WO1996039799A1 - Procede de preparation d'une culture a l'aide d'un champ magnetique et composition contenant cette culture - Google Patents

Procede de preparation d'une culture a l'aide d'un champ magnetique et composition contenant cette culture Download PDF

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Publication number
WO1996039799A1
WO1996039799A1 PCT/US1996/009574 US9609574W WO9639799A1 WO 1996039799 A1 WO1996039799 A1 WO 1996039799A1 US 9609574 W US9609574 W US 9609574W WO 9639799 A1 WO9639799 A1 WO 9639799A1
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WIPO (PCT)
Prior art keywords
culture
microorganisms
composition
resulting
odor
Prior art date
Application number
PCT/US1996/009574
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English (en)
Inventor
Craig Jones
D. Michael Bitz
Original Assignee
E.K.M.A., Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US08/476,374 external-priority patent/US5866112A/en
Priority claimed from US08/569,769 external-priority patent/US5843427A/en
Application filed by E.K.M.A., Inc. filed Critical E.K.M.A., Inc.
Priority to AU62625/96A priority Critical patent/AU6262596A/en
Publication of WO1996039799A1 publication Critical patent/WO1996039799A1/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N13/00—Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L9/00—Disinfection, sterilisation or deodorisation of air
    • A61L9/01—Deodorant compositions
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
    • C05F5/00—Fertilisers from distillery wastes, molasses, vinasses, sugar plant or similar wastes or residues, e.g. from waste originating from industrial processing of raw material of agricultural origin or derived products thereof
    • C05F5/006—Waste from chemical processing of material, e.g. diestillation, roasting, cooking
    • C05F5/008—Waste from biochemical processing of material, e.g. fermentation, breweries
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
    • C05F9/00—Fertilisers from household or town refuse
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G5/00—Fertilisers characterised by their form
    • C05G5/20—Liquid fertilisers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
    • Y02A40/10—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in agriculture
    • Y02A40/20—Fertilizers of biological origin, e.g. guano or fertilizers made from animal corpses

Definitions

  • the present invention relates, in general, to a biocatalyst and to a method of stimulating biological activity using same.
  • the present invention relates to a composition capable of 5 stimulating plant growth and to a method of preparing such a composition.
  • the invention also relates to a composition capable of reducing or eliminating odors and to a method of odor reduction or elimination based on same.
  • the invention further relates to a method of 0 effecting bioremediation.
  • compositions capable of stimulating biological activity, including microbial activity have application in a number of areas. Such compositions can be used, for example, to stimulate plant growth and thus are useful in farming and commercial and residential landscape maintenance.
  • Various plant growth stimulating compositions are available that are derived from natural or from synthetic sources.
  • the compositions of the present invention have constituents from both sources and have the advantage of superior growth stimulatory properties.
  • the compositions of the present invention have the further advantage that they can be tailored so as to be optimum for a particular plant type growing under particular soil conditions.
  • compositions that stimulate biological activity can also be used to reduce or eliminate odors .
  • odors are sufficiently offensive to be problematic for individuals having to work in proximity with the odor producing material. Examples of such situations include cattle feed lots, swine barns, poultry houses and the like. In these settings, the ability to reduce or eliminate odor production would greatly enhance the quality of the working environment.
  • the present invention provides a composition suitable for use in such settings.
  • compositions that stimulate biological activity can also be used to effect bioremediation.
  • the present invention relates to a method of preparing a culture of microorganisms .
  • the method comprises : i) obtaining a starting culture sample of microorganisms from the gastrointestinal track of a mammal; ii) culturing the sample in a medium comprising sodium, potassium, calcium, magnesium, inorganic phosphorus and chlorine or salts thereof; iii) culturing the sample resulting from step (ii) in the presence of a food source comprising a grain or a grass; and iv) separating the culture of microorganisms resulting from step (iii) from the food source.
  • the culture resulting from step (iv) is exposed to a magnetic field.
  • microorganisms are removed from the culture resulting from step (iv) , with or without exposure to a magnetic field.
  • the present invention relates to a biological activity stimulatory composition
  • a biological activity stimulatory composition comprising a component produced by the above method and a formulation comprising Na, Cl, P, Mg, Ca, S, Zn, Cu, Co, I, Se, Fe, K, Mn, Mo, Si, B, Ni and Rb.
  • the present invention relates to such a formulation.
  • the present invention relates to method of stimulating the growth of a plant.
  • the method comprises administering to the plant the above composition under conditions such that the stimulation is effected.
