NZ625321B2 - Processing biomass for use in fuel cells - Google Patents

Abstract

Disclosed is a method of processing biomass by the saccharification of a lignocellulosic or cellulosic material (e.g., contacting the material with an enzyme) to produce a carbohydrate solution that can be used in a fuel cell, e.g., a direct glucose fuel cell, an indirect sugar fuel cell and a biological fuel cell. Particularly disclosed is method for producing electricity, the method comprising: providing a fuel to a fuel cell, wherein the fuel is produced by saccharification of a treated cellulosic or lignocellulosic material, and wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or lignocellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation. gical fuel cell. Particularly disclosed is method for producing electricity, the method comprising: providing a fuel to a fuel cell, wherein the fuel is produced by saccharification of a treated cellulosic or lignocellulosic material, and wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or lignocellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation.

Description

PROCESSING BIOMASS FOR USE IN FUEL CELLS RELATED APPLICATIONS This application claims priority to U.S. Provisional ation Serial No. 61/579,568, filed December 22, 2011, the complete disclosure therein incorporated herein by reference.

BACKGROUND Cellulosic and lignocellulosic materials (e.g., biomass materials) are produced, processed, and used in large quantities in a number of ations. Often such materials are used once, and then discarded as waste, or are simply considered to be waste materials, e.g., , bagasse, sawdust, and stover.

A typical biomass al contains cellulose, hemicellulose, and lignin plus lesser amounts of proteins, extractables and minerals. The complex carbohydrates ned in the cellulose and hemicellulose fractions can be sed into fermentable sugars by saccharification, using a cellulolytic enzyme, and the sugars can then be used as an end product or intermediate, or converted by further bioprocessing, e.g., tation, into a variety of products, such as alcohols or organic acids. The product obtained depends upon the microorganism utilized and the conditions under which the bioprocessing occurs.

This invention relates to methods of processing (e.g., saccharifying) carbohydrate-containing materials (e.g., biomass materials or biomass-derived materials) to produce sugar (e.g., glucose) solutions that can be used in fuel cells such as direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells. The invention also s to utilizing the carbohydrate-containing material derived sugars solutions in fuel cells.

In one aspect of the present invention there is ed a method comprising: utilizing, in a fuel cell, a sugar solution comprising saccharified cellulosic or ellulosic material, wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or ellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the al with ng radiation.

In another aspect of the present invention there is provided a fuel cell comprising: an electrode and saccharified cellulosic or lignocellulosic s, wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or lignocellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation.

In another aspect of the present invention there is provided a method for producing electricity, the method comprising: ing a fuel to a fuel cell, wherein the fuel is ed by saccharification of a treated cellulosic or lignocellulosic material, and wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or ellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation.

In some implementations, the sugar solution is produced by the saccharification of a lignocellulosic or cellulosic al, for example by contacting the al with an enzyme (e.g., a cellulase). In other implementations the recalcitrance of the lignocellulosic or cellulosic material has been reduced relative to [Text continued on page 2] that of the material in its native state prior to saccharification. In some cases, reducing the recalcitrance of the feedstock includes treating the feedstock with a treatment.

The treatment can be, for example, ionization radiation (e.g., electron beam radiation), sonication, pyrolysis, oxidation, steam explosion, chemical treatment, various mechanical treatments and combinations of any of these treatments. The physical treatment can comprise any one or more of the treatments disclosed herein, applied alone or in any desired combination, and applied once or multiple times. When treating with ionizing radiation, e.g., electron beam radiation, the dosage can be at least 10 Mrad and less than about 200 Mrad (eg., 50 Mrad to lSOMrad).

In some implementations, the method can include using an ve in the fuel cell. For example additives can be acids, bases, buffers (e.g., phosphate buffers), minerals, colloids, emulsions, emulsifiers, ulates, nano-particles, cations, anions, metal ions (e.g., Fe2+’ Fe3+, Mn2+, Cu2+, K+, Na+), ns, enzymes, peptones, extracts, surfactants, nutrients, gases (e.g., en, en, helium, argon, carbon de, carbon e) sources (e.g., ammonia, urea), , chemicals, nitrogen pigments, fragrances, anionic polymers, cationic polymers, non-ionic polymers, oligomers, lipids, fats, surfactants, dispersants, anti-foam agents, bacteriostatic , antimicrobial agents, microorganisms, viscosity modifiers, oxidizing agents (e.g., peroxides, chlorates), reducing agents, anti-scale agents, ion inhibitors, antifouling agents, precipitating agents, coagulants added in any combination and sequence.

A typical biomass ce contains cellulose, hemicellulose, and lignin plus lesser amounts of proteins, extractables and ls. In some implementations, cellulosic or lignocellulosic material includes paper, paper products, paper waste, paper pulp, pigmented papers, loaded papers, coated papers, filled papers, magazines, printed matter, printer paper, polycoated paper, card stock, cardboard, paperboard, offal, cotton, wood, particle board, forestry wastes, sawdust, aspen wood, wood chips, grasses, switchgrass, thus, cord grass, reed canary grass, grain residues, rice hulls, oat hulls, wheat chaff, barley hulls, ltural waste, silage, canola straw, wheat straw, barley straw, oat straw, rice straw, jute, hemp, flax, , sisal, abaca, corn cobs, corn stover, n stover, corn fiber, alfalfa, hay, coconut hair, sugar sing residues, bagasse, beet pulp, agave bagasse, algae, seaweed, manure, sewage, arracacha, buckwheat, banana, , cassava, kudzu, oca, sago, sorghum, , sweet potato, taro, yams, beans, favas, lentils, peas, and mixtures of any of these.

The cellulosic or lignocellulosic material can be mechanically treated to reduce the bulk density of the cellulosic or lignocellulosic material and/or increase its surface area. In some implementations, the method includes mechanically treating the feedstock before and/or after reducing its recalcitrance. Mechanical treatments e, for example, cutting, g, e.g., hammermilling, pressing, grinding, shearing and chopping. For example, comminuting the biomass material can be effective treatment d to the biomass material. Mechanical treatment can reduce the bulk density of the feedstock and/or increase the surface area of the feedstock. In some ments, after mechanical treatment the material has a bulk density of less than 0.75 g/cm3, e.g., less than about 0.7, 0.65, 0.60, 0.50, 0.35, 0.25, 0.20, 0.15, 0.10, 0.05, or less, e.g., less than 0.025 g/cm3. Bulk density is determined using ASTM D1 895B.

In one aspect, the invention features utilizing, a sugar solution d from a cellulosic or lignocellulosic material as described above in a fuel cell, e.g., a direct sugar fuel cell, an indirect sugar fuel cell and/or a biological fuel cell. Optionally the fuel can be a sugar or an alcohol derived from the saccharification of the osic or lignocellulosic material.

Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.

DESCRIPTION OF GS is a diagram illustrating the enzymatic hydrolysis of cellulose to is a flow diagram illustrating conversion of a biomass feedstock for use in a fuel cell. is a m showing various processes using saccharified feedstock in fuel cells. is a diagrammatic view of an example of a direct glucose fuel cell. is a diagrammatic view of an example of an ct sugar fuel cell. is a diagrammatic view of a half-cell of a biological fuel cell.

DETAILED DESCRIPTION Referring to during saccharification, a cellulosic or lignocellulosic substrate is initially hydrolyzed by endoglucanases at random locations producing oligomeric ediates. These intermediates are then substrates for litting glucanases such as cellobiohydrolase to produce cellobiose from the ends of the cellulose polymer. Cellobiose is a water-soluble 1,4-linked dimer of glucose. Finally cellobiase cleaves iose to yield glucose. The glucose, or other sugars, d from saccharification can be utilized in a fuel cell as will be described in detail herein.

Referring to a process for manufacturing sugar solutions for incorporation into a fuel cell system can include, for example, optionally mechanically treating a cellulosic and/or lignocellulosic feedstock (step 110). Before and/or after this treatment, the feedstock can optionally be treated with another physical treatment, for example irradiation, to reduce or r reduce its recalcitrance (step 112). A sugar solution is formed by rifying the feedstock (step 114) by, for example, the addition of one or more enzymes (step 111).

