Patent Application
20080182309
The present invention relates to improved fermentation. Fermenters or bioreactors are used, for example, to optimize growth conditions of various strains of bacteria and tissue culture cells. Fermentation is used in numerous applications, including production of fuel alcohols such as ethanol, production of distilled beverages, food manufacturing, textiles, and pharmaceuticals. Despite the widespread use and availability of fermenters, it is still desired to be able to improve fermentation processes such as by increasing fermentation rates and yields and exercise more control over fermentation processes.
Therefore, it is a primary object, feature, or advantage of the present invention to improve over the state of the art.
It is a further object, feature, or advantage of the present invention to provide an improved fermentation process.
A still further object, feature, or advantage of the present invention is to improve the ability to control a fermentation process while maintaining a chemical free environment.
A further object, feature, or advantage of the present invention is to provide a fermenter which is self-regulating in order to optimize the fermentation process.
Another object, feature, or advantage of the present invention is to provide a biofermenter capable of providing improved ethanol production from biological material including plant material such as corn.
Another object, feature, or advantage of the present invention is to ferment carbohydrates which may be used for food or in the production of ethanol.
One or more of these and/or other objects, features, or advantages of the present invention will become apparent from the specification and claims that follow.
According to one aspect of the present invention, a method for magnetic fermentation is provided which includes subjecting a biological material in a medium to a magnetic field in order to affect fermentation of the biological material into a fermented product. The fermentation reaction may occur in an acidic or alkaline medium. The magnetic field may be a positive or negative magnetic field. The magnetic field or other parameters associated with the fermentation process may be monitored with one or more sensors and the magnetic field may be modulated accordingly.
According to another aspect of the present invention, a method for producing ethanol includes subjecting to a positive magnetic field of about 2000 to about 3000 Gauss a medium comprising a biological material and a microorganism or an enzymatic preparation or a combination thereof, wherein the microorganism or the enzymatic preparation is capable of fermenting the biological material to produce ethanol. The biological material may be any of a number of different types of fermentable materials.
According to another aspect of the present invention, a method for magnetic fermentation includes controlling a fermentation process by subjecting a biological material in a medium to a DC magnetic field having a magnitude of between 2000 to 3000 Gauss and electronically monitoring the fermentation process to generate fermentation data. The step of controlling comprises adjusting the magnitude of the DC magnetic field.
According to another aspect of the present invention a magnetic fermentation system includes a fermentation vessel for containing biological material and a medium, a magnetic field component for applying a magnetic field to the medium during a fermentation process within the fermentation vessel, wherein the magnetic field component is configured to create a magnetic field having a magnitude of between 2000 to 3000 Gauss. The magnetic fermentation system may include an intelligent control electrically connected to the magnetic field component and adapted for controlling the magnetic field applied by the magnetic field component. The magnetic fermentation system may be configured for applying the magnetic field to medium within staging operatively connected to the vessel. The staging may include a recycle stage.
According to another aspect of the present invention, a magnetic field generating device for applying a magnetic field to a fluid flowing through a conduit is provided. The device includes a plurality of magnetic modules, each of the magnetic modules being substantially U-shaped for positioning on the conduit, each of the magnetic modules having a base and legs extending from the base with an electromagnet at the base and permanent magnets near the legs. The plurality of magnetic modules being arranged in alternating fashion such that the base of each magnetic module is opposite the base of any immediately adjacent magnetic module.
The present invention provides for method and devices which may be used for applying and/or controlling magnetic fields during fermentation processes. Without wishing to be bound by this theory, it is contemplated that the use of a magnetic field and the ability to modulate the magnetic field during fermentation confers several advantages. These include but are not limited to any of the following: speeding up the dissolving of oxygen in the medium/water, increasing the rate of enzyme or bacteria's cell division, and disrupting the water's hydrogen bonding to generate unstructured water with fewer hydrogen bonds to provide a more reactive environment. Importantly, cells convert a larger faction of the sugar substrate towards cell mass production as the amount of oxygen available to the cells speeds up which allows the fermentation rate to increase.
The methods of the present invention may be used in producing any number of products whose production employs fermentation. Examples include without limitation: cellulosic ethanol that uses hydrolysis of cellulose followed by fermentation of the generated free sugars; ethanol produced by methods such as the simultaneous saccharification and fermentation of a biological material such as glucose or inulin (Ohta et al. Production of high concentrations of ethanol from inulin by simultaneous saccharification and fermentation using Aspergillus niger and Saccharomyces cerevisiae. Appl Environ Microbiol. 1993 March; 59(3):729-33); polymeric hexose and pentose sugars in cellulose and hemicellulose; glucose, lactic acid produced by the fermentation of sugars and the like. Accordingly, methods of the present invention may be used in the fermentation of a biological material to ethanol, the Simultaneous Saccharification and Fermentation (SSF) of a biological material to ethanol, and fermentation of a biological material to lactic acid.
The present invention provides for applying a magnetic field to affect a fermentation process. To assist in describing the invention, the basic process of applying magnetic fields to fermentation is described. Next, various embodiments for producing the magnetic field are described. Finally, examples of the process are provided and results from various fermentation processes are given. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Unless mentioned otherwise, the techniques employed or contemplated herein are standard methodologies well known to one of ordinary skill in the art. The materials, methods and examples are illustrative only and not limiting. The following is presented by way of illustration and is not intended to limit the scope of the invention.