  • the present invention relates to a method of preparing a culture of microorganisms for use as a constituent of a plant growth stimulating composition comprising exposing the culture to a magnetic field under conditions such that thickening of the cell walls of the microorganisms, as determined by light microscopy, is effected.
  • the present invention relates to an odor-reducing composition
  • an odor-reducing composition comprising an acid component, an iron component and a nitrogen componen .
  • the present invention relates to a method of reducing or eliminating an offensive odor comprising contacting the compound responsible for the odor with the above odor-reducing composition under conditions such that the odor is eliminated or reduced.
  • the present invention relates to a method of inhibiting odor production at a source comprising contacting the source with the above odor-reducing composition under conditions such that the inhibition is effected.
  • the present invention relates to a method of effecting bioremediation.
  • the method comprises applying an amount the above-described biological activity stimulatory composition to a site in need of bioremediation sufficient to effect bioremediation.
  • Figure 1 Diagram of fermentation tanks for preparation of microbial cultures.
  • Figure 2 Diagram of orientation of magnets relative to recirculation tube.
  • the present invention relates to a method of processing microbial cultures for use as constituents of compositions that stimulate biological activity, for example, of microorganisms present at a particular site.
  • the invention relates to plant growth stimulating compositions.
  • the invention also relates to nutrient formulations to be used in combination with such processed cultures in the stimulating compositions.
  • the cultures and formulations of the invention can be used to stimulate the growth of a variety of plant types including sugar cane, vegetables, fruits, grasses and tropical plants.
  • the cultures and formulations can also be used to advantage on ornamental plants.
  • the present invention also relates to a method of reducing or eliminating odors and inhibiting the production thereof, and to compositions suitable for use in effecting that reduction/elimination or inhibition.
  • the odor-reducing compositions of the invention can be used to reduce or eliminate odor production in a variety of settings, including barns, poultry houses, kennels and other animal holding areas, feed lots, and areas where water is contained, such as lagoons. These compositions can also be used in waste control area such as landfills, trash transfer centers, leachate reservoirs and animal disposal areas.
  • the composition of the present invention comprises an acid component or salt thereof such as citric acid, acetic acid, ascorbic acid, malic acid or tartaric acid, an iron component, such as a ferric salt, for example, ferric chloride, and a nitrogen source, such as urea.
  • the composition can further comprise one or more of the following: a molybdenum component, for example, molybdic acid or salt thereof, a copper component, for example, copper sulfate, aloe vera and a gum component, such as xanthan gum or guar gum.
  • composition of the invention can include an enzyme component derived from a microbial culture supernatant or prepared from polysaccharide hydrolases such as starch hydrolases, including ⁇ -amylase and glucoamylase, and galactomannan hydrolases, such as hemicellulase.
  • polysaccharide hydrolases such as starch hydrolases, including ⁇ -amylase and glucoamylase, and galactomannan hydrolases, such as hemicellulase.
  • Enzyme components of each of the compositions of the present invention can be derived from a microbial culture or supernatant thereof using the following preparative procedure.
  • Starting cultures suitable for use in preparing microbial cultures appropriate for various aspects of the invention can be obtained by combining isolates of specific microbial strains or by obtaining a mixed culture from an animal, for example, from the gastrointestinal track of an animal, preferably, a mammal, more preferably a herbivore, most preferably a cow (e.g., a lactating cow) .
  • Starting cultures can be obtained, for example, from a sample aspirated from the stomach of an animal (e.g. from the rumen of a herbivore) or from a fecal sample taken from the intestinal track of an animal.
  • the starting culture is cultured for an initial period (e.g.
  • a medium that can be prepared from natural sources (e.g. from the saliva of a herbivore (e.g. a cow)) or from chemicals (culturing can be carried out in a container such as Tank 1 of Figure 1) .
  • a bolus e.g. about 1 liter
  • a filter e.g. about an 80 micron filter
  • the bolus is washed with warm water (about 10 liters of water per liter of bolus) and the filtrate (pH preferably about 6.3 to 6.8) is obtained and used as the initial culture medium.
  • warm water about 10 liters of water per liter of bolus
  • the filtrate pH preferably about 6.3 to 6.8
  • synthetic medium it is preferably formulated so as to contain the following:
  • a culture medium containing the following can be used:
  • concentrations of the culture medium components can vary depending on the starting culture and on the target, however, typically concentrations vary, for example, by plus or minus 45%, preferably, plus or minus 20% from the above.
