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Chapter 6 Fuel and Energy Microorganisms have a historic role in the fermentation of a variety of organic materials to alcohols, acids, and CO2, mainly related to the manufac- ture of foods, beers, and wines. Today alcohol production is largely synthetic, that is, nonmicrobial, although the rising costs of petroleum have created renewed interest in the production of ethanol by fermentation for use as a fuel. For example, ambitious programs for production of large quantities of fuel ethanol by fermentation have been undertaken in Brazil, India, and several other countries. Similarly, biogas is being produced as a source of energy in several countries. The most practical process to produce fuel for farm and community use by microbial processes is the generation of biogas (Table 6.1~. Ethanol for fuel requires a capital investment more in keeping with regional or large farm-cooperative manufacture. The technology for both methane and ethanol manufacture is readily accessible. The generation of hydrogen and methanol through microbial processes is still in the laboratory stage. Although there are organisms that yield hydrogen and methanol from organic substrates, much more development work is re- quired to make these processes economically feasible. TABLE 6.1 Characteristics of Fuels from Microbial Processes Approximate Gross Typical Fuel Energy Content Sources Process Considerations Liquids Methanol 10,000 BTU/lb Methane None commercially 23.8 MJ/kg available Ethanol 13,000 BTU/lb Molasses, grains, Requires significant 30.6 MI/kg plans biomass capitalinvestment Gases Methane 24,000 BTU/lb Animal, human, Practical for farm and 55.5 MJ/kg and agricultural community use wastes Hydrogen 61,000 BTU/lb Algae-nutrient None commercially 142 MJ/kg system available 107

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108 Ethanol MICROBIAL PROCESSES The production of ethanol from residues with a high sugar content may soon be economically practical as a means of offsetting the rising costs of petroleum. Ethanol can be used alone or blended with gasoline or diesel fuels. For this use it need not be of high purity or entirely free of water. The alcohol yield depends on the amount of starch or fermentable sugars present in the substrate; sugar-cane is a suitable raw material because of the large amounts of this crop available in most parts of the world. The economics of ethanol production improve as the size of the plant increases, to the point where costs of collecting raw material to sustain a huge processing unit become too large. For a plant with a capacity of 20 million gallons per year, capital investment would be about $30 million. Plant sche- matics and material requirements for the conversion of molasses and corn to ethanol are shown in Figures 6.1 and 6.2. Estimates of the areas required for various crops to support a 100,000 t per year plant are shown in Table 6.2. The fermentations shown in Figures 6.1 and 6.2 are conducted at atmos- phenc pressure. However, for the production of volatile products like ethanol, both rapid fermentations using a vacuum, and Me recycling of mi- crobial cells have significant advantages over older conventional methods. When a vacuum of approximately 50 mm of mercury is applied to a fer- mer~ter operating at 35C, the ethanol can be removed continuously as it is produced by yeast. The removal of ethanol overcomes inhibition of the fer- mentation. Reductions in yield and productivity and suppression of yeast growth occur at ethanol concentrations of 7-10 percent. High substrate con- centrations can be used in vacuum fermentations and good yields still TABLE 6.2 Crop Area to Support a 100,000 T Per Year Ethanol Plant Africa (hectares, in thousands) South America Near East Far East Corn 349 253 154 333 Wheat 457 26 8 370 305 Rice 213 213 76 157 Cassava 87 49 135 73 Sugar Cane: Molasses only 120 125 78 145 Total Cane Juice 36 37 23 43 Source: Leo Hepner. 1977. Feasibility of producing basic chemicals by fermentation. In Microbial energy conversion, H. G. Schlegel and J. Barnea, eds. Oxford: Pergamon Press. p. 550