The present inventions now will be described more fully hereinafter with reference to the accompanying drawings, in which some, but not all embodiments of the invention are shown. Indeed, these inventions may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. Like numbers refer to like elements throughout.
Many modifications and other embodiments of the inventions set forth herein will come to mind to one skilled in the art to which these inventions pertain having the benefit of the teachings presented in the foregoing descriptions and the associated drawings. Therefore, it is to be understood that the inventions are not to be limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims. Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.
A magnetic field is created and applied in order to affect a fermentation process. Different types of magnetic fields are used in different embodiments. The magnetic fields may be created by permanent magnets or electromagnets, although variable DC controlled electromagnets provide for convenient adjustment of magnetic fields. Generally, the magnetic fields applied are monopole or substantially monopole. The monopole magnetic fields may be positive or negative, depending upon the effect desired. The magnetic fields are controlled in a manner that assists the fermentation process, by increasing the rate of enzyme or bacteria's cell division. When the magnetic field removes positively charged calcium ions that help bind them together, this is loosening the membrane structure and is increasing its permeability, resulting in extra free calcium leaking into the cell from outside (normally there is about a thousand times greater concentration of calcium on the outside than on the inside). This process stimulate metabolism and cell multiplication (cells normally regulate their rate of metabolism by controlling their internal calcium concentration). By “chemical free environment” it is understood that applying a magnetic field does not involve use of any chemicals. A feedback loop can be established so that the intensity of the magnetic field can be varied as data regarding the fermentation process is monitored. A submersible Gauss meter or other type of magnetic field sensor can be placed in the fermenter or otherwise used to measure the magnetic field being applied. In addition oxidation reduction potential (ORP) and pH can be monitored. The magnetic fermenter essentially controls the pH levels through the amplifying process of the magnetic field. The magnetic fields can be created by generating magnetic fields within a fermentation vessel, outside of the fermentation vessel with the magnetic field directed inwardly, or during staging operations that occur outside of the fermentation vessel, such as applying the magnetic field to a pipe associated with a recycle stage.
All fermentation data received from the fermenter's computer system or other intelligent control system may be monitored and used to adjust the application of the magnetic fields. For example, where a DC power supply is used, the fermentation data may be conveyed as data correction for the electromagnets DC Power Supply. A bridge software program may interpret the data and modulate the magnetic field created in order to affect the fermentation process. In this way, the fermenter becomes self-regulating; the magnetic field for enzymatic stimulation, the pH (by increasing the voltage or by reversing the electric field polarity to the magnets) may be regulated based on a production curve and the magnetic field can be modulated such as to maintain a desired pH.
As shown in
Where magnetic fermentation is applied using a monopole positive field, it has been observed that dissolved oxygen (D) increases by about 26 percent in the first 30 minutes. It has further been observed that a positive and substantially monopole magnetic field increases the enzymes' metabolic activity by about 25 percent. Thus, the application of the positive field provides significant benefits and advantages.
Various embodiments of devices for producing magnetic fields may be used. These include embodiments where permanent magnets are used, where electromagnets are used, where the magnetic field is applied from outside of a vessel inwardly, where the magnetic field is applied from within the vessel, and where the magnetic field is applied in a staging such as the recycle stage, as opposed to within the fermentation vessel.
Thus, as previously explained, the magnetic field may be applied to the contents of the fermentation tank from outside of the fermentation tank. Alternatively, the magnetic field may be applied to the contents of the fermentation tank by placement of the magnets within the fermentation tank. In addition, the magnetic fields may be generated in various ways including through the use of permanent magnets and/or electromagnets.
The present invention also contemplates that instead of creating the magnetic field within the tank, the magnetic field may be created to affect the moving fluid within piping or staging connected to the tank or otherwise associated with the fermentation process. By “fluid” it is intended to mean the fluid mixture, including the medium and other material present with the medium. For example,
The device 150 is modular in design, and includes one or more magnetic modules 154. Each magnetic module includes an electromagnet as will be explained in greater detail. A tie rod 160 extends through each of the magnetic modules with a hex nut 162 on each end of the tie rod 160. In addition, module retention straps 156, 158 are used to secure the magnetic modules to one another. Although a configuration of five magnetic modules 154 is shown in
The magnetic modules 154 are further illustrated in
There is a gang housing 184 adjacent or otherwise proximate the electromagnet coil assembly 170 which includes an electromagnetic coil 171. A magnetic nose cover 170 and a stop plate are also shown. Pole pieces 176 are both provided on the same side of the device 154. A magnetic retention cover 178 is placed over each polepiece 176 to maintain it in place. As best shown in
Another method for creating the magnetic field is to use a solenoid device 250 as shown in
Although various embodiments have been provided showing how magnetic fields may be created, the present invention contemplates that any number of devices or methods may be used. One alternative design is to use an electromagnetic formed from a high energy coil. However, when a high energy coil is used, cooling of the coil may also need to be performed, such as through placement of the coil in mineral oil, periodically switching the electromagnet off or otherwise cooling the coil. Another alternative design is to place plastic casings containing permanent magnets and coil around the vessel. In such an embodiment, permanent magnets may be placed in a plastic casing. Subsequently, coils may be wrapped around the plastic casing and a second casing may be placed around the first casing and the coils, thus creating an electromagnetic device whose magnetic field affects the complete volume of the fermenter.