  • the starting culture sample is incubated in the culture medium, preferably, at a pH in the range of 6.9 to 7.3. Adjustments in pH can be made at this point and throughout the process using a variety of acids and bases, sulfuric, hydrochloric and citric being the preferred acids, citric being more preferred, and potassium hydroxide, calcium hydroxide and sodium bicarbonate being the preferred bases, sodium bicarbonate being most preferred.
  • air is introduced (e.g. by compressed air injection) to maintain an oxygen content in the range of 3 to 5 ppm (oxygen, nitrogen and argon being major components of compressed air) .
  • oxygen, nitrogen and argon being major components of compressed air
  • Cells increase in number, nutrient content and in cell-wall content.
  • a food source (substrate) is subsequently (e.g. after about 23 to 28 hrs, preferably 24 hrs) added to the medium/starting culture sample mixture.
  • the food source can comprise a mixture of feed grains and grasses. Preferably, at least three of the following are added in approximately equal parts by weight:
  • a mixture of crushed corn, oats, alfalfa and whole peanuts is preferred. The same is advantageous for tropical plants.
  • grasses are the target plant (e.g. in the case of golf course maintenance)
  • a mixture of crushed corn, oats, alfalfa and flax is preferred.
  • alfalfa, wheat soy beans and barley are preferred.
  • the food source is typically added to the medium/starting culture sample together with a further volume of liquid (i.e., culture medium and water (e.g., in a ratio of about 1:5 to 1:4)) in a ratio of about 1 kg of dry matter to about 7-8 liters of liquid. Multiple additions of food source and liquid to the original medium/starting culture sample can be made, 2 additions at approximately 24 hour intervals being preferred.
  • an approximately neutral pH is maintained, a pH in the range of about 6.9 to about 7.3 being preferred.
  • the temperature is maintained, preferably, in the range of about 34° to 41°C, 37° to 40°C being preferred.
  • the resulting broth is well mixed, for example, by recirculating the broth in a recirculation tank.
  • the recirculation is typically for a period of about 24 hours, after which time the broth is allowed to stand for a period sufficient to allow the particulate matter to settle out.
  • An aliquot of the broth supernatant is then removed and placed in a second container (e.g., a tank such as Tank 2 in Figure 1) .
  • a second container e.g., a tank such as Tank 2 in Figure 1 .
  • the pH of the transferred aliquot is slowly reduced (e.g., over a period of several hours) to about 4.5 to about 6.3, 5.1/5.8 to 6.3 being preferred, 5.1 to 6.1 being more preferred (the pH can in fact range from 3.4 to 9.0) .
  • the temperature is maintained in the range of about 34°C to 41°, 37° to 41°C being preferred.
  • a minimal amount of a second food source is added (e.g., about l%-3% v/v of the aliquot, 3% being preferred) .
  • the second food source is, for example, molasses (e.g., sugar cane or citrus molasses) , aloe vera, papaya juice, stearate or glycogen.
  • Sugar cane molasses is preferred when sugar cane or grass is the target plant, citric molasses being preferred in the case of citrus and vegetable crops as well as tropical plants (papaya juice can also be advantageous in the case of tropical plants) .
  • Glycogen, aloe and citrus molasses are preferred when the composition is to be used for odor reduction or elimination.
  • the number of cells per ml is, advantageously, in the range of 700,000 to 1.5 million per ml, about 850,000 cells/ml to 900,000 cells/ml being preferred, around 890,000 cells/ml being most preferred.
  • the cell count can be increased by delaying the transfer of the aliquot from the first tank to the second.
  • an aliquot of the culture e.g., transferred to a tank such as Tank 3 of Figure 1
  • a magnetic field preferably twice
  • the field is created using an electromagnet or permanent magnets, for example, rare earth magnets.
  • an appropriate field can be created, for example, by two opposing magnets.
  • Magnets suitable for use in the present invention have a strength in the range of 1200 to 4500 gauss, about 3500 gauss magnets being preferred.
  • Figure 2 includes a diagram of a preferred orientation of such magnets. While, in the Figure, like poles (i.e., north poles) are shown to face either side of the recirculation tube, such need not be the case (e.g. opposite poles can also face the tube) .
  • magnetic fields can also be generated by particle movement.
  • the exposure of the microorganisms to the magnetic field results in an increase in the thickness of the microbial cell wall and an increase in cell mobility, as viewed under light microscopy.
  • the invention contemplates the use of magnetic fields that can achieve these ends.
  • the temperature is, advantageously, maintained between 18° and 42°C.
  • the cell concentration is, preferably, held between 100,000 and 150,000 cells/ml.
  • a formulation of nutrients is added that can include Na, Cl, P, Mg, Ca, S, Zn, Cu, Fe, K, Mn, Mo, Si, B, Ni, and Rb, preferably, also Co, I, or Se.