Optionally, the method may also include transporting, e. g., by pipeline, railcar, truck or barge, the sugar on (or the ock, enzyme and water, if saccharification is performed en route) to a manufacturing plant (step 116). For example, the methods of transporting and sing biomass as discussed in U.S. Patent 8,318,453 filed Jan 21, 2009 can be used herein; the te disclosure of which is orated herein by reference. If desired, the steps of measuring lignin content (step 118) and g or adjusting process parameters based on this measurement (step 120) can be performed at various stages of the process, for example, as described in U.S. Application Number 12/704,519, filed on February 11, 2011, the complete sure ofwhich is incorporated herein by reference. The sugar solution is then incorporated into a fuel cell or fuel cell system (122). Optionally, products produced in the process can be r processed and/or modified, for example, if sugars from the process are fermented to products, the product can be purified, for example by lation (124).

The products produced in these processes can also be utilized in a fuel cell system. is a diagram showing processes using saccharified feedstock in fuel cells. The sugar solution from the saccharified feedstock can be used in a direct sugar fuel cell, an indirect sugar fuel cell or a biological fuel cell. Glucose and xylose are often the most abundant sugars available from biomass and sugar solutions derived from biomass can include a mixture of xylose and glucose in various ratios. For example, only e can be present or only xylose can be present, especially in cases when the sugars have been isolated and/or d. When desired, other ratios can be utilized, for example as a percent of total glucose and xylose, the amount of glucose can be between 100% and 90%, 90% and 80%, 80% and 70%, 70% and 60%, 60% and 50%, 50% and 40%, 40% and 30%, 30% and 20%, 20% and 10%, 10% and 0%. gh, the e and xylose are often abundant biomass derived sugars, often providing more than 10 wt. % of the sugar to biomass (e.g., more than 20 wt.%, more than 30 wt.%, more than 40 wt.%, more than 50 wt.%, more than 60 wt.%, or even more than 70 wt.%) and are useful in these different fuel cells, other sugars and polysaccharides can also be useful. For example, arabinose, mannose, galactose and rhamnose, cellulose, starch, xylan, glucuronoxylan, arabinoxylan, glucomannan and xylogulcan can be used. Mixtures of any of these sugars can be utilized. In addition the sugars described herein can be isomerized (e.g., to fructose) and then used in a fuel cell. These different fuel cells and their use are discussed in more detail below.

Direct sugar fuel cells, for example glucose fuel cells, generally include a cathode electrode, an anode electrode, and a separator (e.g., an anion-exchange membrane (AEM) or a diffusion layer.) The fuel cell may be acidic or ne. In the e shown in an AEM is sandwiched between an anode electrode and a cathode electrode, with flow fields being provided between each of the electrodes and the AEM. In some cases, the cell has a two-cylinder uction in which one ode (e. g., the anode) is in the form of an inner cylinder and the other (e.g., the cathode) is in the form of an outer cylinder.

The active component of the anode may be, for example, PdNi or Pd—Pt, and the active component of the cathode may be, for example, a combined catalyst of cobalt porphyrin x (CoTPP) and spinel (MnC0204) or other suitable catalyst.

In the embodiment of a direct sugar fuel cell shown in a fuel solution containing glucose and, generally, potassium hydroxide (KOH), is fed into the anode flow channel, e.g., by a peristaltic pump (not shown), while oxygen is fed to the cathode flow field. Glucose is oxidized at the anode and the reduced product flows away through an anode exit channel. The electrons flow from the anode and through a load. Oxygen is d at the e and exhaust gas is vented from the cathode flow field. 2012/070624 Direct sugar fuel cells may or may not completely oxidize the sugar fuel to carbon dioxide and water while generating electricity. For example, the reaction for the total oxidation of glucose as shown here may occur.

Anode reaction: C6H1206 +24OH' 9 18H20 + 6C02 + 18e- Cathode on: 602 + 12H20 + 24e- 9 24OH' Overall on: C6H1206 + 602 9 6C02 + 6H20 AH = -2830 KJ/mol Since the total oxidation of e proceeds through many intermediates (e.g., gluconic acid, glucaric acid, gamma-gluconolactone, gamma-glucaro e, 2- ketogluconic acid, arabinose, trihydroxyglutaric acid, tartaric acid, hydroxyl malonic acid and oxalic acid) any of these intermediates can also be used in a fuel cell. Any of these intermediates, if produced by some of the processed described herein, (e.g., saccharification, fermentation) can be useful in generating electricity in a fuel cell.

Optionally, any sugar products not used in generating electricity in the direct sugar fuel cell can be further processed as shown in for example they can be ted to an alcohol and the l isolated by distillation. In some cases the fuel cell can use one sugar, for example glucose, and does not use other sugars, for example xylose, and the second sugar can be used in subsequent processes. In some other instances, a process, for example fermentation, only uses one sugar (e.g., glucose) leaving other sugars which can be then used in a fuel cell. In many cases, only a partial oxidation of glucose occurs in a direct sugar fuel cell. For example, the oxidation of glucose to gluconic acid occurs quickly providing 2 electrons and releasing at most about 200 KJ/mol of energy (e.g., about 7% of the available energy).

In terms of usable energy, the fuel cells convert at least 1% of the fuel to electric energy (e.g., at least 5%, at least 7%, at least 10%, at least 14%, at least 20%, at least %, at least 30 %, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%). In some cases between about 1% and 90% of the fuel is converted to electric energy (e.g., between about 1% and 70%, between about 1% and 50%, n about 1% and 20 %, between about 1% and 10%, n about 5% and 90%, n about 5% and 70%, between about 5% and 50%, between about 5% and 25%, between about 5% and 10%, between about 10% and 90%, between about % and 70%, between about 10% and 50%, between about 10% and 30%, between about 10% and 20%, between about 20% and 70%).

In addition to sugars and products such as alcohols, the solutions derived from biomass by the processes described herein can include various solids or dissolved compounds and/or materials. For example the solutions can include enzymes (e.g., parts of enzymes, active enzymes, denatured enzymes), amino acids, nutrients, live cells, dead cells, cellular debris (e.g., lysed cells, yeast extract) salts , acids, bases, (e.g., halides, sulfates, and phosphates, alkai, alkali earth, transition metal salts), partial hydrolysis products (e.g., cellulous and hemicellulose fragments), , lignin residues, inorganic solids (e.g., siliceous als, clays, carbon black, ), remnants of saccharified biomass and ations thereof.

Enzymes that can be present can be the intact or denatured enzymes utilized in the sing, or derived from these enzymes (e. g., proteins and amino acids). These can be dissolved/and or precipitated and suspended solids. For example, the sugar ons can have up to about 10 wt.% enzymes (e.g., up to 9 wt. %, up to 8 wt. %, up to 5 wt. %, up to 2 wt. %, up to 1 wt. %, between about 0.1 and 5 wt. %, between about 1 wt. % and 5 wt. %, between about 2 wt. % and 5 wt. %, between about 0.1 wt. % and 1 wt. %, between about 0.01wt.% and 1 wt.%, between about 0.001 wt.% and 0.1 wt.%).

During rification of a biomass, an optimal pH can often be in the acidic region and therefore the solutions used, if used directly in a fuel cell system, can have a pH n about 2 and 5 (e.g., between about 4 and 5). The pH can also be adjusted up or down after saccharificaion and or saccharification done at a higher or lower pH . In some embodiments the solution used in the fuel cell can therefore have pH values selected from a broad range. For example, the pH can be selected from a range of about 2 to about 10 (e.g., between about 2 and 5, between about 3 and 5, between about 3 and 6, between about 4 and 6, n about 5 and 10, between about 6 and 10 between about 8 and 10).

The sugar ons derived from the processes described herein and used in fuel cells systems can include non-sugar suspended or dissolved solids present at concentrations up to about 50 wt.%, for e between about 1 and 50 wt.%, 2 and 40 wt. %, 3 and 25 wt.%, 5 and 25 wt.%, 40 and 50 wt.%, 30 and 40 wt.%, 10 and 20 wt.%, 1 and 5 wt.%, 10 and 40 wt.%, less than about 50 wt.%, less than about 40 wt.%, less than about 30 wt.%, less than about 20 wt.%, less than about 10 wt.%, less than about 5 wt.%, less than about 1 wt.%, less than about 0.5 wt.%, less than about 0.01 wt.%. These solutions can have high turbidity, for example at least about 5 nephelometric turbidity units (NTU) (e. g., at least abouth NTU, at least about 50 NTU, at least abouthO NTU, at least about 100 NTU at least about 200 NTU, at least about 300 NTU, at least about 400 NTU and even greater than about 500 NTU).