In accordance with the present invention, methods and apparatus are described for the fermentation of a biological material, such as a fermentable carbohydrate, in a medium using a magnetic field to produce a fermented product, such as ethanol. Advantageously, use of the methods of the present invention increase the yield and/or production rate of a fermented product. Experiments subjecting Ethanol Red yeast and glucose to a positive magnetic field results in the yeast multiplying twice as fast as yeast not exposed to a magnetic field as well as producing ethanol 1.4 times faster than conventional fermentation methods. Use of fermentation methods of the present invention will impart significant savings to the industry. Without wishing to be bound by this theory, it is contemplated that the use of a magnetic field and the ability to modulate the magnetic field during fermentation confers several advantages. These include but are not limited to any of the following: speeding-up the dissolving of the oxygen in the medium/water, disrupting the water's hydrogen bonding to generate unstructured water with fewer hydrogen bonds to provide a more reactive environment, and increasing the rate of enzyme or bacteria's cell division. Importantly, cells convert a larger faction of a sugar substrate towards cell mass production as the amount of oxygen available to the cells speeds-up (increases). It has been observed that trace oxygen can serve as a nutrient during the anaerobic fermentation of sugars, allowing the fermentation rate to increase as more cells are produced.
In one aspect, the invention provides for a method of producing a fermented product that includes subjecting a biological material in a medium to a magnetic field to affect fermentation of the biological material.
In one aspect, the biological materials include but are not limited to sugars, plant extracts, inulin, biomass, biomass containing cellulose, hemicellulose, lignin, carbohydrates, starches, egg shell membranes, mash, pharmaceutics, nutrients, biominerals, fruit sugars, cane press juice (high test molasses), corn/grain based starches/sugars, cellulose hydrolysate, any source of glucose, sucrose, maltose, fructose, sucrose fractions, betaine fractions, xylose fractions, residual fractions, recycled fractions, betaine, xylose, galactose, rhamnose, mannose, xylonic acidbetaine, sugar alcohols, monosaccharides, hexoses, pentoses, xylose arabinoses, lignosulphonates, oligosaccharides, complex structural polymers containing cellulose, pectins, lignins, lignocelluloses, lignin-celluloses or cellulose-containing plants, the products of cellulose-containing plants, stems, hulls, husks, cobs of plants, unprocessed plant materials, lawn clippings, leaves, fibers, pulps, hemps, sawdusts, newspapers, agricultural crops, grasses, cotton, cotton stalks, corn stalks, corn cobs, wheat straws, oat straws, rice straws, cane sugars (bagasses), soybean stalks, peanut plants, pea vines, sugar beet wastes, sorghum stalks, tobacco stalks, maize stalks, barley straws buckwheat straws, quinoa stalks, cassayas, potato plants, legume vines stalks, vegetable inedible portions, weeds, vines, kelps, flowers, algaes, bioenergy crops, trees, agricultural residues, wood residues, cellulosic fiber fines, waste papers, commercial waste products containing cellulose, paper, cotton clothes, bagasse wallboards, wood products, trees, shrubs, corn, husks, municipal solid wastes, waste papers, yard wastes, biomass high in starch, grains, fruits, vegetables, branches, bushes, canes, forests, herbaceous crops, barks, needles, logs, roots, saplings, short rotation woody crops, switch grasses, vines, hard soft woods, or organic waste materials generated from agricultural processes including farming and forestry activities, specifically including forestry wood waste and any biological material suitable as a substrate for fermentation.
One skilled in the art would be knowledgeable in the selection of the biological material for use in producing a particular fermented product. For example, complex saccharide substrates may be used as a starting source for depolymerization and subsequent fermentation for use in simultaneous saccharification and fermentation process.
In one aspect, the medium may include any medium such as water, a broth, a nutrient solution or any other medium that comes in contact with the biological material and is capable of facilitating the fermentation process. The term “broth” includes a solution as well as a suspension. The broth may be largely inorganic and include any number of minerals in solution to provide the major nutrient ions such as sodium, potassium, phosphate, sulfate, magnesium and iron and optionally an organic chelating agent to keep iron from precipitating. The absolute concentrations of the nutrients in the broth are not critical as long as they are present in adequate amounts for the microorganisms to grow but not so high as to inhibit growth. Broths include standard bacteriological growing media which can be modified in any standard manner. Typical nutrient solutions, which are well known to skilled artisans, may include minimum nutrient broth, yeast extract, corn steep liquors and the like. The appropriate medium to use in conjunction with a particular fermentation process may be determined by one skilled in the art and the medium prepared from any number of routine protocols, such as the one described in Example 1.