  • the formulation has the following composition and the concentration ranges listed (g/1) reflect the increase in concentration of the components in the culture upon addition of the formulation to the culture:
  • Chloroacetic acid .0001-.04 .0005-.03
  • Formulations advantageous for tropical plants, vegetables and grass are as follows (expressed in g/1 of culture, the form in which each is added being as indicated above) (see Example for sugar cane values) : Tropical Golf Course
  • the values for tropical plants can vary, for example, by plus or minus 59%, preferably, plus or minus 28%; the values for vegetables by plus or minus 430%, preferably, plus or minus 22%; the values for grass by plus or minus 450%, preferably, plus or minus 20%; and the values for sugar cane by plus or minus 45%, preferably, 20%.
  • the resulting composition can be processed (e.g., filtered or centrifuged) so as to remove microorganisms, used immediately or stored, for example, for as long as two years.
  • Stored compositions containing the microorganisms can be processed to remove the microorganisms prior to use.
  • the pH is maintained, preferably, at about 5 (e.g., 4.9 to 5.2) , however, a pH range of 5.5 and 6.5 can also be used.
  • the temperature can be held between 5°C and 45°C, a temperature in the range of 34-41°C being preferred. Storage in the absence of ultra violet light is preferred.
  • aliquots of the composition can be centrifuged, for example, first at a low speed (e.g., about 3500 rpm) and for a short duration (e.g., about 5-10 min) .
  • the resulting supernatant can then be recentrifuged (preferably under refrigeration) at a greater speed (e.g., about 20,000 rpm or up to about 45,000 g) for a longer time (e.g., about an hour or longer) to pellet cell walls and other cellular debris that may be present.
  • a greater speed e.g., about 20,000 rpm or up to about 45,000 g
  • a longer time e.g., about an hour or longer
  • a variety of filtration processes can be used to remove the microorganisms.
  • a series of Thompson E1300 series stainless steel filters ranging from 30 down to 150 mesh can be used first to effect filtration, followed, for example, by a second filtration using the same filter with a 5 ⁇ filter paper and then a third filtration using the same filter with a I ⁇ filter paper.
  • the resulting supernatant can be assayed to ensure that appropriate concentrations of enzymes (e.g., amylase, and hemicellulase and/or glucanase) are present.
  • enzymes e.g., amylase, and hemicellulase and/or glucanase
  • amylase activity is assayed using a colorimetric visual endpoint determination. Kits are commercially available for conducting such assays, one such kit being available from Sigma Chemical Company (e.g., catalog numbers 577-250 and 577-M) .
  • the time required for sucrose-starch mixtures, when treated with iodine solution, to change from blue to reddish-brown is inversely proportional to amylase activity.
  • the amylase activity of the supernatant (filtrate) is preferably at least 45,000-45,4000 ct/ml .
  • Amylase levels can be adjusted by combining aliquots of the composition resulting from the culture process described above, processed to remove intact microorganisms and cellular debris.
  • Hemicellulase levels in the resulting supernatant (filtrate) are advantageously at least 2,000 ct/mg.
  • Hemicellulase levels can be measured using a ⁇ galactase dehydrogenase system (locust bean gum substrate) .
  • a commercial kit for such purpose can be used (e.g., Sigma H 0771) .
  • Table 1 below includes preferred components of the composition of the invention and concentration ranges at which those components can be present in the composition.
  • Table II includes a specific odor-reducing composition suitable for use in the present invention.
  • Citric Acid Food grade
  • HMIS Anhydrous .002%
  • Ferric Chloride Anhydrous .00011%
  • Viscosity can be measured using a viscometric tube, stop watch and temperature using carboxy methylcellulose as the standard.
  • the composition of the invention can be specifically formulated to suit a specific application need.
  • the composition when an odor-producing source is exposed to UV light, the composition can be formulated to include a UV protectant (e.g., sodium alginate, a microbial gum) .
  • a UV protectant e.g., sodium alginate, a microbial gum
  • the composition when the odor-producing source is exposed to drying conditions, the composition can be formulated (e.g., with a gum) so as to render it capable of retaining its moisture content.
  • the composition of the present invention can be used alone or in combination with disinfectants and/or perfumes, depending on the odor problem to be addressed. When such disinfectants, perfumes, etc are used, they can be applied separately or co-formulated with the composition of the invention using known formulating techniques .
  • composition of the present invention can be carried out in a variety of ways as convenient in view of the odor-producing source.