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FUEL AND ENERGY Molasses 1~ acid ~ hank 14 Ammonium su If ate Water _ . . _ . . _ _ _ ~ Carbon 5: dioxide AIdehydes Fermenter | Water Fusel oil ~ r ~ l Beers Yeast L ~ cu Itu re ~!~J machine Material and Utility Requirements r ~ rig cn Stop ~ TIC Benzene Basis1,000 gal 95% ethyl alcohol .= c ~~ E ~ ' al a, Water Ethyl alcohol (95%) ~ 0 Ethyl alcohol (absolute) plus 4 gal fusel oil, 4,800 lb carbon dioxide, 1,000 lb carbon, and 900 lb potash 109 Molasses (blackstrap) 2,400 gal Process water 10,000 gal Sulfuric acid (60 Be) 170 lb Cooling water 42,000 gal Ammonium sulfate 15 lb Electricity 1 10 kwhr Steam 50,000 lb Source: W.L. Faith; D. B. Keyes; and R. L. Clark, 1974. Industrial chemicals. New York: John Wiley and Sons. FIGURE 6.1 Ethanol from molasses by fermentation. achieved. When both vacuum fermentation and cell recycling are practiced, productivities from 10 to 12 times higher than conventional batch processes are achieved. Increases in productivity reduce capital costs and energy re- quirements for fermenter operation. Vacuum distillate from the fermentation liquid contains up to 20 percent ethanol. The cost of distillation to achieve an ethanol concentration of 80-95 percent is then significantly less than that of conventional methods. Applica- tion of these techniques for the production of ethanol should make it more attractive as a substitute for fossil fuel. Production of ethanol and methane from cellulosic wastes by thennophilic microbes is promising for future development. Various lignocellulose sources including rice straw, corn stover, bagasse, etc., may be the ultimate choices for feedstocks because of their lack of alternative value as foods.

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110 Corn Water Steam Vacuum I ~ ~ __. . . . .. _ Entrainment r ~ se pa rato r Sprain Milll _ r ~ ~ l ~ _ _ Degerminator _ ~~ . Lit ~ Press cake Malts l r Converterl and cooler ~ Corn oil 1 - 1 ~ c, Yeast_ _ 4.- l ~ ~ Water Material Requirements MICROBIAL PROCESSES Carbon ~ ioxide Water ~ _ ~ Fusel oil S lop Basis-1,000 gal 95~O ethyl alcohol _ - A~dehydes - _ Ethyl alcohol . ~r ~ (95%) c E _' ~ ~ C: ~ ~ ~ _ _ 3 ~ r __ - Water ~ r plus 400 lb corn oil, 10 gal fusel oil, 4,800 lb carbon dioxide, 750 lb press cake, and 4,400 lb stock feed (dry residue) Corn 372 bu Barley malt 83 bu Yeast Variable Process water 17,000 gal Source: W. L. Faith; D. B. Keyes; and R. L. Clark, 1974. Industrial chemicals. New York: John Wiley and Sons. FIGURE 6.2 Ethanol from corn to fermentation. Lim itations Village production of fuel-grade ethanol (95 percent and above) is inappro- priate because there are no economical small-scale concentration techniques available. Large-scale production requires harvest and transport of the crops over a fairly broad area and costs for these operations must be included. Research Neecis The following research and development activities are needed to facilitate widespread microbial production of ethanol: A survey of the types and characteristics of raw materials for fermenta- tion in various climates and locales; Agricultural research to improve yields of crops, such as cassava and sorghum, which are candidate substrates for alcohol fermentations; Improvements in methods for harvesting and preparation of ferment- able substrates from a variety of agricultural, forestry, and other organic wastes and crops;

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FUEL AND ENERGY 111 Development of equipment and processes for fermentation and recov- ery of alcohol that require less capital, energy, and labor than conventional processes; and Socioeconomic studies to develop optimal systems for harvests trans- port, and processing of crops. Utilization of Cellulose Several anaerobic bacteria of the genus Clostridium have been used ire fermentations of cellulose. C. thermocellum, which has simple nutritional requirements, is the only known thermophilic species that degrades cellulose. Because it grows at higher temperatures (above 50C) than most bacteria, it has the advantage of being less prone to contamination and also has a faster reaction rate than microbes growing at lower temperatures. In pure culture fermentations, the chief products from cellulose are cell mass, acetate, etha- nol, lactate, hydrogen, and carbon dioxide. In a mixed culture of C. therrno- cellum and Methanobacterium thermoautotrophicum, the major products from cellulose are cell mass, methane, and acetate. One can envision the use of C. thermocellum in pure cultures for ethanol production or in mixed cultures for the production of biogas from cellulosic wastes. Limitations The accumulation of acetic acid during fermentation limits growth, and since C. thermocellum cannot decompose lignin, many natural substrates such as wood must be pretreated by acid hydrolysis to make the cellulose available for fermentation. Research Neecis The following requirements are necessary to facilitate the utilization of cellulose as an energy source: Process development for specific substrates and end-products as well as for an optimal fermenter design; Better understanding of the biochemistry of the process; and Development of a means for converting the acetate to nontoxic pro- ducts. Methane Microbiological conversion of organic materials to methane (biogas) is a natural process, providing energy in a clean, gaseous form. Although this