In some cases the solids are tely or partially removed prior to being use in the fuel cells. For example the solids can be removed by filtration, centrifuging, settling, floatation and combinations of these. In some cases the solids are derived from a previously soluble material that has been precipitated, for example an enzyme that has been red. After removing the solids the turbidity of the solutions can be reduced by up to about 500 NTU (e.g., reduced by up to about 100 NTU, reduced by up to about 50 NTU, reduced by up to about 5 NTU).

In addition to being turbid, the sugar solutions produced from the processes described herein can be d due to colored impurities. For example some metal ions and polyphenols and lignin derived products produced or released during the processing of a lignocellulosic s can be highly colored. The solutions can be used ly in the fuel cell systems described herein or can be lly or completely decolorized prior to being used. For example the colored impurities can be filtered out of the solution, destroyed (e.g., by chemical decomposition) and/or precipitated out of the solution.

The ionic strength of the s derived sugar solutions can be high due to the source of the biomass as well as the processing of the biomass as described herein.

The solutions can be used directly or fully, selectively or partially de-ionized prior to being used in the fuel cell s described herein.

In some embodiments the fuel cell can include biomass (e. g., lignocellulosic biomass, cellulosic s, starch) as well as a rifying enzyme. For example, sugar can be utilized in a fuel cell system while it is being produced by the action of a saccharifying enzyme on a biomass material.

In yet other embodiments the sugar solutions used in the fuel cells herein described can include an additive. Such additives can modify properties of the solutions such as the pH, viscosity, chemical potential, e tension, thermal properties, color, odor, opacity, ionic strength, conductivity, sterility and/or nutrient value. For example additives can be acids, bases, buffers (e.g., phosphate buffers), minerals, colloids, emulsions, fiers, particulates, nano-particles, cations, anions, metal ions (e.g., Fe2+’ Fe3+, Mn2+, Cu2+, K+, Na+), vitamins, enzymes, peptones, extracts, surfactants, nutrients, gases (e.g., hydrogen, en, helium, argon, carbon monoxide, carbon dioxide), chemicals, nitrogen sources (e.g., ammonia, urea), pigments, fragrances, anionic polymers, ic polymers, non-ionic rs, oligomers, lipids, fats, tants, dispersants, anti-foam agents, bacteriostatic agents, antimicrobial agents, microorganisms, viscosity rs, oxidizing agents (e.g., peroxides, chlorates), reducing agents, anti-scale agents, corrosion inhibitors, antifouling agents, precipitating agents, coagulants added in any combination and sequence. ves can be added in ranges from a few parts per million to several percents. For example 1 to 1000 ppm, 5 to 500 ppm, 5 to 100 ppm, 50 to 100 ppm, 100 to 1000 ppm. 1 to 10 wt.%, 2 to 10 wt.%, 5 to 10 wt.%, 2 to 5 wt.%). In some embodiments including cations, anions, metal anions the amounts are between 1 to 1000ppm. In some embodiments where acids, bases or s are added, the final pH after addition of the additive can be chosen to be between pH 2 and 10, (e.g., between about 4 and 8, between about 5 and 7, between about 6 and 8, between about 4 and 5, between about 7 and 8, between about 8 and 10 and between about 2 and 4).

Additives can also be metered and added in amounts between about 1 uM to 5 M amounts (e.g., between about 1 mM and l M, between about 5 mM and 100 mM, between about 100 mM and 1 molar, n about 10 mM and 100mM). is a diagrammatic view of an example of an indirect sugar fuel cell. lly, the indirect sugar fuel cell uses a biological s to convert a primary fuel to a secondary fuel and the secondary fuel generates a current using a fuel cell.

The primary fuel (1) is brought into t with a bio-component (2) where it produces a secondary fuel (3) and waste (4). The secondary fuel is brought into the fuel cell and comes into contact with the anode (5) where it is oxidized, producing a reduced product (8), releasing an on to an external circuit, and providing a proton. The proton travels in the fuel cell through an ion selective membrane (6) to the cathode (7). Oxygen is supplied to the cathode where it is reduced by ons supplied from the external circuit and combines with the proton producing water. In another possible design, the bio-component resides within the fuel cell, so that the production of t and electricity all occur within the fuel cell. In some cases, the ion selective membrane is also not required. In other cases the oxidant may be oxidants other than dioxygen (e.g., hydrogen peroxide, organic peroxides and inorganic peroxides).

The y fuel used in the indirect fuel cell can be sugars (e.g., glucose and xylose) as well as polysaccharides that can be ed through the saccharification ofbiomass as previously discussed. The secondary fuel can be a fermentation product ofthe primary fuel. For example, the secondary fuel can be an alcohol or other fermentation product (e.g., ethanol, methanol, butanol, polyols, acetic acid, lactic acid and H2 ). Generally, the primary and secondary fuels can be ed from the intermediates and products discussed below. The bio-component can be a microbial material including but are not limited to, any naturally occurring or genetically modified microorganism or organism, for example, protists, e.g., animal protists (e.g., protozoa such as flagellates, amoeboids, ciliates, and sporozoa) and plant protists (e.g., algae such alveolates, chlorarachniophytes, cryptomonads, euglenids, glaucophytes, haptophytes, red algae, stramenopiles, and viridaeplantae). Other examples include seaweed, plankton (e.g. , macroplankton, mesoplankton, microplankton, nanoplankton, picoplankton, and femptoplankton), phytoplankton, bacteria (e.g., gram positive bacteria, gram ve bacteria, and extremophiles), human cells, mammalian cells, yeast and/or mixtures of these. There may be several bio-components, for example there may be several bacteria specialized to te ent ts useful for producing a current from different or the same components ofthe fuel. For e, fermentation methods and ting organisms discussed herein can be utilized to produce the secondary t.

Some species of microorganisms that can be used to produce the secondary fuel in the indirect sugar fuel cell are: idium saccharobulylacetonicum, Clostridium saccharoperbulylacetonicum, Clostridium saccharobulylicum, Clostridium um, Clostridium beijernckii, Clostridium acetobulylicum, idium aurantibulyricum, Clostridiumfelsineum, Clostridium bulyricum, Geobacter species, s of the genus Sacchromyces spp. e.g., Sacchromyces cerevisiae ’s , Saccharomyces icus, Saccharomyces uvarum, strains ofthe genus Kluyveromyces, e.g., species Kluyveromyces mamianus, Kluyveromyces fragilis, strains of the genus Candida, e.g., Candida pseudotropicalis, and Candida brassicae, Pichia stipitis (a relative of Candida shehatae), strains of the genus Clavispora, e.g., species Clavispora lusitaniae and Clavispora opuntiae, strains of the genus olen, e.g., species Pachysolen tannophilus, and strains of the genus Bretannomyces, e.g., species Bretannomyces clausem'i (Philippidis, G. P., 1996, Cellulose bioconversion technology, in Handbook on Bioethanol: Production and Utilization, Wyman, C.E., ed., Taylor & Francis, Washington, DC, 2).

Commercially available yeasts the may be used for the formation of secondary fuel include, for example, Red Star®/Lesaffre Ethanol Red (available from Red Star/Lesaffre, USA) FALI® (available from Fleischmann’s Yeast, a division of Burns Philip Food Inc., USA), SUPERSTART® (available from Alltech, now Lalemand), GERT STRAND® (available from Gert Strand AB, Sweden) and FERMOL® (available from DSM Specialties).

Biological fuel cells are devices capable of ly transforming a chemical fuel to electrical energy via electrochemical reactions involving biochemical ys. Generally this involves enzymes or active parts of enzymes for catalysis.

The enzymes can be within a living organism (e.g., microbial fuel cells) or can be outside of a living cell (e.g., enzyme fuel cells). shows a diagrammatic view of a generalized half-cell for a biological fuel cell. A supplied fuel is contacted with a ical component which oxidizes the fuel and s waste that is removed. The electrons released from the fuel are transferred from the biological component to a mediator which either diffuses to or is associated with the electrode where the mediator is oxidized to its original state releasing an electron to an external circuit.