  • the composition can be provided as an aerosol into the atmosphere surrounding the odor-producing source, thereby effecting direct contact with the odor-producing molecules in the air and, upon settling to the ground, with the odor-producing source.
  • the composition can likewise be directly applied to the odor-producing source by, for example, pouring, spraying, etc, the composition onto the source. It will be appreciated that the number of applications of the compositions will vary with the particular odor problem being addressed. In the case of water containment situations such as lagoons, where short carbons chains are involved, the application can be, for example, 1 oz/1000 gallons per week.
  • the odor-producing sources to which the present method and composition have applicability are contaminated with microbes (e.g., anaerobes and aerobes) , the metabolism of which is believed to result in the release of aromatic compounds into the environment that are offensive to the olfactory system. It is also believed that the composition, on contact with the airborne odiferous aromatic compounds, chemically reacts therewith in a manner that results in decomposition of the aromatic compounds to products having an acceptable odor.
  • microbes e.g., anaerobes and aerobes
  • composition of the present invention upon contact with the odor-producing source (e.g., microbially contaminated animals, feces, feed and the like) , is believed to alter the microbial population thereof in favor of aerobic organisms (or metabolism in the case of facilitative anaerobes) . Aerobic metabolism of the source by the microbes present therein is believed to result in the release of more acceptable (from an odor standpoint) products into the environmen .
  • the odor-producing source e.g., microbially contaminated animals, feces, feed and the like
  • the regimen used to apply the composition can be optimized for any particular plant.
  • 1.5 gallons of the composition can be applied per acre to a sugar cane crop per year in approximately four equal applications; about two gallons can be applied per acre of citrus grove per year in two equal applications; about 3 gallons can be applied per acre of golf course grass in two equal applications; and for vegetable crops, about 2-2.5 gallons can be applied per acre per year in two equal applications.
  • about 1 gallon can be applied per acre per month.
  • the composition is, advantageously, diluted about 20:1 with water and applied by the spraying of the diluted composition, however, other modes of application (e.g., irrigation) can also be used.
  • composition results in a significant stimulation of plant growth.
  • the composition can also be applied directly to seed and it can be used as a root drench to decrease transplantation shock and to increase root production
  • the composition can be.applied alone or with other agents, eg an insecticide, herbicide or fungicide.
  • the present invention relates to a composition suitable for enhancing sucrose accumulation in plants (e.g., fruits and vegetables, particularly sugar cane and pineapple) .
  • the composition comprises a culture prepared essentially as described above. During the process of recirculation, for example, through the magnetic field, however, a different formulation is added than that described above.
  • molybdic acid (Mn0 3 ) (0.00001 g/1) ; sulphur (0.00006 g/1) ; iron (Fe3/2 H 2 0 4 S) (0.00006 g/1) ; manganese (MnS0 4 H 2 0) (.030 g/1); boric acid (H 3 Bo 3 ) (.0000002 g/1); nickel carbonate (NiC0 3 ) (.0000005 g/1); cobalt (CoCCH 3 COO) 2 4H 2 0 ( .00000005/gl) ; copper (CuS0 4 ) (0.0000002 g/1); zinc stearate (Zn(C 18 H 35 0 2 ) 2 (0.0004 g/1); silica (Si) (0.004 g/1) and selenium (4(HN0 3 )Se) (0.00000003 g/1).
  • the bacteria can be removed from the composition by centrifferrite, a stearate (Zn(C 18 H 35 0
  • the invention is described herein primarily in the context of a plant growth stimulator or odor- reducer, it is, in a broader sense, a microflora accelerator. It is in this context that the invention further relates to a method of effecting bioremediation and composition suitable for use therein.
  • the composition of the invention targets the sulphur links of short and long chain carbon structures present at a bioremediation site.
  • bioremediation sites include animal waste sites, animal feedlots, lagoons, enclosed facilities (such as dairy parlors, swine barns, chicken houses, aviaries, kennels, etc), human waste treatment plants, land fills, and leachate water runoffs from sanitary facilities.
  • Bioremediation sites can exist in fresh or salt water marine environments. Bioremediation is particularly important when fuel oils, coal tars, napthas or benzene are present in the soil or water (or in the atmosphere) . The present invention is applicable to all such situations.
  • the composition used to effect bioremediation is prepared essentially as described above. Concentrations of specific components, however, will vary depending on the target site.
  • composition invention to a site in need of bioremediation can be made by any suitable application protocol, spraying and fogging being examples.
  • the composition can be simply poured onto and mixed into the contaminated soil or water at a rate that will vary with the nature of the contamination.