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112 MICROBIAL PROCESSES process will not meet the total energy demands of modern society, it may economically supplement other sources of fuel. Its use will depend on factors such as the cost of fossil fuels, availability and degradability of organic sub- strates, and the availability of trained personnel. Biogas production occurs in marry natural microbial ecosystems such as organic sediments of aquatic systems, marshes and soil, arid in the rumen and large intestine, especially in herbivorous animals. It involves a complex mix- ture of anaerobic bacteria, which convert up to 90 percent of the combustible energy of the degradable organic matter to methane and carbon dioxide. Anaerobic treatment of complex organic mixtures may be considered a three-stage process, as shown in Figure 6.3. In the first stage, a group of facultative microorganisms acts upon the organic substrates. By enzymatic hydrolysis, the complex substances are solubilized and serve as the substrates for microorganisms in the second stage. In the second stage, these soluble organic compounds are converted to organic acids. The acids (primarily acetic) serve as substrates for the final stage of decomposition accomplished by the methanogenic bacteria. These bacteria can produce methane either by converting acetic acid to methane and carbon dioxide or by reducing carbon dioxide to methane, using hydrogen or formate produced by over bactena. Only about 10 percent of the energy is converted into microbial cells that obtain energy for growth during the conversion. With organic waste materials such as cattle manure or urban organic refuse, in theory 30-50 percent of the combustible energy could be converted to methane. With an efficiently operated digester and substrate materials such as cattle waste, as much as 4.5 liters of methane can be produced per liter of reactor material each day. With some vegetable materials or forages, even higher rates are possible, and as much as 70 percent of the energy can be converted to methane. , Protein-decomposing / PROTEINS . organisms 1~3~ I organisms lll Elm ~ I organ isrns \ I Soluble I I STAGE 3 - .-1 AC3d ~1 Compounds I Bacteria Lids I I Bacteria | CO2 | 1 I ll Source: National Academy of Sciences. 1977. Methane generation from human, animal, and agricultural wastes. Washington, D.C.: National Academy of Sciences. P. 28. FIGURE 6.3 Anaerobic fermentation of organic solids. ll

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FUEL AND ENERGY 113 The economically valuable substances such as ammonia nitrogen, phos- phate, and microbial cells are retained in the reactor effluent and residue, and these may have value as fertilizer or as an animal feed supplement. The residue also has value as a soil conditioner, and it usually does not attract insects or have the disagreeable odors often associated with animal manure. Such residues can also be dried and burned to obtain additional heat energy. The fermentation process can be applied to sewage effluent as ~ step in water recycling by removing nutrients and facilitating subsequent water purification (see Chapter 8~. The gas produced can be an economic incentive for waste treatment. In addition, the process also converts malodorous and pathogenic waste into an innocuous, potentially useful sludge. The technology of biogas production is highly developed and can be applied economically to many organic substrates, depending on their bio- degradability, their alternative uses, the possible economic value of by- products, and the competitive use of fossil fuel. Thousands of small-scale plants of family, farm, or village size have been operated, especially in parts of Asia and in Europe. A typical methane plant in India based on cow manure is Mown in Figure 6.4. 1 1 MODEL OF COWDUNG GAS PLANT __: I.A.R.I. E , it\ ~ o ,, ,,., rail , . . . ~ . . . ~ 1 A BRICK WALL B GAS HOLDER C I RON ROD D PULLEY E COWDUNG INLET PIPE F SLURRY EXIT CHANNEL G COWDUNG MIXING TANK H DRYING BED I ANGLE IRON POSTS J GAS OUTLET PIPE K COUNTERPOISE WEIGHTS L GROUND LEVEL M EARTH PLATFORM N GAS MOISTURE EXIT TAP O SLURRY LEVEL P F E RME NTATI ON TAN K Q PLATFORM R LEDGE S G AS COC K Source: National Academy of Sciences. 1977. Methane generation from human, animal, and agricultural wastes Washington, D.C.: National Academy of Sciences. P. 70. FIGURE 6.4 Biogas plant designed by Acharya, developed at Indian Agricultural Research Institute.