In one aspect, the method includes the use of a microorganism, such as yeast, bacteria, or fungus or combinations thereof to facilitate the conversion of the biological material to a fermented product. More than one biological material or microorganism may be placed within the apparatus depending on the type of fermentation process to be carried out. Any suitable microorganism or combination of microorganisms which ferments a biological material to produce a fermented product such as ethanol may be used with the methods of the present invention. “Conversion” includes any biological, chemical and/or bio-chemical activity which produces a fermented product, such as ethanol or a byproduct from the biological material, such as biomass. Suitable microorganisms for use with the methods of the present invention include yeast or bacteria that consume sugars derived from the hydrolysis of biomass. See U.S. Pat. No. 6,927,048, herein incorporated by reference in its entirety.
Genetically engineered or natural strains of microorganisms maybe used in the conversion of the biological material to a fermented product, for example, bacteria, such as Zymomonas mobilis and Escherichia coli; yeasts such as Saccharomyces cerevisiae or Pichia stipitis; and fungi that are natural producers of the fermented product, such as ethanol. The term microorganism further encompasses mutants and derivatives, such as those produced by known genetic and/or recombinant techniques, for example, those that contain pyruvate decarboxylase and/or alcohol dehydrogenase genes, those that have been produced and/or selected on the basis of enhanced and/or altered the production of the desired fermented product, e.g. ethanol.
In one aspect of the invention, an enzyme preparation may be used with the methods described herein. The enzyme preparation may include any enzyme that ferments a biological material into a fermented product. The preparation may be contain isolated or recombinant enzymes. See for example, U.S. Pat. No. 7,226,776, herein incorporated by reference in its entirety. In one aspect, a microorganism and an enzyme preparation may be used in combination in accordance with the methods of the present invention.
In one aspect, microorganisms and/or enzyme preparations, concentration of the microorganisms and/or enzyme preparations, and conditions (e.g. pH, fermentation medium, levels of nutrients, and temperature) are selected in accordance with standard techniques and may be optimized for both yield and efficiency (for example, ethanol production rate). Suitable microorganisms are well-known and commercially available.
In one aspect of the invention, the temperature of the medium comprising the biological material may be monitored using a temperature sensor and modulated in accordance with the desired operating conditions. Typically, fermentation is carried out at a temperature within the range of from about 25° C. to about 40° C., preferably within the range of from about 30° C. to about 35° C. In one embodiment, the medium having the microorganism and biological material may be heated to optimize production of the fermentation product or alternately cooled to prevent the temperature from rising to a temperature that would be stressful to the microorganisms, for example, kill them.
As described herein, the optimum pH may achieved through monitoring the pH of the medium using a pH sensor and modulating the pH by modulating the magnetic field. The optimum pH can vary from about 4.5 or lower for some microorganisms such as some yeasts, up to about 6.0 to about 6.7 or higher for other microorganisms, such as recombinant organisms. Determining the optimum pH for any given microorganism is well within the routine skill of the skilled artisan.
Using the methods and apparatus of the present invention, the biological material may be fermented to produce any number of products. In one aspect, the fermented product may include any fermented product or by-product such as ethanol, citric acid, butanol, isopropanol, lactic acid, collagen, pharmaceutics, such as antibiotics, for example, penicillin G, penicillin V and Cephalosporin C. (See K. Matsumoto, Bioprocess. Techn., 16, (1993), 67-88, J. G. Shewale & H. Sivaraman, Process Biochemistry, August 1989, 146-154, T. A. Savidge, Biotechnology of Industrial Antibiotics (Ed. E. J. Vandamme) Marcel Dekker, New York, 1984, or J. G. Shewale et al., Process Biochemistry International, June 1990, 97-103), and the like.
A determination is made with respect to whether a positive or negative magnetic field should be applied to the biological material. A positive magnetic field and a negative magnetic field will generally have the opposite effects on the fermentation process. Consideration should be given to whether the fermentation reaction occurs in an acidic or alkaline medium. For example, for reactions that take place in an alkaline (basic) medium such as oxidoreductase catalysis, a negative static magnetic field should be applied. Conversely, for reactions that take place in an acidic medium, such as fermentation, a positive static magnetic field should be applied. Without wishing to be bound by this theory, it is believed that a positive static magnetic field increases the activity of transferases, hydrolases or both. Both oxidation phosphorylation and fermentation catalysis are energized by static magnetic fields; however, the reactions are energized by opposite magnetic poles. It is believed that oxidation phosphorylation is energized by a negative static magnetic field in an alkaline-hyperoxic medium, whereas fermentation is energized by a positive static magnetic field in an acid-hypoxic medium.
Any suitable apparatus may be used in conjunction with the methods of the present invention so long as the apparatus can be used to ferment the biological material and a source of a magnetic field can be applied to the biological material. Any suitable apparatus can be used, for example, batch, fed-batch, cell recycle, continuous process or multi-step bioreactors. Accordingly, any suitable technique may be used to expose the biological material to the magnetic field, for example, a magnetic field may be applied internally or externally with respect to the apparatus, passing through the apparatus if needed to reach the biological material placed within the apparatus. In one aspect, the magnetic field may be placed to take advantage of the circulating biological material as it is believed that aerating or suspending the microorganisms may improve the magnetic effect.