  • 5 to 7 oz of the composition of the invention can be applied per 1,000 gallon of leachate; 5 to 15 gallons can be applied per acre foot of soil to be remediated; about 3 gallons of the present composition can be applied per area foot of liquid state petroleum reservoir; and about 5 oz can be applied per 5,000 cubic feet of air.
  • the optimum application protocol for a particular remediation site can be readily determined, as can the optimum composition.
  • the advantages of the present invention result, at least in part, from the effects of components of the present composition on nitrofication which in turn enhances sulphur metabolism.
  • alcohols, aldehydes, organic acids, esters, ketones, phenols and sulphur compounds are believed to be produced.
  • the sulphur compounds are of particular importance.
  • boron and magnesium are expected to stimulate the fermentation process, along with oxygen, nitrogen, and argon (the major components of compressed atmospheric air) . When these constituents are added, they are believed to facilitate the nitrogen cycle of yeast present in the culture.
  • yeast convert sugar in the food substrate and the second food source, alcohols, esters and gums are formed (gums can include alginate, microbial gums, plant exudate and bean gum) .
  • Groups of carbohydrates of particular interest include such gums and cellulose compounds, for example, those derived from the food substrate.
  • the ability of yeast to uptake nitrogen is augmented by certain aerobic bacteria, azotabactors and cyanobactors, along with other nitrogen fixers. These bacteria are believed to be stimulated by molybdenum, boron and magnesium, which elements are believed to be important to the production of gums which, in turn, are important in sulfur metabolism. The importance of molybdenum, boron and magnesium is believed to result from the role played by these elements in the following enzymatic processes.
  • the process of nitrogen fixation requires the nitrogenase complex which consists of a reductase (which provides electrons with high reducing power) and a nitrogenase (which uses these electrons to reduce N 2 to NH4+) .
  • Each component is an iron-sulfur protein in which iron is bonded to the sulfur atom of a cysteine residue and to inorganic sulfide.
  • the nitrogenase component of the complex also contains one or two molybdenum atoms.
  • the conversion of N 2 into NH 4+ by the nitrogenase complex requires ATP and a powerful reductant .
  • the source of high potential electrons in this six-electron reduction is reduced ferredoxin.
  • ATP binds to the reductase and shifts the redox potential of the enzyme from -0.29V to -0.40V by altering its conformation.
  • ATP is hydrolyzed and the reductase dissociates from the nitrogenase component.
  • N 2 bound to the nitrogenase component of the complex is reduced to NH + .
  • a kinase catalyzes the transfer of a phosphoryl groups from ATP to an acceptor.
  • Hexokinase catalyzes the transfer of a phosphoryl group from ATP to a variety of six-carbon sugars.
  • Hexokinase requires Mg 2+ (or another divalent metal ion such as Mn 2+ ) for activity. The divalent metal ion forms a complex with ATP.
  • Acetylcholinesterase is an enzyme that catalyzes the hydrolysis of the ester bond in acetylcholine.
  • Acetate and choline are two important substances in the formation of gums and waxes .
  • Sulfation is defined as any process of introducing an S0 4 group into an organic compound in which the reaction product (sulfate) exhibits the characteristic -OSO 3 - molecular configuration. Sulfation involves the reaction wherein a -COS- linkage is formed by the action of a sulfating agent on an alkene, alcohol, or phenol.
  • the alcohol sulfates are readily susceptible to hydrolysis in acidic media. Sulfation of fatty alcohols and polyalkoxy reductases occurs in the present process and the sulfation products lend themselves to detergent action as emulsifiers . Gums that are produced by the present process can store and stabilize products of microbial sulphur metabolism. Gums also serve generally to stabilize the fermentation components and thus facilitate storage of the product of the present method. The stabilization of the sulphur metabolism components allows immediate reaction with hydrocarbon chains in the environment by removing the sulfur link from those hydrocarbon chains for detergent reaction with associated alcohols.
  • Magnetism can alter the end-product production.
  • bio ⁇ catalysts e.g., proteins, vitamins, trace elements
  • desired sulfur containing compounds are enriched, including ⁇ -linked purines, biotin, sulfinated carbohydrates, etc, and the levels of undesirable gases can be decreased.
  • Magnetics when combined with the present formulation, apparently provide for a more rapid conversion through the nitrogen and sulphur cycles to end complexes of sulphur metabolism.
  • a one liter sample is aspirated from the rumen of a 7.5 year old lactating Holstein cow using a rumen aspirator (Johnson and Johnson) .
  • the sample is taken about 12-14 hours after feeding. Observed microscopically, the sample includes Clostridia, Bacillus, Azotobacter and protozoa (at least 100 cells of each per ml of sample) .