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114 MICROBIAL PROCESSES Methane can be separated from other contaminating gases, such as carbon dioxide and hydrogen sulfide, and burned to generate electricity and heat and to produce steam, or for cooking. Biogas can be added as a supplement to natural gas pipelines, stored in tanks under atmospheric pressure, or com- pressed for storage. Capital and maintenance costs of biogas production units vary with the size of the plant, which is a function of local resources and needs. Table 6.3 shows some cost estimates for various sizes of methane plants in India. TABLE 6.3 Capacities and Costs of Biogas Plants Daily Production Rate (ma) 6 10 20 45 85 140 Estimated Cost in 1975 (in Rupees) 3,016 4,175 6,100 11,500 20,740 38,800 58,000 Approximate Head of Cattle or Swine Required 3-4 6-10 16-20 35-40 60-70 1 10~140 400-450 Source: National Academy of Sciences. 1977. Methane generation from human, animal, and agricultural wasted Washington, D.C.: Na- tional Academy of Sciences. P. 120. Limitations Temperature is a very important factor in biogas production. The rate of fermentation decreases rapidly below 20C. Maximum rates are obtained at about 40C (optimum for many mesophilic bacteria), but for thennophilic bacteria the optimum temperature is near 60C. The most effective tempera- ture for operation of a given digester depends on a number of factors such as insulation, climate, rate of conversion of the substrate, and concentrations of nutrients and other substances formed from the breakdown of the substrate. If dissolved materials in the effluent from the fermentation are low and the substrate being used is high in dry matter, it is possible to recycle the effluent and thereby conserve heat and nutrients. If nutrient concentrations are high (as in animal wastes), recycling of effluent liquids is not practical, and effluent heat value can be conserved only through heat exchangers. The system must be maintained in the neutral range (pH 6.8-7.8) for optimum rates. The pH may vary according to concentrations of acids or ammonia in the reactor. The percentage of solids and the biodegradability of substrates in the feed are important. With a high proportion of solids, the energy requirements for mixing, which is essential for effective fermentation, may be too high and soluble substances such as ammonia or organic acids may reach toxic concentrations. On the other hand, dilute substrates require large digesters. Nutrient elements such as nitrogen, phosphorus, sulfur, and

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FUEL AND ENERGY 115 iron may need to be supplied in an appropriate form and concentration; many materials such as human and animal wastes (but not urban refuse) contain adequate quantities of these substances. The reaction time, or rate of substrate use measured as flow per volume of reactor per day, needs to be short for rapid methane production. But if the reaction time is too short, less organic material is degraded and less methane is produced. Bacteria are sensitive to abrupt changes in loading rate and retention time, and they lose the potential for rapid activity during long periods of inactivity when substrate is not added. An efficient, rapid methane-producing system requires continuous or semicontinuous feeding, without drastic changesin operating conditions. Because of heating and other demands, larger digesters are generally more economical. The suitability of organic substrates for methane production depends on many factors. For example, removal or addition of water from or to relatively wet or dry substrates may be expensive. The inherently higher water content of some substrates will increase transportation costs and require a larger reactor. Collection costs and incentives for collection may also be important. Organic sugars fermented anaerobically produce roughly equal volumes of methane and carbon dioxide. The CO2 reduces the energy content of the biogas. It may be removed by `'scrubbing" it by passing it through dilute alkali. Many organic materials such as cattle wastes, forages, and urban wastes contain large amounts of lignin, silicates, and waxes that are indigestible under anaerobic conditions. These substances may greatly reduce the extent of conversion of cellulosic materials to methane. Treatment to increase the fermentability of such materials may be expensive. Some substrates, such as urban refuse, require separation of nonorganic wastes like glass and metals, and some may require particle-size reduction for effective fermentation. Many effective substrates, including whole corn, alfalfa, sugarcane, and cassava, may be more economically used as food or animal feed. Residual sludge disposal may be a problem, and aqueous effluent may cause pollution. But the residue and effluent may also serve as substrates for the growth of algae, which in turn may be an effective substrate for the production of methane gas. Even at the farm or village level, the effective production of methane requires competent management, but this should not be a problem if ade- quate teaching and extension activities are available. Research Needs The following research and development efforts are needed to facilitate greater production and use of biogas: Research leading to effective, economically produced substrates with emphasis on those that are photosynthetically derived;