The magnetic field may be applied using any source of such radiation, for example, permanent magnets or electromagnets. In general, the magnetic source is applied to the biological material in the apparatus for an appropriate time and at an appropriate level to affect fermentation. Depending on the effect desired, a negative field may also be applied. In one aspect, the range of the applied magnetic field is from about 2000 to about 3000 Gauss and it is preferred that the field is generally uniform, although it is recognized that some variation may be present. In one aspect, the magnetic field is from about 2200 to about 2400 Gauss. In one aspect, a DC electromagnetic “monopole positive” flux density of about 2200 to about 2400 Gauss is applied on the medium or fermenter's fluid/solids. In one aspect, the medium with the biological material/fluid is passing through a pipe at no more than 0.1 to 5 seconds at any given time. In another aspect, the medium with the biological material/fluid is passing through a pipe at less than 20 minutes of the fluid cycle. Any level of magnetic field, length of exposure to the magnetic field or flow rate of the medium/fluids may be used so long as the fermentation reaction is able to take place. According to the present invention, a method of fermenting a biological material includes contacting a medium with a biological material and a source of a magnetic field for a time sufficient for the fermentation to occur. The present invention contemplates providing the magnetic field for a first time period, and then turning the magnetic field off for a second time period. For example, the magnetic field may be turned on and off every 30 seconds. Of course, other time periods may be used, and the first time period during which the magnetic field is applied need not be the same duration as the second time period where the magnetic field is turned off. It is preferred that any amount of medium/water/solids or fluid is not exposed continuously to a magnetic field for more than 30 seconds.
Optimal levels of magnetic fields may be determined. It has been observed that in some instances, applying magnetic fields greater than 3000 Gauss decreases ethanol production from yeast. As previously explained, a magnetic field sensor may be coupled to a feedback arrangement and a controller for modulating the magnetic field until the magnetic field is at or above a predetermined magnetic field value. The magnetic field may be monitored and compared to a previous level of the magnetic field, for example, using a submersible Gauss meter and feedback loop. In one aspect, one or more detectors are used to measure the intensity of the magnetic field. In one aspect, the detector is placed within the apparatus. Alternately, the detector may be placed external to the apparatus opposite the source of the magnetic field depending on the strength of the field. The intensity of the field may be modulated to achieve a given parameter. The adjustment may be automated or manual. The magnetic field may be monitored and compared to a previous level of the magnetic field, for example, using a Gauss meter and feedback loop.
According to one aspect of the invention, the methods include modulating the level or intensity of the magnetic field during the fermentation process. As used herein, the term “modulate”, “modulates” or “modulating” refers to a change, i.e. an increase or decrease in the magnetic field.
The biological material is exposed to medium comprising one or more sources of the appropriate magnetic field and optionally a pH sensor. The medium is exposed to the source of magnetism for a time sufficient for the reaction to take place. As the reaction occurs, the pH of the medium will change, Accordingly, in one aspect, the methods include using one or more pH sensors to detect a change in the pH of the medium as the fermented product is produced. For example, as an acidic product is produced, the medium becomes more acidic and the pH decreases. Conversely, when an alkalinic product is produced, the medium becomes more basic and the pH increases. The change in the pH of the medium may be detected using any number of methods. The differences in change of pH may be compared to a control that is not subjected to a magnetic field.
As discussed previously, the pH of the medium may be assayed to determine whether a positive or negative magnetic field should be applied to the biological material. The biological material is exposed to medium comprising one or more sources of a magnetic field and optionally a pH sensor.
Accordingly, in one aspect, the method includes determining the pH of the medium comprising a biological material so that the pH can be modulated using a magnetic field. If the fermentation takes place in an acidic medium, then the pH can be monitored and, if desired, regulated to maintain an acidic pH, for example, within a desired pH range. For example the pH of the medium containing the biological material may be adjusted, for example, to a pH of about 3 to about 4.5 when fermenting glucose to produce ethanol. As a fermented product such as ethanol is produced, the medium becomes more basic. If the fermentation takes place in a basic medium, then the pH can be monitored and, if desired, regulated to maintain a basic pH.
The medium may be evaluated for a change in pH, for example, using a pH sensor. The sensor may operate continuously or at frequent time intervals to monitor the pH. Prior to any detection, an initial pH level of the medium may be determined. In one aspect, the change in the pH may be monitored at various time points for example, at an initial starting point and then at various time points thereafter and compared to the previous pH reading. Time points may vary from hours to days depending on the criteria of the experimental design and the type of fermentation product being produced. Such criteria include but are not limited to the amount of biological material in the medium, the amount of medium, the temperature and the type of microorganism. If a reading from the pH sensor detects an unacceptable pH level, the intensity of the magnetic filed can be modulated. Note that there is no need to add an acid or base to alter pH levels of the medium. These affects can be achieved in a chemical free manner.
In one aspect, a pH sensor is coupled to a feedback arrangement and a controller for modulating the magnetic field until the medium is at or above a predetermined pH. The change in the pH of the medium may vary depending on the medium and biological material present in the medium, the microorganism and the amount of fermented product produced. A variety of detectors such as a magnetic field sensor, pH sensor, temperature sensor, oxidation reduction potential sensor, ethanol or glucose sensor can be selected to provide a number of measurements for use in the methods and/or apparatus of the present invention, which measurements will depend on the type of fermentation reaction and the parameters being controlled.