  • the one liter sample is added to a culture medium (169 liters) that includes:
  • the sample and the culture medium (Mixture A) are maintained in Tank 1 (see Figure 1) at a temperature of 37°C and at a pH in the range of 6.9-7.3 for a first 24 hour period. At this stage, and throughout the process, pH adjustments are made using citric acid or sodium bicarbonate, as appropriate. During this first 24 hr period, Mixture (A) is agitated by the injection of compressed air which results in the presence in Mixture (A) of about 3-5 ppm oxygen.
  • Mixture (B) is added to Mixture (A) in Tank 1.
  • Mixture (B) comprises 140 liters of water, pH 7.0-7.1, and 30 liters of the culture medium described above into which air has been injected to achieve an oxygen content of 3-5 ppm.
  • Mixture (B) also includes approximately 20 kg of a substrate comprising the following in approximately equal parts by weight:
  • Substrate crushed corn oats alfalfa whole peanuts
  • the pH of Mixture (B) is maintained at about 7.1 to 7.2, the temperature at about 34-40°C, and the oxygen content at about 3-5 ppm (by injection of compressed air) . These same conditions are maintained after the addition of Mixture (B) to Mixture (A) to form
  • Mixture (C) is maintained in Tank 1 at about 37°C for a second 24 hr period with agitation by compressed air injection.
  • Mixture (D) is added to Mixture (C) in Tank 1 to yield Mixture (E) .
  • Mixture (D) like Mixture (B) , comprises 140 liters of water, pH 7.1 to 7.2, and 30 liters of culture medium. Mixture (D) also includes 20 kg of the substrate described above.
  • Mixture (E) is maintained in Tank 1 for a third 24 hr period with agitation by compressed air injection (temperature 40°C; pH 7.1; oxygen content 3-5 ppm) .
  • Mixture (E) is recirculated for ten minute periods.
  • Tank 1, and the recirculation system associated therewith, is designed such that complete recirculation of Mixture (E) can be effected in the ten minute period. That recirculation is carried out at two hour intervals for a fourth 24 hour period.
  • Mixture (E) is then allowed to stand for a time sufficient to permit particulate matter to settle out.
  • E-T2 (175 liters) is transferred to a further recirculation tank, Tank 3 (see Figure 1) and a "micronutrient" package is added.
  • the contents of the package is formulated so that the addition thereof to the 175 liters results in the following concentrations, expressed as g/1 of Mixture (E-T2) :
  • the content of Tank 3 (Mixture (E-T3)) is recirculated, and, during recirculation, is passed though a magnetic field (an 80 gal/min pump is used in the recirculation process) .
  • the field is generated by six 3500 gauss rare earth magnets oriented as shown in Figure 2 with respect to a 1W diameter PVP 80 gauge recirculation tube, a 1/32" phenolic band being located between the magnets and the tube.
  • the pH is maintained at between 5.5 and 6.5 and the temperature at about 37°C.
  • Compressed air is injected during recirculation to maintain an oxygen content of 3-5 ppm.
  • the composition resulting after 10 minutes of recirculation is stored for about 24 hours at a temperature of 35-38°C.
  • composition resulting from the foregoing process is applied to sugar cane by spraying four times per year for a total annual application of 1.5 gallons per acre.

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  • General Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • Biomedical Technology (AREA)
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  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
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Abstract

La présente invention concerne, de manière générale, un procédé de stimulation de l'activité microbienne ainsi qu'une composition pouvant être utilisée dans cette stimulation. Cette invention concerne en particulier une composition capable de stimuler la croissance de végétaux, ainsi qu'un procédé de préparation d'une telle composition. Cette invention concerne en outre un procédé permettant d'effectuer une biorestauration.