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116 MICROBIAL PROCESSES Development of more economical collection and processing proce- duresfor example, by dewatering or desaltingalong with economical means of increasing fermentability; Research on separation of the cellulose-lignin complex; Identification of economic uses for lignin and other residues; Investigation of the relationship of the chemical composition of sub- strates to their efficiency in conversion to methane; and Design of improved and less-expensive digesters or reactors and their components, for instance, solar energy and better heat exchangers, to reduce the energy required to maintain temperatures at which fermentation is most effective. Methanol Methane gas is produced in enormous amounts in some areas of the world as a by-product of oil recovery and refining. It is also found in mud at the bottom of marine and freshwater environments and as a product of anaerobic fermentation of organic wastes. Methane is often difficult to transport from areas where it is produced to parts of the world where it can be used as an energy source. It must either be distributed by pipelines or refrigerated at -162C for transport by tanker. Since tanker transport is too expensive and hazardous to justify in most cases, the gas is simply flared off in many oil fields. By contrast, methanol has a relatively high energy content per unit vol- ume, and its transport is less expensive. Methanol is used in many parts of the world for heating, lighting, cooking, and power. It also has potential as a nonpolluting industrial fuel. The conversion of methane to methanol as an intermediate in the oxida- tion of methane to carbon dioxide occurs in all methane-oxidizing bacteria. Methanol accumulates in small amounts in many pure culture fermentations. The use of inhibitors (iodoacetate, phosphate buffers, and EDTA) of the microbial enzyme methanol dehydrogenase has been demonstrated. These materials inhibit methanol utilization without preventing the preceding methane oxidation step. In addition, some work has been conducted on "leaky" mutants of methane oxidizers that excrete (rather than further oxi- dize) methanol. Yields are poor, however, even in the best cases. The bio- conversion is believed to involve the following reaction In all methane-oxid~z- ing bacteria: CH4 + O2 + XH2 ~ CH3OH + H2O +X Methane Methanol where XH2 is an agent that reduces one-half of the O2 molecule in the bio- chemical process. Thus, a reducing agent must be available, which may b

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FUEL AND ENERGY 117 obtained by concurrent oxidation of inexpensive substrates or, ideally, by biophotolysis of water. Limitations The biochemistry of methane oxidation was poorly understood until re- cently, and it is necessary to establish the energy balance of the system before the process can be evaluated. The methanol produced will necessarily be in an aqueous solution, and the economics of recovery by distillation may preclude its use as a fuel unless cheaper means are found. It may be more practical to use the methanol as a fermentation substrate for single-cell protein production than to distill it to obtain a fuel. Research Needs When improving the feasibility of methanol production by microbial pro- cesses, the following preliminary steps should be taken: Elucidation of the biochemical pathways of the processes; Identification of new microorganisms and mutants that have greater potential for producing methanol; and Exploration of the use of inhibitors of methanol oxidation. Hydrogen The enormous amounts of solar radiation that reach the earth's surface greatly exceed the world's foreseeable needs for energy. The use of solar energy to produce fuels as well as biomass directly is, therefore, an inviting technical challenge. All green plants and algae use solar radiation in the reduc- tion of CO2 by water. The oxygen is released as molecular oxygen and the concurrent reducing equivalents (hydrogen ions and electrons) are used to reduce carbon dioxide in order to synthesize cellular constituents. It is possible under certain conditions to modify this biological process and cause the production of hydrogen from the biologically produced reducing equiv- alents. Biophotolysis, the production of hydrogen from water using the radiant energy of sunlight, has been demonstrated in a large number of algal cultures. It is theoretically possible to produce hydrogen from water using any plant or algae that contains the hydrogenate enzyme. The use of algae as a means of trapping solar energy to expand fuel supplies is attractive for reasons other than its potential as a cheap energy