The oxidant side of the fuel cell is not shown. Some oxidants can be, for example, 02, ed by air, or peroxides (e.g., hydrogen peroxide, organic peroxides, inorganic peroxides). Some biological fuel cells do not require a mediator; in such cells the electron is erred directly from the biological component to the electrode. In some biological fuel cells the anode reaction with oxygen is zed by a biological component. Some biological fuel cells have been described, for example by Derek Lovely in “The microbe ic: conversion of organic matter to electricity”, Current Opinions in Biotechnology, 2008, Volume 19, pages 1-8, or in US. patent No. 8,283,076 filed May 18, 2007; the entire sure in these references are orated by reference herein.

The fuel used in the biological fuel cell can be the rification products from biomass as previously discussed. Especially in cases where organisms are utilized (e.g., ial fuel cells) other nutrients and media components can be added to the fuels, for example ions (e.g., sodium, potassium, iron, magnesium, manganese, zinc, copper), ates, sulfates, ammonia, urea, amino acids, proteins, ns, buffers, organic acids, nic acids, organic bases, inorganic bases, nutrient rich extracts(e.g., yeast extracts, meat extracts and vegetable extracts). Additionally the temperature and pH can be controlled. For example atures between 10 and 70 °C can be used (e.g., between about 10 and 60 °C between about 10 and 50 °C, between about 10 and 40 °C, between about 20 and 70 °C, between about 20 and 60 °C, n about 20 and 50 °C, between about 20 and 40 °C, between about 30 and 70 °C, between about 30 and 60 °C, between about 30 and 50 °C, between about 30 and 40 oC). The pH can be between about 3 and 10 (e.g., between about 3 and 9, between about 3 and 8, n about 3 and 7, between about 3 and 6, between about 3 and 5, between about 4 and 9, between about 4 and 8, between about 4 and 7, between about 5 and 9, between about 5 and 8, between about 5 and 7). es of organisms that can be useful in biological fuel cells are species of geobacter, proteus vulgaris, Desulphovibrio huricans, E. coli, Actinobacillus succinogenes, Desulphovibrio vulgaris, Shewanella putrefaciens and Rhodoferax ferrireducens.

Examples of enzymes that can be useful in ical fuel cells are glucose oxidase, alcoholdehydrogenase, dedehydrogenase, formate dehydrogenase, oxidoreductase, diaphorase, flavor-oxido-reductase, laccase, microperoxidase, glucose dehydrogenase, hydrogenase, peroxidases, reconstituted enzymes from this list and combinations thereof.

MECHANICAL TREATMENTS Biomass feedstock (e.g., cellulosic and/or lignocellulosic material) can be mechanically d prior to or after other ents. s of mechanically ng the carbohydrate-containing material include, for example, milling or grinding. Milling may be performed using, for example, a hammer mill, ball mill, colloid mill, conical or cone mill, disk mill, edge mill, Wiley mill, grist mill or other mills. Grinding may be performed using, for example, a cutting/impact type grinder.

Some exemplary grinders include stone grinders, pin grinders, coffee grinders, and burr grinders. Grinding or milling may be provided, for example, by a reciprocating pin or other element, as is the case in a pin mill. Other mechanical treatment methods include mechanical ripping or tearing, other methods that apply pressure to the fibers, and air attrition milling. Suitable mechanical treatments further include any other technique that ues the disruption of the internal structure of the material that was initiated by the previous processing steps.

Mechanical feed preparation systems can be configured to produce streams with specific characteristics such as, for example, c maximum sizes, specific length-to-width, or specific e areas ratios. Physical preparation can increase the rate of reactions, improve the movement of material, e the irradiation profile of the material, improve the radiation uniformity of the material, or reduce the processing time required by opening up the als and making them more accessible to ses and/or reagents, such as reagents in a solution.

The bulk density of feedstocks can be controlled (e.g., increased). In some situations, it can be ble to prepare a low bulk density material, e.g., by densifying the material (e.g., densification can make it easier and less costly to transport to another site) and then reverting the material to a lower bulk density state (e.g. , after transport). The material can be densified, for example from less than about 0.2 g/cc to more than about 0.9 g/cc (e.g., less than about 0.3 to more than about 0.5 g/cc, less than about 0.3 to more than about 0.9 g/cc, less than about 0.5 to more than about 0.9 g/cc, less than about 0.3 to more than about 0.8 g/cc, less than about 0.2 to more than about 0.5 g/cc). For example, the material can be densified by the methods and equipment disclosed in U.S. Pat. No. 7,932,065 to Medoff and International Publication No. (which was filed October 26, 2007, was published in English, and which designated the United States), the full disclosures of which are incorporated herein by reference. Densified materials can be processed by any of the methods described herein, or any material processed by any of the methods bed herein can be subsequently densified.

In some embodiments, the material to be sed is in the form of a fibrous al that includes fibers provided by shearing a fiber source. For example, the shearing can be performed with a rotary knife .

For example, a fiber source, e.g., that is recalcitrant or that has had its recalcitrance level reduced, can be sheared, e.g., in a rotary knife cutter, to provide a first fibrous material. The first fibrous material is passed through a first , e.g., having an average opening size of 1.59 mm or less (1/16 inch, 0.0625 inch), provide a second fibrous material. If desired, the fiber source can be cut prior to the shearing, e.g., with a er. For example, when a paper is used as the fiber source, the paper can be first cut into strips that are, e.g., 1/4- to 1/2-inch wide, using a shredder, e.g., a counter-rotating screw shredder, such as those manufactured by Munson (Utica, N.Y.).

As an alternative to shredding, the paper can be reduced in size by cutting to a desired size using a guillotine cutter. For example, the tine cutter can be used to cut the paper into sheets that are, e.g., 10 inches wide by 12 inches long.

In some embodiments, the shearing of the fiber source and the passing of the resulting first fibrous material through a first screen are performed concurrently. The shearing and the passing can also be performed in a batch-type process.

For example, a rotary knife cutter can be used to concurrently shear the fiber source and screen the first fibrous material. A rotary knife cutter includes a hopper that can be loaded with a shredded fiber source prepared by shredding a fiber source.

In some implementations, the feedstock is physically treated prior to rification and/or fermentation. Physical treatment processes can include one or more of any of those bed herein, such as ical treatment, chemical treatment, ation, sonication, oxidation, pyrolysis or steam explosion. Treatment methods can be used in combinations of two, three, four, or even all of these technologies (in any order). When more than one treatment method is used, the methods can be applied at the same time or at different times. Other processes that change a molecular structure of a biomass feedstock may also be used, alone or in combination with the processes disclosed herein.

Mechanical treatments that may be used, and the characteristics of the mechanically treated feedstocks, are described in r detail in U.S. Serial No. 13/276,192, filed October 18, 2011, the full disclosure of which is hereby incorporated herein by reference.

In on to this size reduction, which can be performed lly and/or later during processing, mechanical treatment can also be advantageous for ng up,” “stressing,” breaking or shattering the feedstock materials, making the cellulose of the materials more susceptible to chain scission and/or disruption of lline structure during the structural modification treatment.

The biomass can be in a dry form, for e with less than about % moisture content (e.g., less than about 20 %, less than about 15 %, less than about 10 % less than about 5 %, less than about 4%, less than about 3 %, less than about 2 % or even less than about 1 %). The biomass can also be red in a wet state, for example as a wet solid, a slurry or a suspension with at least about 10 wt% solids (e.g., at least about 20 wt.%, at least about 30 wt. %, at least about 40 wt.%, at least about 50 wt.%, at least about 60 wt.%, at least about 70 wt.%).

The processes sed herein can utilize low bulk density materials, for example cellulosic or lignocellulosic feedstocks that have been physically pretreated to have a bulk density of less than about 0.75 g/cm3, e.g., less than about 0.7, 0.65, 0.60, 0.50, 0.35, 0.25, 0.20, 0.15, 0.10, 0.05 or less, e.g., less than about 0.025 g/cm3.

Bulk density is determined using ASTM D1895B. Briefly, the method es filling a measuring cylinder ofknown volume with a sample and obtaining a weight of the sample. The bulk y is calculated by dividing the weight of the sample in grams by the known volume of the cylinder in cubic centimeters. If desired, low bulk y materials can be densified, for example, by methods described in US. Pat. No. 7,971,809 to Medoff, the full disclosure of which is hereby incorporated by reference.

