fig. 5.14. Some of the possible methods of supplying hydrogen to petrol stations.

used at present for petrol and diesel. In the second option the hydrogen is generated at the filling station either by electrolysis or from natural gas and stored. In the third option hydrogen is supplied to the filling station by pipeline where it is compressed or liquefied. All these options have some of the problems outlined above, including the energy required to compress or liquefy the hydrogen, its storage and its dispen — sion. A well-to-wheel assessment of the supply of hydrogen and storage on the vehicle concluded that the energy used to deliver hydrogen in a liquid form was similar to that for petrol and diesel.

The on-site generation was much more costly in terms of energy. In both cases 80% of the energy use was expended on hydrogen production and liquefaction or compres­sion (de Wit and Faaij, 2007). When the fuel delivery and driving costs were considered, on-site generation was again the most expensive. The driving costs were higher than petrol and diesel in all cases.

Once the hydrogen has been made available there are a number of options for its use in the vehicle for both internal combustion engines and fuel cells (Ogden et al., 1999). Compressed hydrogen or liquid hydrogen will have to be stored in

cylinder/tanks at pressure of up to 600 psi or at -253°C in insulated tanks. Either method of storage will pose problems when filling the vehicle. There are options to avoid the on-board storage of hydrogen which includes the on-board produc­tion of hydrogen from methanol or petrol (Fig. 5.15). Both methanol and petrol are liquid and can easily be supplied by the present infrastructure, but petrol is non-sustainable and methanol would need to be produced in a sustainable manner. Both systems would add weight, complication, increase fuel consumption and cost to the vehicle.

The two critical stages in product development are the harvesting of the microorgan­isms and the extraction of the product as these unit operations can add considerable costs to the process. For microalgae, there are a number of methods for harvesting

the cells including centrifugation, filtration, flocculation and settling. Flocculation can be used to improve the other methods of harvesting. Flocculation can be carried out using multivalent metal salts (Molina Grima et al., 2003) or cationic polymers. Centrifugal recovery is rapid and expensive but has been used for many microalgae. Filtration using filter presses has proved unsuccessful with the smaller microalgae, and membrane filtration has not been extensively used.

Extraction

Intracellular oils are difficult to extract from wet biomass (Belarbi et al., 2000), but can be more easily extracted from freeze-dried cells or cell paste. Oil has been extracted from Phaeodactylum tricormutum (diatom) and Monodus subterraneus (green alga) with solvents under pressure (Belarbi et al., 2000). In the case of C. protothecoides, the cells were freeze-dried before solvent extraction as were the oils from Isochrysis galbana (Molina Grima et al., 1994). Microalgal cells may be disrupted to extract the oils using a number of microbial cell disruption methods, but these methods can be expensive. Free fatty acids have been extracted from wet biomass using a potassium hydroxide — ethanol mixture (Molina Grima et al., 2003). Whole cells of Dunaliella tertiolecta have been liquefied at 300°C and 10 MPa to form oil comparable to fuel oil. Both supercriti­cal CO2 (Mendes et al., 2003; Gouveia et al., 2007) and thermochemical liquefaction have also been used to produce biodiesel from macroalgae (Aresta et al., 2005). In addi­tion, whole microalgal cells containing high levels of oil have been used directly in diesel and biodiesel in emulsion fuels (Scragg et al., 2003). In general, all methods both mechanical and solvent based are expensive and will affect the cost of the biofuel.

In the USA, the main crop for ethanol production is maize and since 1997 genetic manipulation has increased the yield of maize per hectare from 7.5 to 9.0 Mt/ha which is above the increase expected by traditional plant breeding (McLaren, 2005). This has been achieved by engineering maize to be resistant to the European corn borer by inserting a gene from Bacillus thuringiensis (Bt) that codes for a protein with insecticide activity. Another example is the pollen beetle which is a pest affecting the pollen of rapeseed, the main European source of biodiesel. There are no natural plants resistant to this pest capable of breeding with rapeseed and so genetic manipu­lation was the only solution. Transgenic rapeseed plants have been produced carrying a pea lectin gene and the lectin was found to be toxic to the pollen beetle larvae. The pea lectin was placed under the control of a pollen-specific promoter so that the lectin is not present in the rest of the plant (Ahman et al., 2006).

One of the important characteristics of some of the perennial grasses is the possession of C4 metabolism rather than C3. C4 and C3 metabolism refer to the pathways used

to assimilate carbon dioxide during photosynthesis. C4 plants are more efficient at higher light and temperatures compared to C3 plants. The C4 plants have a lower moisture content, require less fertilizer input and are twice as efficient with water. C4 assimilation of carbon is theoretically 350 kg/ha/day compared with 200 kg/ha/day for C3 plants (Venturi and Venturi, 2003). All these features make C4 plants more suitable for biomass fuel planting than C3 plants.