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l 118 MICROBIAL PROCESSES alternative. Hydrogen as a fuel is nonpolluting. The substrate, water, and the energy source, sunlight, are inexhaustible. Many algae have simple nutritional requirements and can be cultivated on dilute waste materials. They can potentially be utilized as a food or fertilizer after use as a catalyst for hydrogen production. Algae as catalysts for the process are easily renewed and can perhaps be preserved in an active state. The most important considerations for future energy processes using algae include: 1) identification of strains that produce molecular hydrogen at the highest rates and use radiant energy most efficiently, and 2) genetic manipula- tion of these organisms and alteration of their metabolic processes to increase hydrogen production. A partial list of algae capable of evolving hydrogen is shown in Table 6.4. TABLE 6.4 Algae Capable of Evolving Hydrogen Scenedesmus obligaus Scenedesmus quadricauda Chlorella vulgaris Chlorella fusca Chlorella autotrophica Chlamydomonas moewasii Chlamydomonas debaryana Chlamydomonas dysosmos Chlamydomonas humicola Chlamydomonas reinhardii Ankistrodesmus brauni Ankistrodesmus stipitatum Dunaliella sp. Chondrus crispus Corallina officinals Ceramium niobium Porphyridium aerugineum Hydrogen production has been demonstrated from cell fractions of algae supplemented with essential enzymes. The major theoretical advantages of using this approach, which requires purification of all the fractions necessary for biophotolysis, would be a reduced system size for trapping light energy and improved efficiency. Isolated cell fractions, on the other hand, are re- newed with difficulty and present significant technical problems in storage and preservation of catalytic activity. The production of hydrogen by fermentation of organic substrates using nonphotosynthetic microorganisms has been discussed as a possible process for energy production. Hydrogen is also an end-product of organic fermenta- tion by many anaerobic bacteria. This process as an alternative to methane production has little value for energy conservation. The maximum amount of energy that can be conserved in the hydrogen produced by fermentation is 33 percent of the energy avail- able in the best substrates. The fermentation requires stringent precautions to ensure culture stability. By contrast, the energy conserved in the methane fermentation can exceed 80 percent of the energy available in organic matter, and this fermentation process has much greater potential for efficiency and economy.

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FUEL AND ENERGY Limitations 119 Biophotolysis for the production of hydrogen requires large reactors that are transparent to radiant energy and impermeable to hydrogen. The cost of any reactor must allow the process to be competitive with alternative energy sources. The production of hydrogen by most organisms is sensitive to oxy- gen, which must be rigorously excluded. The algae must be grown under adequate nutritional conditions, and the environment must be modified so that biomass production ceases and hydro- gen production proceeds. Hydrogen evolution can be accomplished by removal of carbon dioxide or nitrogen in some cultures. The optimum condi- tions will vary with the organism selected and other factors. Research Needs The present state of the technology for biophotolysis precludes its applica- tion in the near future. To further its potential, the following research should be undertaken: Examination of many kinds of algae for their potential as catalysts for hydrogen production; Determination of optimum conditions for efficient hydrogen produc- tion; and Research on microencapsulation and stabilization of sub cellular frac- tions capable of hydrogen evolution. Bacterial Leaching Thiobacillus ferrooxidans is a bacterium that lives in acid environments and obtains energy for growth by oxidizing reduced (ferrous) iron in various metal sulfides, sulfur, and soluble sulfur compounds. Many insoluble metal sulfides can be oxidized to corresponding metal sulfates. The oxidation of elements in pyrite ore can lead to the production of oxidized iron, sulfuric acid, and metal salts of sulfuric acid. The oxidized iron and sulfuric acid produced by the bacteria can be used for the extraction of uranium and other metals from raw ores. Large-scale leaching of uranium ores is employed in India, Canada, and the Soviet Union. In India, the leach liquor is percolated through a descending system of terraces containing pyritic uranium oxides. Exploitation of bac- terial leaching enables the recovery of uranium from low-grade ore (0.01-0.05 percent U3O~), uneconomical to process by other means. The process can

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120 MICROBIAL PROCESSES also be applied to high-grade material such as uranium-rich pillars supporting the roof of a worked-out mine. Recovery of uranium is accomplished by employing resin columns, and the oxidized iron in the liquid is recycled through the ore or slag. The cost of recovering uranium from low-grade ores by leaching is less than by conventional processes. In some cases, improved methods have been developed involving the production of acid ferric sulfate by a bacterial oxida- tion of pyrite, with the acid ferric sulfate used to leach ground ore. Continu- ous-culture methods for uranium leaching have also been described. The pro- cess with the greatest commercial potential is one that requires production of ferric iron leach liquid by a bacterial process and then utilizes the liquor for chemical leaching of uranium or other metals. The control of the bacterial populations is relatively easy because the acid content of the leach liquor and the substrates available for growth limit the kinds of organisms that can grow. These are practical, proven processes for recovering uranium that remain underexploited in many parts of the world. The attractions of leaching over conventional methods include its simplicity of operation, the lower capital requirements for materials, and lower energy costs. Bacterial leaching has also been applied to the recovery of hydrocarbons from oil shale. Research Needs Considerable pilot-scale research has been devoted to defining the ideal conditions for maximum rates of extraction of uranium and other metals. More attention needs to be given to scale-up of the extraction process. Leaching underground should be developed to obviate bringing ore to the surface. References and Suggested Reading Ethanol Anderson, Earl V. 1978. Gasohol: energy mountain or molehill? Chemical and Engineer- ingNews 56:8-12, 15. Cysewski, G. R., and Wilke, C. R. 1977. Rapid ethanol fermentation using vacuum and cell recycle. Biotechnology and Bioengineering 19:1125-1143. Faith, W. L.; Keys, D. B.; and Clark, R. L. 1974. Industrial chemicals. New York: John Wiley and Sons, Inc. Gall, Norman. 1978. Noah's ark: energy from biomass in Brazil. Report No. 30. Han- over, New Hampshire: American Universities Field Staff. Hepner, Leo. 1977. Feasibility of producing basic chemicals by fermentation. InMicro- bial energy conversion, H. G. Schlegel and J. Barnea, eds., pp. 531-553. Oxford: Pergamon Press