PCT/US1996/009574 1995-06-07 1996-06-05 Procede de preparation d'une culture a l'aide d'un champ magnetique et composition contenant cette culture WO1996039799A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU62625/96A AU6262596A (en) 1995-06-07 1996-06-05 A method of preparing a culture using a magnetic field and a compositon containing the same

Applications Claiming Priority (6)

Application Number Priority Date Filing Date Title
US47610995A 1995-06-07 1995-06-07
US08/476,109 1995-06-07
US08/476,374 1995-06-07
US08/476,374 US5866112A (en) 1995-01-20 1995-06-07 Methods of odor treatment
US08/569,769 1995-12-08
US08/569,769 US5843427A (en) 1995-01-20 1995-12-08 Method of using a plant growth stimulating composition

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102972124A (zh) * 2012-12-15 2013-03-20 安徽科技学院 一种采用稀土植物生长灯进行萝卜芽苗菜室内生产的方法
RU2479494C2 (ru) * 2008-09-22 2013-04-20 Вильям Стивен ЛОПЕС Установка обработки магнитным полем для кондиционирования текучих сред
CN103086793A (zh) * 2013-01-10 2013-05-08 山西昌鑫生物农业科技有限公司 一种氨基酸生物水溶肥料及其制备方法
CN105944827A (zh) * 2016-04-21 2016-09-21 洛南县恒丰非金属矿业有限公司 一种从选钼尾矿再选稀土预选抛尾的方法

Citations (12)

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SU324680A1 (ru) * Е. Гак , Т. Г. Жгенти Способ обработки объектов малого объема магнитными полями большой неоднородности
US2196361A (en) * 1936-07-31 1940-04-09 Liebesny Paul Method of promoting the propagation and the activity of microorganisms
US2767072A (en) * 1951-04-13 1956-10-16 Coanda Henri Soil regeneration
US3095359A (en) * 1959-11-16 1963-06-25 New England Inst For Medical R High-frequency treatment of matter
US3623265A (en) * 1969-11-26 1971-11-30 Nitron Inc Method of fixing nitrogen in the atmosphere and the soil
US3675367A (en) * 1970-07-27 1972-07-11 Raymond D Amburn Apparatus for magnetically treating seeds
US3871961A (en) * 1973-03-15 1975-03-18 Matilde Gianessi Method for accelerating the growth and increasing the yield of microorganisms
US4487766A (en) * 1982-02-26 1984-12-11 Lohmann Tierernahrung Gmbh Growth stimulants and compositions containing same
US4508625A (en) * 1982-10-18 1985-04-02 Graham Marshall D Magnetic separation using chelated magnetic ions
US4879045A (en) * 1986-01-13 1989-11-07 Eggerichs Terry L Method and apparatus for electromagnetically treating a fluid
US4915915A (en) * 1988-10-11 1990-04-10 Treharne Richard W Water-powdered piezoelectric unit for producing nitrogen fertilizer
US4938875A (en) * 1987-01-28 1990-07-03 Philippe Niessen Method and apparatus for magnetically treating a liquid

Patent Citations (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
SU324680A1 (ru) * Е. Гак , Т. Г. Жгенти Способ обработки объектов малого объема магнитными полями большой неоднородности
US2196361A (en) * 1936-07-31 1940-04-09 Liebesny Paul Method of promoting the propagation and the activity of microorganisms
US2767072A (en) * 1951-04-13 1956-10-16 Coanda Henri Soil regeneration
US3095359A (en) * 1959-11-16 1963-06-25 New England Inst For Medical R High-frequency treatment of matter
US3623265A (en) * 1969-11-26 1971-11-30 Nitron Inc Method of fixing nitrogen in the atmosphere and the soil
US3675367A (en) * 1970-07-27 1972-07-11 Raymond D Amburn Apparatus for magnetically treating seeds
US3871961A (en) * 1973-03-15 1975-03-18 Matilde Gianessi Method for accelerating the growth and increasing the yield of microorganisms
US4487766A (en) * 1982-02-26 1984-12-11 Lohmann Tierernahrung Gmbh Growth stimulants and compositions containing same
US4508625A (en) * 1982-10-18 1985-04-02 Graham Marshall D Magnetic separation using chelated magnetic ions
US4879045A (en) * 1986-01-13 1989-11-07 Eggerichs Terry L Method and apparatus for electromagnetically treating a fluid
US4938875A (en) * 1987-01-28 1990-07-03 Philippe Niessen Method and apparatus for magnetically treating a liquid
US4915915A (en) * 1988-10-11 1990-04-10 Treharne Richard W Water-powdered piezoelectric unit for producing nitrogen fertilizer

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
RU2479494C2 (ru) * 2008-09-22 2013-04-20 Вильям Стивен ЛОПЕС Установка обработки магнитным полем для кондиционирования текучих сред
CN102972124A (zh) * 2012-12-15 2013-03-20 安徽科技学院 一种采用稀土植物生长灯进行萝卜芽苗菜室内生产的方法
CN103086793A (zh) * 2013-01-10 2013-05-08 山西昌鑫生物农业科技有限公司 一种氨基酸生物水溶肥料及其制备方法
CN105944827A (zh) * 2016-04-21 2016-09-21 洛南县恒丰非金属矿业有限公司 一种从选钼尾矿再选稀土预选抛尾的方法

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