The amount of biological material or fermented product produced by the methods of the present invention may be determined. The differences in the amount of biological material or fermented product may be compared to a control that is not subjected to a magnetic field to determine yield or efficiency for the fermentation methods. The yield or rate of fermented product produced using fermentation methods of the present invention may be determined and compared to the yield or rate relative to another fermentation method that does not use magnetic fermentation using, for example, qualitative, quantitative, or statistical evaluation.
As used herein, “yield” may include reference to the amount of fermented product produced, for example, the amount of fermented product produced (gr/l), such as ethanol or lactic acid, divided by the amount of biological material consumed (gr/l), such as glucose. One skilled in the art will be able to determine yield for a particular fermented product. For example, the medium may be removed from the apparatus to facilitate determination of the level of biological material or fermented product in the medium. Levels may be determined using for example High Performance Liquid Chromatography (HPLC), a Biochemistry Analyzer such as YSI 2700 (YSI Inc, Yellow Springs, Ohio) or Cobas Mira Biochemistry Analyzer (F. Hoffmann-La Roche, Ltd, Nutley, N.J.), or mass spectrometry. A “control” may comprise, for example: (a) medium that contains the same starting biological material but which has not been subjected to a magnetic field (b) a medium that does not contain a starting biological material. Thus, a “control” may be used to provide a reference point for measuring changes in pH, yield, or production rate, or concentration of biological material or fermented product when using the fermentation methods of the present invention as compared to more conventional fermentation methods.
The fermentation methods described herein can include any number of steps, for example, a feeding step where the biological material is broken down or consumed, a recycling or circulation phase, and a product recovery phase where the fermented product is recovered. Other products of cellulose-containing plants may be recovered using the methods of the present invention such as waxes, gums, oils, sugars, wood alcohol, agar, rosin, turpentine, resins, rubber latex, dyes, glycerol, etc.
Advantageously, at least some of the steps during the fermentation process can occur sequentially, continuously, or simultaneously. In one aspect, the method includes membrane filtration, for example, for use in a saccharification stage, byproduct recovery stage or fermentation stage to retain enzymes, carbohydrates, salts, or microorganism to enhance the rate of fermentation. In another aspect, membrane filtration may be used to recover byproducts produced in some fermentation processes such as glycerol, lactic acid and others and or to reduce the amount of solids going to an evaporator.
The fermentor used with the methods of the present invention is typically an anaerobic fermentor which may be continuous, batch fed, or simple batch. Carbon dioxide, which is byproduct of fermentation, can be removed continuously from the fermentor. If a continuous or batch fed fermentor is used, then optionally, on a continual basis, fluid having ethanol may be drawn off from the fermentor and treated to recover ethanol, for example, by evaporation and/or distillation. The ethanol concentration above which the fermenting organisms will decrease or cease production will depend upon the particular microorganism used. Accordingly, the fermented product produced by the methods and/or apparatus of the present invention may be recovered using any suitable method, for example, ethanol may be removed from the medium by evaporation or by membrane filtration technology.
In one aspect, the fermentation method of the present invention includes monitoring the ORP of the medium using an ORP sensor. In one aspect, the ORP may be adjusted to optimize the ORP for the specific fermentation reaction. See U.S. Pat. No. 7,078,201 to Burmaster describing oxidant addition (such as air or oxygen sparging, peroxide etc), reductant substitution (such as ammonia with caustic), or reductant elimination (such as oxidation of sulfite) to adjust the ORP. Optimization of ORP is well within the skill of one skilled in the art.
Without wishing to be bound by this theory, it is contemplated that the use of a magnetic field and the ability to modulate the magnetic field during fermentation confers several advantages. These include but are not limited to any of the following: speeding up the dissolving of oxygen in the medium/water, increasing the rate of enzyme or bacteria's cell division, and disrupting the water's hydrogen bonding to generate unstructured water with fewer hydrogen bonds to provide a more reactive environment. Importantly, cells convert a larger faction of the sugar substrate towards cell mass production as the amount of oxygen available to the cells speeds up which allows the fermentation rate to increase.
In one aspect, the methods of the present invention include increasing the yield of a fermented product. In another aspect, the methods of the present invention include increasing the production rate of the fermented product. In another aspect, the methods of the present invention include shortening the length of time to produce the fermented product. In another aspect, the methods of the present invention include increasing the growth rate of the microorganism.
As described previously, methods of the present invention may be used produce fermented products that have application in various industries, such as but not limited to food containing lactic acid such as yogurt, and alcohol in distilled beverages, such as potable beers, wines, and grain alcohols, as well as industrial and fuel alcohol such as ethanol, pharmaceutics, textile industries, and biodegradable plastics (Brown, S. F., 2003, Fortune, 148:92 94; Datta, R., et al., 1995, FEMS Microbiol. Rev. 16:221 231).
The methods of the present invention may be used in producing any number of products whose production employs fermentation. Examples include without limitation: cellulosic ethanol that uses hydrolysis of cellulose followed by fermentation of the generated free sugars; ethanol produced by methods such as the simultaneous saccharification and fermentation of a biological material such as glucose or inulin (Ohta et al. Production of high concentrations of ethanol from inulin by simultaneous saccharification and fermentation using Aspergillus niger and Saccharomyces cerevisiae. Appl Environ Microbiol. 1993 March; 59(3):729-33); polymeric hexose and pentose sugars in cellulose and hemicellulose; glucose, lactic acid produced by the fermentation of sugars and the like.