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FUEL AND ENERGY 121 Jackson, E. A. 1 976. Brazil's national alcohol program. Process Biochemistry 11: 29-30. Paturau, J. M. 1969. By-products of the cane sugar industry. New York: Elsevier-North Holland Publishing Company. Rao, M. R. K., and Murthy, N. S. 1963. Alcohol as a fuel for diesel engines. Paper Presented at the Symposium on New Developments of Chemical Industries Relating to Ethyl Alcohol, Its By-products and Wastes, 14-16 October, at New Delhi Utilization of Cellulose Cooney, C. L., and Wise, D. L. 1975. Thermophilic anaerobic digestion of solid waste for fuel gas production. Biotechnology and Bioengineering 17 :1119-1135. Ng, T. K.; Weimer, P. J. P.; and Zeikus, J. G. 1977. Cellulolytic and physiological properties of Clostridium thermocellum. Archives of Microbiology 1 14 :1-7. Weimer, P. J. P., and Zeikus, J. G. 1977. Fermentation of cellulose and cellobiose by Clostridium thermocellum in the absence and presence of A~Iethanobacterium thermo- autotrophicur~ Applied and Environmental hI crobiology 33:289-297. Methane Bryant, M. P. 1979. Microbial methane productiontheoretical aspects. Journal of Ani- malScience48:1. Golueke, C. G. 1977. Biological reclamation of solid wastes. Emmaus, Pennsylvania: Ro dale Press, Inc. Gould, R. F., ed. 1971. Anaerobic biological treatment processes. Advances in Chemistry Series, No. 105. Washington, D.C.: American Chemical Society. Jewell, W. J.; Davis, H. R.; Gunkel, W. W.; Lathwell, D. J.; Martin, J. A., Jr.; McCarty, T. R.; Morris, G. R.; Price, D. R.; and Williams, D. W. 1976. Biocon version of agricultural wastes for pollution control and energy conversion. Final Report TID 27164, for the U.S. Department of Energy under the National Science Foundation Contract No. ERDA-NSF-741222 A01. Ithaca, New York: Cornell University, Divi- sion of Solar Energy. National Academy of Sciences. 1977. Methane generation from human, animal, and agricultural wastes. Report of an Ad Hoc Panel of the Advisory Committee on Technology Innovation, Board on Science and Technology for International Develo~ meet, Commission on International Relations. Washington, D.C.: National Academy of Sciences. Pfeffer, J. T., and Liebman, J. C. 1976. Energy from refuse by big-conversion, fermenta- tion and residue disposal processes. Resource Recovery and Conversion 1:295. Schlegel, H. G., and Barr~ea, J., eds. 1976. Microbial energy conversion: Report of the United Nations Institute for Training and Research. Oxford: Pergamon Press. Van Soest, Peter J., and Mertens, D. P. 1974. Composition and nutritive characteristics of low quality cellulosic wastes. Federation Proceedings (published by Federation of American Societies for Experimental Biology) 33:1942-1944. Methanol Anthony, C. 1975. The biochemistry of methylotrophic microorganisms. Science Prog- ress (kondon) 62:167-206. Foo, E. L., and Heden, C.-G. 1977. Is biocatalytic production of methanol a practical proposition? In Microbial energy conversion, H. G. Schlegel and J. Barnea, eds., pp. 267-280. Oxford: Pergamon Press. Heden, C.~. 1974. Microbial aspects of the methanol economy. Annual Review of Microbiology 24:137-150. Ribbons, D. W.; Harrison, J. E.; and Wadsinski, A. M. 1970. Metabolism of single carbon compounds. Annual Review of Microbiology 24:135-158. Whittenbury, R. J.; Dalton, E. J.; and Reed, H. L. 1975. The different types of methane oxidizing bacteria and some of their more unusual properties. In Microbial growth on C-compounds, pp. 1-9. Kyoto, Japan: The Society of Fermentation Technology.