In some cases, the biomass can be screened through a mesh or perforated plate with a desired opening size, for example, less than about 6.35 mm (14 inch, 0.25 inch), (e.g., less than about 3.18 m (1/8 inch, 0.125 inch), less than about 1.59 mm (1/16 inch, 0.0625 inch), is less than about 0.79 mm (1/32 inch, 0.03125 inch), e.g., less than about 0.51 mm (1/50 inch, 0.02000 inch), less than about 0.40 mm (1/64 inch, 0.015625 inch), less than about 0.23 mm (0.009 inch), less than about 0.20 mm (1/128 inch, 0.0078125 inch), less than about 0.18 mm (0.007 inch), less than about 0.13 mm (0.005 inch), or even less than about 0.10 mm (1/256 inch, 0.00390625 .

Screening of biomass material can also be by a manual method, for example by an operator or mechanoid (e.g., a robot equipped with a color, reflectivity or other ) that removes unwanted material. ing can also be by magnetic screening wherein a magnet is disposed near the conveyed material and the magnetic material is removed magnetically.

Optionally, prior to further processing, the biomass material can be heated, for e by IR radiation, microwaves, combustion (e.g., gas, coal, oil, biomass), ive heating and/or inductive heating. Heating can be, for example, for the purpose of drying the material. In the case of drying the material, this can also be facilitated, with or without heating, by the movement of a gas (e.g., air, oxygen, nitrogen, He, C02, Argon) over and/or through the biomass.

WO 96452 ally, the s can be cooled prior to or after mechanical treatment.

Cooling material is described in US Pat. No. 7,900,857 to Medoff, the disclosure of which in incorporated herein by reference.

RADIATION TREATMENT In some cases, the feedstock may be irradiated to modify its structure and thereby reduce its recalcitrance. Irradiation may, for example, reduce the average lar weight ofthe feedstock, change the crystalline structure ofthe feedstock, change the fimctionalization ofthe ock (e.g., by oxidation) and/or increase the surface area and/or porosity of the feedstock.

Various other irradiating devices may be used in the methods disclosed herein, including field tion sources, electrostatic ion separators, field ionization generators, thermionic emission sources, microwave discharge ion sources, recirculating or static accelerators, dynamic linear rators, van de Graaff accelerators, and folded tandem accelerators. Such devices are sed, for example, in U.S. Pat. No. 7,931,784 to Medoff, the complete disclosure ofwhich is incorporated herein by reference.

A beam of electrons can be used as the radiation source. A beam of electrons has the advantages of high dose rates (e.g., l, 5, or even 10 Mrad per second), high hput, less containment, and less confinement equipment. Electron beams can also have high electrical efficiency (e.g., 80%), allowing for lower energy usage relative to other radiation methods, which can translate into a lower cost of operation and lower greenhouse gas emissions corresponding to the smaller amount of energy used. Electron beams can be generated, e.g., by ostatic generators, cascade generators, transformer generators, low energy accelerators with a scanning system, low energy accelerators with a linear cathode, linear rators, and pulsed accelerators.

Electrons can also be more efficient at causing changes in the molecular ure of carbohydrate-containing als, for example, by the mechanism of chain scission. In addition, electrons having energies of 0.5-10 MeV can penetrate low density materials, such as the biomass materials described herein, e.g., materials having a bulk density of less than 0.5 g/cm3, and a depth of 03-10 cm. Electrons as an ionizing radiation source can be useful, e.g., for relatively thin piles, layers or beds of materials, e.g., less than about 0.5 inch, e.g., less than about 0.4 inch, 0.3 inch, 0.25 inch, or less than about 0.1 inch. In some ments, the energy of each electron ofthe electron beam is from about 0.3 MeV to about 2.0 MeV (million electron volts), e.g., from about 0.5 MeV to about 1.5 MeV, or from about 0.7 MeV to about 1.25 MeV. s of irradiating materials are discussed in U.S. Pat. App. Pub. 2012/01000577 A1, filed October 18, 2011, the entire disclosure ofwhich is herein incorporated by reference.

Electron beam irradiation devices may be procured commercially from Ion Beam ations, Louvain-la-Neuve, Belgium or the Titan Corporation, San Diego, CA. Typical on energies can be 0.5 MeV, 1 MeV, 2 MeV, 4.5 MeV, 7.5 MeV, or 10 MeV. l electron beam irradiation device power can be 1 KW, 5 KW, 10 KW, 20 KW, 50 KW, 60 KW, 70 KW, 80 KW, 90 KW, 100 KW, 125 KW, 150 KW, 175 KW, 200 KW, 250 KW, 300 KW, 350 KW, 400 KW, 450 KW, 500 KW, 600 KW, 700 KW, 800 KW, 900 KW or even 1000 KW.

Tradeoffs in considering electron beam irradiation device power specifications include cost to operate, capital costs, depreciation, and device footprint. Tradeoffs in considering exposure dose levels of electron beam irradiation would be energy costs and environment, safety, and health (ESH) concerns. Typically, generators are housed in a vault, e.g. of lead or concrete, especially for production from X-rays that are generated in the process. Tradeoffs in considering electron energies include energy costs.

The electron beam irradiation device can produce either a fixed beam or a scanning beam. A scanning beam may be advantageous with large scan sweep length and high scan speeds, as this would ively replace a large, fixed beam width.

Further, available sweep widths of 0.5 m, 1 m, 2 m or more are available.

In some embodiments, two or more radiation sources are used, such as two or more ng radiation sources. For example, samples can be treated, in any order, with a beam of electrons, followed by gamma radiation and UV light having wavelengths from about 100 nm to about 280 nm. In some embodiments, s are treated with three ionizing radiation sources, such as a beam of ons, gamma radiation, and energetic UV light. The biomass is conveyed through the irradiation zone (354 in where it can be irradiated, for example by electrons. It is generally preferred that the bed of biomass material has a relatively m thickness, as previously described, while being irradiated. 2012/070624 Effectiveness of changing the lar/supermolecular structure and/or reducing the itrance of the carbohydrate-containing biomass depends on the electron energy used and the dose applied, while re time depends on the power and dose.

In some embodiments, the irradiating (with any radiation source or a combination of sources) is performed until the material receives a dose of at least about 0.05 Mrad, e.g., at least about 0.1, 0.25, 0.5, 0.75, 1.0, 2.5, 5.0, 7.5, 10.0, 15, 20, , 30, 40, 50, 60, 70, 80, 90, 100, 125, 150, 175, or 200 Mrad. In some embodiments, the irradiating is performed until the al receives a dose of between 0.1 - 100 Mrad, 1 - 200, 5 — 200, 10 — 200, 5 — 150, 50 - 150, 100 - 200, 100 - 150, 80 - 120, 5 — 100, 5 — 50, 5 — 40,10 — 50,10 — 75,15 — 50, 20 — 35 Mrad.

In some embodiments, the irradiating is performed at a dose rate of between .0 and 1500.0 kilorads/hour, e.g., n 10.0 and 750.0 kilorads/hour or between 50.0 and 350.0 kilorads/hours. In other embodiments the irradiation is performed at a dose rate ofbetween 10 and 10000 kilorads/hr, between 100 and 1000 kilorad/hr, or between 500 and 1000 ds/hr.

In some implementations, it is desirable to cool the material during irradiation.

For example, the material can be cooled while it is being conveyed, for example by a screw extruder or other conveying equipment.

Radiation can be applied while the cellulosic and/or lignocellulosic material is exposed to air, oxygen-enriched air, or even oxygen itself, or blanketed by an inert gas such as nitrogen, argon, or helium. When maximum oxidation is desired, an oxidizing environment is utilized, such as air or oxygen and the distance from the radiation source is optimized to maximize ve gas formation, e.g., ozone and/or oxides of nitrogen.

SONICATION, PYROLYSIS, OXIDATION, STEAM EXPLOSION If desired, one or more tion, pyrolysis, oxidative, or steam explosion processes can be used in addition to or instead of irradiation to reduce the recalcitrance of the feedstock. These processes are bed in detail in U.S. Serial No. 12/429,045, the full disclosure ofwhich is incorporated herein by reference.

SACCHARIFICATION The d biomass materials can be saccharified, generally by combining the al and a cellulase enzyme in a fluid medium, e.g., an aqueous solution. In some cases, the material is , steeped, or cooked in hot water prior to saccharification, as described in U.S. Pat. App. Pub. 1000577 A1, filed October 18, 2011.