Accordingly, methods of the present may be used in the fermentation of a biological material to ethanol, the simultaneous saccharification and fermentation of a biological material to ethanol, and fermentation of a biological material to lactic acid.
In one embodiment, the methods and apparatus of the present invention may be used to ferment sugars or starches to ethanol using a positive magnetic field. In one aspect, a microorganism such as yeast is used to carry out the enzymatic conversion. In one aspect, microorganisms, concentration of the microorganisms, selection of biological materials and conditions (e.g. pH, fermentation medium, levels of nutrients, and temperature) are selected in accordance with standard techniques and may be optimized for both yield and efficiency (ethanol production rate). See U.S. Pat. No. 4,349,628 to English et al; see also U.S. Pat. No. 5,932,456 to Van Draanen et al., U.S. Pat. No. 4,400,470 to Zeikus et al; U.S. Pat. No. 5,000,000 to Ingram et al; U.S. Pat. No. 5,028,539 to Ingram et al; and U.S. Pat. No. 5,162,516 to Ingram et al, disclosing the conversion to ethanol of polymeric hexose and pentose sugars in cellulose and hemicellulose, all of which are incorporated herein by reference.
In one embodiment, the fermentation of ethanol from a biological material using a magnetic field may be part of a dry grind process, modified dry grind process or wet mill process. In one embodiment, the ethanol production facility utilizes grain as a starting biological material. In one embodiment, the grain is selected from the group consisting of sorghum, wheat, barley, oats and rice. The liquid medium processing stream can include heavy steep water, an uncooked slurry, a cooked mash, a liquefied mash, and (for a dry grind process) whole stillage, thin stillage and wet cake.
Those skilled in the art will appreciate, and readily accommodate, without undue experimentation, adjusting the magnetic field, concentration of the microorganisms, and conditions (e.g. pH, fermentation media, levels of nutrients, and temperature) for yield and efficiency all in accordance with the teachings disclosed herein.
It is understood that both the substrate and product of the ethanolic fermentation may inhibit the fermentation process or effect the fermentation rates. Accordingly, in one aspect, the method includes fermenting the sugar to ethanol and removing the resulting ethanol. The fermented product such as ethanol can be recovered using any suitable means, for example, by a ferment stripper, distillation or membrane technology. (See for example, U.S. Pat. Nos. 4,665,027 and 5,141,861, herein incorporated in their entirety) with gas stripping of ethanol from the broth, the vacu-ferm fermentation suggested by Ramalingam and Finn (1977), the coupled fermentation/distillation Biostil process developed by Alpha Laval and then acquired by Chematur Engineering (1994), etc.). Accordingly, ethanol may be separated using for example to remove ethanol as it is produced. The amount of ethanol produced may be analyzed, for example, by High Performance Liquid Chromatography (HPLC), a YSI 2700 Biochemistry Analyzer (YSI Inc, Yellow Springs, Ohio), Cobas Mira Biochemistry Analyzer (F. Hoffmann-La Roche, Ltd, Nutley, N.J.), or mass spectrometry.
In one aspect, methods of the present include a method for the Simultaneous Saccharification and Fermentation (SSF). In SSF, product inhibition of the cellulases can be avoided by conversion of the glucose into ethanol or other desired fermentation product. The SSF philosophy has been used for decades by the ethanol industry with starch enzymes. Research also shows that this concept works for the hemicellulase and cellulase enzyme systems. The Gulf Oil Company developed a method for the production of ethanol from cellulose using a yeast-based process termed simultaneous saccharification and fermentation (SSF) (Gauss et al. (1976) U.S. Pat. No. 3,990,944, herein incorporated in its entirety). Fungal cellulase preparations and yeasts may be used to produce ethanol from a slurry of the cellulosic biological material. Ethanol may be produced concurrently during cellulose hydrolysis.
The invention also provides methods for the production of lactic acid by subjecting a biological material to fermentation using a magnetic field. In one aspect, the biological material is cellulose and hemicellulose. In another aspect, the method includes culturing a microorganism capable of fermenting a biological material under conditions suitable for the production of lactic acid. The method may further comprise the optional step of recovering include L(+)-lactic acid, 1,3-propanediol, 1,2-propanediol, succinic acid, ethanol and D(−)-lactic acid. See U.S. Pat. No. 7,098,009 to Shanmugam et al.
The configuration and components employed in any apparatus using fermentation methods may be coordinated with the application requirements, for example, the scale of the operation and amount of product desired. Various embodiments of the invention, including different configurations and utilizing diverse components for the generation of a magnetic field are possible.
This invention can be better understood by reference to the following non-limiting examples. It will be appreciated by those skilled in the art that other embodiments of the invention may be practiced without departing from the spirit and the scope of the invention as herein disclosed and claimed.
The present invention is further defined in the following Examples, in which parts and percentages are by weight and degrees are Celsius, unless otherwise stated. The disclosure of each reference set forth herein is incorporated herein by reference in its entirety.