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122 MICROBIAL PROCESSES Hydrogen Gaffron, H., and Rubin, J. 1942. Fermentative and photochemical production of hydro- gen in algae. Journal of General Physiology 26: 219-240. Kondratieva, E. N. 1976. Phototrophic microorganisms as a source of hydrogenate for- mation. In Microbial energy conversion, H. G. Schlegel and J. Barnea, eds., pp. 205-216. Oxford: Pergamon Press. Oschepko, V. P., and Krawnovski, A. A. 1976. Photoproduction of molecular hydrogen by green algae. Akademiya Nauk S.S.R. Izvestiya. Seriya Biologicheskaya 87:100. Rao, K. K.; Rosa, L.; and Hall, D. O. 1976. Prolonged production of hydrogen gas by a chloroplast biocatalytic system. Biochemical Biophyszcal Research Communications 68:21-27. San Pietro, A. 1977. Hydrogen formation from water by photosynthesis and artificial systems. In Microbial energy conversion, H. G. Schlegel and J. Barnea, eds., pp. 217-233. Oxford: Pergamon Press. Stuart, T. S., and Gaffron, H. 1972. The mechanism of hydrogen production by several algae. Plan ta (An International Journal of Plant Biology) 106:101-112. Thauer, R. K; Jungermann, K.; and Deker, K. 1977. Energy conservation in chemo- trophic anaerobic bacteria. Bacterzologzical Review 41: 100-180. Bacterial Leaching Guay, P.; Silver, M.; and Torma, A. E. 1977. Ferrous iron oxidation and uranium extraction by Thiobacillus ferrooxidans. Biotechnology and Bioengineering 19:7 27- 740. Kelly, D. P. 1977. Extractions of metals from ores by bacterial leaching. In Microbial energy conversion, H. G. Schlegel and J. Barnea, eds., pp. 329-338. Oxford: Perga- mon Press. Meyer, W. Graig, and Yen, T. F. 1976. Enhanced dissolution of oil shale by bioleaching and thiobacilli Applied Environmental Microbiology 32 :610-613. Touvinen, O. H., and Kelly, D. P. 1974. Use of microorganisms for the recovery of metals. International Metallurgical Renew 19: 21-31. Warren Spring Laboratory. 1975. A bacterially assisted process for uranium extraction. Leaflet No. IME/75F/10c. Stevenage, Hertfordshire (England): Warren Spring Labo- ratory. Research Contacts Ethanol W. D. Bellamy. Department of Food Science, Cornell University, Ithaca, New York 14853, U.S.A. C. R. Wilke, Department of Chemical Engineering, Univexsity of California, Berkeley, California 94720, U.S.A. Utilization of Cellulose Charles L. Cooney, Department of Bioengineenng, Massachusetts Institute of Technol- ogy, Cambridge, Massachusetts 02139, U.S.A. J. G. Zeikus, Department of Bacteriology, University of Wisconsin, Madison, Wisconsin 53706, U.S.A. Methane Jerome Goldstein, Editor, Compost Science/Land Utilization, Box 351, Emmaus, Penn- sylvania 18049, U.S A. W. J. Jewell, Department of Agricultural Chemistry, Cornell University, Ithaca, New York 14850, U.S.A.

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FUEL AND ENERGY 123 P. L. McCarty, Department of Civil Engineering, Stanford University, Stanford, Califor- nia 94305, U.S.A. William J. Oswald, Division of Sanitary Engineering, University of California, Berkeley, California, 94720, U.S.A. J. T. Pfeffer, Department of Civil Engineering, University of Illinois, Urbana, Illinois 60801, U.S.A. Ram Bux Singh, Gobar Gas Research Station, Ajitmal, Etawah, Uttar Pradesh, India. R. J. Smith, Department of Agricultural Engineering, Iowa State University, Ames, Iowa 50010, U.S.A.