The saccharification process can be partially or completely performed in a tank (e.g., a tank having a volume of at least 4000, 40,000, or 500,000 L) in a manufacturing plant, and/or can be partially or completely performed in t, e.g., in a rail car, tanker truck, or in a anker or the hold of a ship. The time required for complete saccharification will depend on the process conditions and the carbohydrate-containing material and enzyme used. If saccharification is performed in a manufacturing plant under controlled conditions, the cellulose may be substantially entirely converted to sugar, e.g., glucose in about 12-96 hours. If saccharification is performed partially or completely in transit, saccharification may take longer.

It is generally preferred that the tank contents be mixed during saccharification, e.g., using jet mixing as described in International App. No. , filed May 18, 2010, which was published in h as WC 2010/135380 and designated the United States, the full disclosure ofwhich is incorporated by reference herein.

The addition of surfactants can enhance the rate of saccharification. Examples of surfactants include non-ionic surfactants, such as a Tween® 20 or Tween® 80 polyethylene glycol surfactants, ionic surfactants, or amphoteric surfactants.

It is generally preferred that the concentration of the sugar solution ing from saccharification be vely high, e.g. than 40%, or r than 50, 60, , greater 70, 80, 90 or even greater than 95% by weight. Water may be removed, e.g., by evaporation, to increase the concentration of the sugar solution. This reduces the volume to be shipped, and also inhibits microbial grth in the solution.

Alternatively, sugar solutions of lower concentrations may be used, in which case it may be desirable to add an antimicrobial ve, e.g., a broad spectrum antibiotic, in a low tration, e.g., 50 to 150 ppm. Other suitable antibiotics include amphotericin B, ampicillin, chloramphenicol, ciprofloxacin, gentamicin, hygromycin B, kanamycin, neomycin, penicillin, puromycin, streptomycin.

Antibiotics will inhibit growth of rganisms during transport and storage, and can be used at appropriate concentrations, e.g., between 15 and 1000 ppm by weight, e.g., between 25 and 500 ppm, or between 50 and 150 ppm. If desired, an antibiotic can be ed even if the sugar concentration is relatively high. Alternatively, other additives with anti-microbial or preservative properties may be used. Preferably the antimicrobial additive(s) are food-grade.

A relatively high concentration solution can be obtained by limiting the amount ofwater added to the carbohydrate-containing material with the enzyme. The concentration can be controlled, e.g., by controlling how much saccharification takes place. For example, concentration can be increased by adding more carbohydrate- containing material to the solution. In order to keep the sugar that is being produced in solution, a surfactant can be added, e.g., one of those discussed above. Solubility can also be increased by increasing the temperature of the solution. For example, the solution can be maintained at a ature of 40-50°C, 60-80°C, or even higher.

SUGARS In the processes described herein, for example after saccharification, sugars (e.g., glucose and xylose) can be ed. For example sugars can be isolated by precipitation, crystallization, chromatography (e.g., ted moving bed chromatography, high pressure chromatography), centrifugation, extraction, any other isolation method known in the art, and combinations thereof.

FERMENTATION Yeast and Zymomonas ia, for example, can be used for fermentation or conversion of s) to alcohol(s). Other microorganisms are discussed below. The m pH for fermentations is about pH 4 to 7. For example, the optimum pH for yeast is from about pH 4 to 5, while the optimum pH for Zymomonas is from about pH 5 to 6. Typical fermentation times are about 24 to 168 hours (e.g., 24 to 96 hrs) with temperatures in the range of 20°C to 40°C (e.g., 26°C to 40°C), however thermophilic microorganisms prefer higher temperatures.

In some ments, e.g., when anaerobic organisms are used, at least a portion of the fermentation is conducted in the absence of , e.g., under a blanket of an inert gas such as N2, Ar, He, C02 or mixtures thereof. Additionally, the mixture may have a constant purge of an inert gas flowing through the tank during part of or all of the fermentation. In some cases, anaerobic condition, can be achieved or maintained by carbon dioxide production during the fermentation and no additional inert gas is needed.

In some embodiments, all or a portion of the fermentation process can be upted before the low molecular weight sugar is completely converted to a product (e.g., ethanol). The intermediate fermentation products include sugar and carbohydrates in high concentrations. The sugars and carbohydrates can be ed via any means known in the art. These intermediate fermentation products can be used in preparation of food for human or animal consumption. Additionally or alternatively, the intermediate fermentation products can be ground to a fine particle size in a ess-steel laboratory mill to produce a flour-like substance.

Jet mixing may be used during tation, and in some cases rification and tation are performed in the same tank.

Nutrients for the rganisms may be added during saccharification and/or fermentation, for example the food-based nutrient packages described in U.S. Pat.

App. Pub. 2012/0052536, filed July 15, 2011, the complete disclosure of which is incorporated herein by reference.

Mobile fermenters can be utilized, as described in Intemational App. No. (which was filed July 20, 2007, was published in English as and designated the United States), the contents ofwhich is incorporated herein in its entirety. Similarly, the saccharification equipment can be . Further, saccharification and/or fermentation may be performed in part or entirely during transit.

DISTILLATION After fermentation, the ing fluids can be distilled using, for example, a “beer column” to separate ethanol and other alcohols from the majority of water and residual solids. The vapor exiting the beer column can be, e.g. , 35% by weight ethanol and can be fed to a rectification . A mixture of nearly azeotropic (92.5%) l and water from the cation column can be purified to pure (99.5%) ethanol using vapor-phase molecular sieves. The beer column bottoms can be sent to the first effect of a three-effect evaporator. The rectification column reflux condenser can provide heat for this first effect. After the first effect, solids can be separated using a centrifuge and dried in a rotary dryer. A portion (25%) of the fuge effluent can be recycled to fermentation and the rest sent to the second and third evaporator effects. Most of the evaporator condensate can be returned to the process as fairly clean condensate with a small portion split off to waste water ent to prevent up of low-boiling compounds.

INTERMEDIATES AND PRODUCTS Using the processes bed herein, the biomass material can be converted to one or more products, such as energy, fuels, foods and als. Specific examples ucts include, but are not limited to, hydrogen, sugars (e.g., glucose, xylose, arabinose, mannose, galactose, fructose, disaccharides, oligosaccharides and polysaccharides), alcohols (e.g., monohydric alcohols or dihydric alcohols, such as ethanol, n-propanol, isobutanol, sec-butanol, tert-butanol or n-butanol), hydrated or hydrous alcohols (e.g., containing greater than 10%, 20%, 30% or even greater than 40% water), biodiesel, organic acids, hydrocarbons (e.g., e, ethane, propane, isobutene, pentane, n-hexane, biodiesel, bio-gasoline and mixtures thereof), co- products (e.g., proteins, such as cellulolytic proteins (enzymes) or single cell proteins), and mixtures of any of these in any combination or relative concentration, and ally in combination with any additives (e.g., fuel additives). Other es include carboxylic acids, salts of a carboxylic acid, a mixture of carboxylic acids and salts of carboxylic acids and esters of carboxylic acids (e.g., methyl, ethyl and n-propyl ), ketones (e.g., acetone), aldehydes (e.g., acetaldehyde), alpha and beta unsaturated acids (e.g., acrylic acid) and olefins (e.g., ethylene). Other alcohols and alcohol derivatives include propanol, propylene glycol, l,4-butanediol, 1,3- propanediol, sugar alcohols (e.g., erythritol, glycol, glycerol, sorbitol threitol, arabitol, ribitol, mannitol, dulcitol, fucitol, iditol, isomalt, maltitol, lactitol, xylitol and other polyols), and methyl or ethyl esters of any of these alcohols. Other products e methyl te, methylmethacrylate, lactic acid, citric acid, formic acid, acetic acid, propionic acid, butyric acid, succinic acid, valeric acid, caproic acid, 3- ypropionic acid, palmitic acid, c acid, oxalic acid, c acid, glutaric acid, oleic acid, linoleic acid, glycolic acid, gamma-hydroxybutyric acid, and mixtures thereof, salts of any of these acids, mixtures of any of the acids and their tive salts. Many of the products obtained, such as ethanol or n-butanol, can be utilized as a fuel for powering cars, trucks, tractors, ships or trains, e.g., as an internal combustion fuel or as a fuel cell ock. Many of the ts obtained can also be utilized to power aircraft, such as planes, e.g., having jet engines or helicopters. In on, the ts described herein can be utilized for electrical power generation, e.g., in a conventional steam generating plant or in a fuel cell plant.