One fermentation protocol, referred to as MF3, includes carrying out fermentation of 5 L or 100 L in a bioreactor containing culture medium with 250 g/L dextrose, 20 g/L peptone (Fisher), 10 g/L yeast extract (Fisher). Flasks may be inoculated with yeast, such as 0.44 g/L of Ethanol Red or Red Star yeast (Fermentis) and incubated at 32° C. with agitation (about 200 rpm) at a pH of 4.5 with aeration (about 0.8 vvm for 100 L or 1.0 vvm for 5 L).
Another fermentation protocol used, referred to as F15MF9TP3, includes carrying out fermentation of 5 L or 100 L in a 14 L bioreactor containing culture medium with 250 g/L glucose, 20 g/L peptone (Fisher), 10 g/L yeast extract (Fisher). Flasks were inoculated with 0.44 g/L of Red Star yeast (Fermentis) and incubated at 32° C. with agitation (about 200 rpm) at a pH of 4.5 with aeration (<0.8 or 1 vvm) with an air LPM of 5.
A magnetic field may be applied to these protocols as described in more detail elsewhere herein.
To monitor cultures, samples from the medium may be removed at various time points to determine the glucose and ethanol concentrations, cell count, dissolved oxygen, optical density and BRIX using High Performance Liquid Chromatography (HPLC), YSI 2700 Biochemistry Analyzer (YSI Inc, Yellow Springs, Ohio), Cobas Mira Biochemistry Analyzer (F. Hoffmann-La Roche, Ltd, Nutley, N.J.), gas chromatography (Dombek et al (1986) Appl. Environ. Microbiol. 52:975 981) or other suitable techniques.
The Cobas data demonstrates that magnetic fermentation has a better growth rate, ethanol yield, and ethanol production rate.
Dry corn is ground, mixed with water producing a slurry, heat-treated through a jet cooker with alpha-amylase to swell the starch and break the starch into smaller polymers. This pasteurized corn mash is then fortified with urea as the nitrogen source, inoculated with active commercial yeast strain, and starch hydrolyzed to glucose by the addition of glucoamylase. The yeast converts one glucose molecule into two ethanol and two carbon dioxide molecules. Typically a 48 to 60 hour yeast fermentation will yield 18% ethanol by volume from a 32% corn mash by solids. The whole fermentation mash is then passed through a distillation column to remove the ethanol. This is followed by a low gravity centrifugation in which the solid portion called distillers grains and the supernatant called thin stillage which is then concentrated by flash evaporation into syrup are recovered. The syrup is added back onto the distiller grains, and dried to produce dried distiller grains with solubles (DDGS). The 95% ethanol recovered from the distiller is passed through a molecular sieve to remove the 5% water which produces fuel grade ethanol. The DDGS is ship to farmers around the world for animal feed, primarily for ruminant livestock.
In one exemplary embodiment, fermentation for use in ethanol production is described. The ethanol market is currently experiencing high growth. Ethanol is generally blended with gasoline at various levels to fuel motor vehicles. Due to limited supplies of crude oil and limitations in refining capacity, concerns over environmental degradation, and the resulting increase in gasoline prices, there appears to be a positive outlook for further growth in the ethanol market. Ethanol can be produced from various sources, including corn, barley, and wheat, as well as cellulose feedstocks. For purposes of this exemplary embodiment, corn is used to produce ethanol.
Cellulose is the most abundant organic compound in the biosphere and it is found in all plant materials as lignocellulose. Cellulose is plant cell wall (30-40%). Lignin is the cell cement that holds plant cells together (20-30%). Hemicellulose is found dispersed outside the plant cell (30-40%). Plant biomass (i.e., switch grass, corn stovers, wood chips, etc.) are dried in the field and stored in a reduced moisture environment. Bails of dried plant biomass is then ground, pretreated to remove microbial and enzymatic inhibitors, pasteurized at high temperatures, then fermented. The fermentation process includes the hydration (water contribution to the protein structure), as protein stability has been directly tied to the equilibrium of structuring water between low-density and higher density forms. It is believed that applying and controlling the magnetic field disrupts the water's hydrogen bonding, thus affecting the protein's structure by speeding-up its un-folding (denaturation).
The fermentation slurry contains pretreated ground plant biomass, cellulosic enzymes and yeast for ethanol bioconversions of glucose from cellulose and of possibly pentoses from hemicellulose. Residual co-products will have value in feed and non-feed applications (i.e., soil amendments, plastics, adhesives, asphalt, etc.).
All publications and patent applications in this specification are indicative of the level of ordinary skill in the art to which this invention pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated by reference.
The invention has been described with reference to various specific and preferred embodiments and techniques. However, it should be understood that many variations and modifications may be made while remaining within the spirit and scope of the invention.
Thus, methods and apparatus related to fermentation have been disclosed. The present invention contemplates numerous variation in the type of fermentation process, whether the magnets used are permanent or magnetic, the number and types of sensors used, the number and types of different controls, the methodology for controlling a DC power supply when used, and other variations. The present invention is not to be limited to this disclosure as these and other variations are contemplated which fall within the spirit and scope of the invention.
This application claims priority under 35 U.S.C. § 119 to provisional application Ser. No. 60/823,023 filed Aug. 21, 2006, herein incorporated by reference in its entirety.
| Number | Date | Country | |
|---|---|---|---|
| 60823023 | Aug 2006 | US |