The development of the C4 metabolism of carbon dioxide assimilation evolved from the Calvin cycle in C3 plants to avoid the loss of carbon dioxide through photorespiration. The fixation of carbon dioxide during photosynthesis takes place in three stages. The addition (carboxylation) of carbon dioxide to ribulose-1,5-bisphosphate (Fig. 4.3) is fol­lowed by the reduction of 3-phosphoglycerate to glyceraldehyde-3-phosphate. Ribulose-1, 5-bisphosphate is then regenerated from glyceraldehyde-3-phosphate. The first step of the Calvin cycle is catalysed by the chloroplast enzyme ribulose bisphosphate carboxylase/ oxygenase known as rubisco. However, another property of the rubisco enzyme is to catalyse the oxygenation of ribulose-1,5-bisphosphate which is the start of light-dependant oxygen uptake and carbon dioxide release, known as photorespiration, which reduces plant yield.

In the C4 metabolism two different types of cell are involved in photosynthesis, the mesophyll and bundle sheath cells (Fig. 4.4). In the mesophyll cells carbon dioxide is used to carboxylate phosphoenolpyruvate (PEP) forming oxaloacetate. The oxalo — acetate is converted to malate (C4) and this is transferred to the bundle sheath cell where the malate is converted into pyruvate and carbon dioxide. The carbon dioxide is then used in the Calvin cycle. The C4 cycle has a higher energy demand but the cycle reduces photorespiration and water loss. The phosphoenolpyruvate (PEP) carboxylase enzyme has a high affinity for the carboxyl ion such that it is saturated and in equilibrium with carbon dioxide gas. Oxygen is not a competitor in the PEP

carboxylase reaction because the substrate is a carboxyl ion. The high activity of the PEP carboxylase allows the plants to reduce the stomatal opening, reducing water loss, while fixing carbon dioxide at an undiminished rate. The high concentration of carbon dioxide in the bundle sheath cells allows the cells to carry out photosynthesis at high temperatures.

Animal and municipal waste

Animal wastes consist of excess slurry and dung from cattle, chickens and pigs. These wastes can be used to generate biogas through anaerobic digestion and there are a number of farm-sized units available. There are also a number of small electricity power stations which run on chicken slurry from large battery chicken farms.

S. cerevisiae is not the only microorganism capable of producing ethanol (Table 6.6). Other yeasts, bacteria and fungi can also ferment sugars to produce ethanol. Ethanol — producing bacteria have been of commercial interest because they have a faster growth rate and can be easily genetically engineered. Escherichia coli uses a different pathway to produce ethanol from pyruvate (Fig. 6.8) with an enzyme pyruvate for­mate lyase forming acetyl-CoA, which is then reduced by two steps with alcohol dehydrogenase to ethanol.

Other ethanol-producing bacteria metabolize glucose via the Entner-Doudoroff pathway which consumes less ATP than glycolysis (Fig. 6.9). One of the best examples

Table 6.6. The substrates that can be utilized by ethanol-producing microorganisms. Organism Substrate utilized

Glucose, fructose, galactose, maltose, maltotriose, xylulose Glucose, fructose, galactose, maltose, maltotriose, xylulose Glucose, galactose, lactose Glucose, xylose, xylulose

Glucose, fructose, sucrose, engineered to use xylose Glucose, cellobiose, cellulose Xylose, no ethanol tolerance Xylose, cellobiose, glucose

Xylose to acetone and butanol, ethanol in small quantities Uses cellobiose faster than glucose Lactose, useful for whey utilization Uses cellobiose if nutrients supplied, glucose, xylose and arabinos

is the bacterium Zymomonas mobilis which has a higher growth rate, is more ethanol- tolerant than yeasts but still only metabolizes glucose, fructose and sucrose.

Biodiesel has been produced using a very wide range of plant oils and animal fats, waste cooking oils and soapstocks. Table 7.17 gives some of the plant oils that have been converted into biodiesel and Table 7.18 the animal fats, waste cooking oil and soapstocks.

All these sources of oil used for biodiesel production have characteristic and dif­ferent fatty acid profiles and these can influence the properties of the subsequent biodiesel.

Fig. 7.19. Effect on the lubricity of conventional diesel when biodiesel is added. The lubricity was measured by ball-on-cylinder lubrication evaluator (BOCLE). (Redrawn from Graboski and McCormick, 1998.)

Table 7.17. Plant oils that have been used to produce biodiesel.

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