Other ediates and products, including food and pharmaceutical products, are described in U.S. App. No. 12/417,900 filed April 3, 2009, the full sure of which is hereby incorporated by reference herein.

CARBOHYDRATE CONTAINING MATERIALS IOMASS MATERIALS As used herein, the term “biomass materials” is used interchangeably with the term "carbohydrate-containing materials”, and includes lignocellulosic, cellulosic, starchy, and microbial materials. Any of the methods described herein can be practiced with mixtures of any biomass materials described herein.

Lignocellulosic materials include, but are not d to, wood, particle board, forestry wastes (e.g., sawdust, aspen wood, wood chips), grasses, (e.g., switchgrass, miscanthus, cord grass, reed canary grass), grain residues, (e.g., rice hulls, oat hulls, wheat chaff, barley hulls), agricultural waste (e.g., silage, canola straw, wheat straw, barley straw, oat straw, rice straw, jute, hemp, flax, bamboo, sisal, abaca, corn cobs, corn stover, soybean stover, corn fiber, a, hay, coconut hair), sugar processing residues (e.g., bagasse, beet pulp, agave bagasse), , algae, seaweed, , sewage, and mixtures of any of these.

In some cases, the lignocellulosic material includes comcobs. Ground or hammermilled comcobs can be spread in a layer of relatively uniform thickness for irradiation, and after irradiation are easy to disperse in the medium for further processing. To facilitate harvest and collection, in some cases the entire corn plant is used, including the corn stalk, corn kernels, and in some cases even the root system of the plant.

Advantageously, no additional nutrients (other than a nitrogen source, e.g., urea or ammonia) are required during tation of comcobs or osic or lignocellulosic materials containing significant amounts of comcobs.

Comcobs, before and after comminution, are also easier to convey and disperse, and have a lesser tendency to form explosive mixtures in air than other cellulosic or lignocellulosic materials such as hay and grasses.

Cellulosic materials include, for example, paper, paper products, paper waste, paper pulp, pigmented papers, loaded papers, coated papers, filled , magazines, printed matter (e.g., books, catalogs, manuals, , calendars, greeting cards, brochures, prospectuses, newsprint), printer paper, polycoated paper, card stock, cardboard, paperboard, materials having a high oc-cellulose content such as cotton, and mixtures of any of these. For example paper products as described in U.S. App.

No. 13/396,365 (“Magazine Feedstocks” by Medoff et al., filed February 14, 2012), the full disclosure of which is incorporated herein by reference.

Cellulosic als can also include lignocellulosic materials which have been de-lignified.

Starchy materials include starch itself, e.g., corn starch, wheat starch, potato starch or rice , a derivative of starch, or a material that includes starch, such as an edible food product or a crop. For example, the starchy material can be arracacha, buckwheat, banana, barley, cassava, kudzu, oca, sago, m, regular household potatoes, sweet potato, taro, yams, or one or more beans, such as favas, lentils or peas.

Blends of any two or more starchy materials are also starchy materials. es of y, cellulosic and or lignocellulosic materials can also be used. For example, a biomass can be an entire plant, a part of a plant or ent parts of a plant, e.g., a wheat plant, cotton plant, a corn plant, rice plant or a tree. The starchy materials can be treated by any of the methods described herein.

Microbial materials include, but are not limited to, any naturally occurring or genetically modified microorganism or organism that contains or is capable of providing a source of carbohydrates (e.g., ose), for example, protists, e.g., animal protists (e.g., protozoa such as flagellates, ids, ciliates, and sporozoa) and plant ts (e.g., algae such alveolates, chlorarachniophytes, cryptomonads, euglenids, glaucophytes, haptophytes, red algae, stramenopiles, and viridaeplantae).

Other examples include seaweed, on (e.g., macroplankton, ankton, lankton, nanoplankton, picoplankton, and femptoplankton), phytoplankton, ia (e.g., gram positive bacteria, gram negative bacteria, and extremophiles), yeast and/or mixtures of these. In some instances, microbial biomass can be obtained from natural sources, e.g., the ocean, lakes, bodies of water, e.g., salt water or fresh water, or on land. Alternatively or in addition, microbial biomass can be obtained from culture systems, e.g., large scale dry and wet culture and fermentation systems.

In other embodiments, the biomass materials, such as cellulosic, starchy and lignocellulosic feedstock materials, can be obtained from enic microorganisms and plants that have been modified with t to a wild type variety. Such modifications may be, for example, h the iterative steps of selection and breeding to obtain desired traits in a plant. Furthermore, the plants can have had genetic al removed, modified, silenced and/or added with respect to the wild type variety. For example, cally modified plants can be produced by inant DNA methods, where genetic modifications include introducing or ing specific genes from parental varieties, or, for example, by using transgenic breeding wherein a specific gene or genes are introduced to a plant from a different species of plant and/or bacteria. Another way to create genetic variation is through mutation breeding wherein new alleles are artificially d from endogenous genes.

The artificial genes can be created by a variety ofways including treating the plant or seeds with, for example, chemical mutagens (e.g., using alkylating agents, es, alkaloids, des, dehyde), irradiation (e.g., X-rays, gamma rays, ns, beta particles, alpha particles, s, deuterons, UV radiation) and temperature shocking or other external stressing and subsequent selection techniques. Other methods ofproviding modified genes is through error prone PCR and DNA shuffling followed by insertion of the desired modified DNA into the desired plant or seed. s of introducing the desired genetic variation in the seed or plant include, for example, the use of a bacterial carrier, biolistics, calcium phosphate precipitation, electroporation, gene splicing, gene silencing, lipofection, microinjection and viral carriers. Additional genetically modified materials have been described in U.S.

Application Serial No 13/3 96,3 69 filed February 14, 2012 the full disclosure of which is incorporated herein by reference.

SACCHARIFYING AGENTS Suitable olytic enzymes include cellulases from species in the genera Bacillus, Caprinus, Myceliophthora, Cephalosporium, Scytalidium, Penicillium, Aspergillus, Pseudomonas, Humicola, Fusarium, via, Acremonium, Chrysosporium and Trichoderma, especially those produced by a strain selected from the species ASpergilluS (see, e.g., EP Pub. No. 0 458 162), la insolenS (reclassified as Scytalidium philum, see, e.g., US. Pat. No. 4,435,307), CaprinuS cinereuS, Fusarium oxySporum, Myceliophthora phila, MeripiluS giganteuS, Thielavia terrestriS, Acremonium Sp. (including, but not limited to, A. perSicinum,A. acremonium,A. brachypenium,A. dichromosporum, A. 0bclavatum,A. toniae, A. roseogriseum, A. incoloratum, and A. furatum). Preferred strains include Humicola insolenS DSM 1800, Fusarium oxySporum DSM 2672, Myceliophthora phila CBS 117.65, Cephalosporium Sp. 2, Acremom'um Sp. CBS 478.94, Acremom'um Sp. CBS 265.95, Acremom'um persicinum CBS , Acremonium acremonium AHU 9519, Cephalosporium Sp. CBS 535.71, Acremom'um brachypenium CBS 866.73, Acremonium dichromosporum CBS 683.73, m'um obclavatum CBS 311.74, Acremom'um pinkertoniae CBS 157.70, Acremom'um roseogriseum CBS 134.56, Acremonium incoloratum CBS 146.62, and niumfuratum CBS 299.70H. Cellulolytic enzymes may also be obtained from Chrysosporium, ably a strain of Chrysosporium lucknowense. Additional strains that can be used include, but are not limited to, Trichoderma (particularly T. viride, T. , and T. koningii), alkalophilic Bacillus (see, for example, US. Pat.

No. 3,844,890 and EP Pub. No. 0 458 162), and Streptomyces (see, e.g., EP Pub. No. 0 458 162).

FERMENTATION AGENTS The microorganism(s) used in fermentation can be lly-occurring microorganisms and/or engineered microorganisms. These fermentation agents can be used, for example, to convert a primary fuel to a secondary fuel to be used for energy generation in an indirect fuel cell. Of the fermentation agents can be used to convert sugars or intermediates not used in fuel cells described in the methods herein. es of microorganism can be a bacterium (including, but not limited to, e.g., a olytic bacterium), a fungus, (including, but not limited to, e.g., a yeast), a plant, a protist, e.g., a protozoa or a fungus-like protest (including, but not limited to, e.g., a slime mold), or an alga. When the organisms are compatible, mixtures of organisms can be utilized.

Suitable fermenting microorganisms have the ability to convert carbohydrates, such as glucose, fructose, xylose, ose, mannose, galactose, oligosaccharides or polysaccharides into fermentation products. Fermenting microorganisms include strains of the genus Sacchromyces spp. (including, but not limited to, S. cerevisiae (baker’s yeast), S. distaticus, S. uvarum), the genus Kluyveromyces, (including, but not limited to, K. marxianus, K. fragilis), the genus Candida (including, but not limited to, C. pseudotropicalis, and C. cae), Pichia stipitis (a relative of Candida ae), the genus Clavispora (including, but not limited to, C. lusitaniae and C. opuntiae), the genus Pachysolen ding, but not limited to, P. hilus), the genus Bretannomyces (including, but not limited to, e.g. B. clausenii ppidis, G. P., 1996, Cellulose bioconversion technology, in Handbook on Bioethanol: Production and Utilization, Wyman, C.E., ed., Taylor & Francis, Washington, DC, 179-212)). Other suitable microorganisms e, for example, Zymomonas mobilis, Clostridium spp. (including, but not limited to, Clostridium thermocellum (Philippidis, 1996, supra), Clostridium saccharobulylacetonicum, Clostridium felsineum,Clostridium saccharobulylicum, Clostridium Puniceum, idium beijernckii, Clostridium acetobulylicum, and Clostridium aurantibulylicum), Moniliella pollinis, Yarrowia tica, Aureobasidium sp., Trichosporonoides sp., Trigonopsis variabilis, Trichosporon sp., Moniliellaacetoabutans sp., Typhula variabilis, a magnoliae, Ustilaginomycetes sp., Pseudozyma tsukubaensis, yeast s of genera Zygosaccharomyces, Debaryomyces, Hansenula and Pichia, and fungi of the dematioid genus Torula.

Many such microbial strains are publicly available, either commercially or through tories such as the ATCC (American Type Culture Collection, Manassas, Virginia, USA), the NRRL (Agricultural Research Sevice Culture Collection, Peoria, Illinois, USA), or the DSMZ che Sammlung von rganismen und Zellkulturen GmbH, Braunschweig, Germany), to name a few.p cially available yeasts include, for example, Red Star®/Lesaffre Ethanol Red (available from Red Star/Lesaffre, USA), FALI® (available from Fleischmann’s Yeast, a division of Burns Philip Food Inc., USA), SUPERSTART® (available from Alltech, now Lalemand), GERT STRAND® (available from Gert Strand AB, Sweden) and FERMOL® (available from DSM Specialties).

Other than in the examples herein, or unless otherwise expressly specified, all ofthe numerical ranges, amounts, values and percentages, such as those for s of materials, elemental contents, times and temperatures of reaction, ratios of amounts, and others, in the ing portion of the specification and attached claims may be read as if ed by the word “about” even though the term “about” may not expressly appear with the value, amount, or range. Accordingly, unless indicated to the contrary, the numerical parameters set forth in the following specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present ion. At the very least, and not as an attempt to limit the application of the doctrine of lents to the scope of the claims, each numerical parameter should at least be construed in light of the number rted significant digits and by applying ordinary rounding techniques.

Notwithstanding that the numerical ranges and ters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently ns error necessarily resulting from the standard deviation found in its underlying respective testing measurements. Furthermore, when numerical ranges are set forth herein, these ranges are inclusive of the recited range end points (i.e., end points may be used). When percentages by weight are used , the numerical values reported are relative to the total weight.

Also, it should be understood that any numerical range recited herein is intended to include all sub-ranges subsumed therein. For example, a range of “l to ” is intended to include all sub-ranges between (and including) the recited minimum value of l and the recited maximum value of 10, that is, having a minimum value equal to or greater than 1 and a maximum value of equal to or less than 10. The terms “one,” “a,” or “an” as used herein are intended to include “at least one” or “one or more,” unless otherwise indicated.

Any patent, publication, or other disclosure material, in whole or in part, that is said to be incorporated by reference herein is incorporated herein only to the extent that the incorporated material does not conflict with existing definitions, statements, or other sure material set forth in this disclosure. As such, and to the extent ary, the disclosure as itly set forth herein supersedes any conflicting material orated herein by reference. Any material, or portion thereof, that is said to be incorporated by reference herein, but which conflicts with existing definitions, statements, or other disclosure material set forth herein will only be incorporated to the extent that no conflict arises between that incorporated al and the existing disclosure material.

While this invention has been particularly shown and described with references to preferred embodiments f, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the invention encompassed by the appended claims.

Throughout the specification and claims, unless the context requires otherwise, the word “comprise” or ions such as ises” or “comprising”, will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

Claims (17)

WHAT IS CLAIMED IS:

1. A method comprising: utilizing, in a fuel cell, a sugar solution comprising saccharified osic or lignocellulosic material, wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or lignocellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation.

2. The method of claim 1, wherein the method further comprises reducing the recalcitrance of the material by ically ng, chemically treating, sonicating, pyrolyzing, oxidizing, steam exploding or combinations thereof.

3. The method of claim 1 or 2, wherein irradiation comprises irradiation with a dose of at least 10 Mrad.

4. The method according to any one of claims 1-3, wherein the ionizing radiation comprises electron beam irradiation.

5. The method ing to any one of claims 1-4, wherein the cellulosic or lignocellulosic biomass is selected from the group consisting of: paper, paper products, paper waste, paper pulp, pigmented papers, loaded , coated papers, filled , magazines, printed matter, printer paper, polycoated paper, card stock, cardboard, paperboard, offal, cotton, wood, particle board, forestry wastes, sawdust, aspen wood, wood chips, grasses, switchgrass, miscanthus, cord grass, reed canary grass, grain es, rice hulls, oat hulls, wheat chaff, barley hulls, ltural waste, silage, canola straw, wheat straw, barley straw, oat straw, rice straw, jute, hemp, flax, bamboo, sisal, abaca, corn cobs, corn stover, n stover, corn fiber, alfalfa, hay, coconut hair, sugar processing residues, bagasse, beet pulp, agave e, algae, seaweed, manure, sewage, arracacha, buckwheat, banana, , cassava, kudzu, oca, sago, sorghum, , sweet potato, taro, yams, beans, favas, lentils, peas, and mixtures of any of these.

6. The method of claim 2, wherein mechanically treating the cellulosic or lignocellulosic material reduces the bulk density of the material and/or se its surface area.

7. The method according to any one of claims 1-6 further sing utilizing an additive in the fuel cell.

8. A fuel cell comprising: an electrode and saccharified cellulosic or lignocellulosic biomass, wherein the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the cellulosic or lignocellulosic al is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the material with ionizing radiation.

9. A method for producing electricity, the method comprising: providing a fuel to a fuel cell, n the fuel is produced by saccharification of a treated cellulosic or lignocellulosic material, and n the fuel cell is selected from the group consisting of direct sugar fuel cells, indirect sugar fuel cells and biological fuel cells and wherein, prior to saccharification, the recalcitrance of the osic or lignocellulosic material is reduced relative to that of the cellulosic or lignocellulosic material in its native state by irradiation of the al with ionizing radiation.

10. The method of claim 9 further comprising treatment of the cellulosic or lignocellulosic material by sonication, pyrolysis, oxidation, steam explosion or combinations thereof.

11. The method of claim 9 r comprising ting the cellulosic ir lignocellulosic material with an enzyme.

12. The method of claim 11 wherein the enzyme is a cellulase.

13. The method of claim 9 wherein the fuel comprises a sugar.

14. The method of claim 9 further comprising fermenting the saccharified treated lignocellulosic material to produce the fuel.

15. The method of claim 14 wherein the fuel is an l.

16. The method according to any one of claims 9-15, wherein irradiation comprises irradiation of the material with a dose of at least 10 Mrad.

17. The method according to any one of claims 9-16, wherein the ionizing radiation comprises an electron beam.