microbial products and their uses to enhance oil recovery

专利摘要:
the present invention relates to compositions and methods of enhanced microbial recovery of oil using biochemicals producing microbes. in specific embodiments, the methods of the present invention comprise the application of a biotensoactive-producing bacterium and / or a by-product of its growth to an oil production site. in preferred modalities, the bacterium is a strain of bacillus in the form of spores. in some embodiments, the methods further comprise applying bacteria with a yeast fermentation product, an alkaline compound, a polymer, a non-biological surfactant, and / or one or more chelating agents. advantageously, the present invention can be useful for stimulating the flow of oil from a well, as well as dissolving scale present in an oil-containing formation.
公开号:BR112019020271A2
申请号:R112019020271
申请日:2018-04-09
公开日:2020-04-22
发明作者:Alibek Ken；Adams Kent；Milovanovic Maja；Farmer Sean；MAZUMDER Sharmistha；Dixon Tyler；Chen Yajie
申请人:Locus Oil Ip Company Llc；
IPC主号:

专利说明:

Descriptive Report on the Invention Patent for MICROBIAL PRODUCTS AND USES OF THE SAME TO IMPROVE OIL RECOVERY.
CROSS REFERENCE TO RELATED ORDERS [001] This claim claims the benefit of the provisional US order. No. 62 / 483,425, filed on April 9, 2017, which is incorporated herein by reference in its entirety.
BACKGROUND OF THE INVENTION [002] The high demand for fossil fuels requires efficient oil production. As the oil wells mature, it becomes more difficult and expensive to continue pumping oil at an economically viable rate. Therefore, it is necessary to develop improved methods of oil recovery. One of these mechanisms uses microbes and their by-products.
[003] Oil exists in small pores and narrow cracks within the body of the reservoir rocks under the earth's surface. The natural pressure of the reservoir causes the oil to flow to the surface, thus providing primary production; however, as oil production progresses, reservoir pressure is reduced to a point where elevation or artificial pumping is required to maintain an economic rate of oil production.
[004] When it is necessary to supply external energy to the reservoir to obtain additional oil recovery (secondary recovery), the extra energy can be introduced by injecting gas (gas injection) and / or water (flooding of water). After a few years of operation in a field, the injected fluids preferentially flow through highly permeable layers that cause these fluids to deviate from saturated areas of oil in the reservoir. Therefore, an increasing amount of water (or gas) increases with oil and, decreasing the oil / water ratio, ends up not becoming
Petition 870190096868, of 9/27/2019, p. 10/117
2/50 it is economical to continue the process; at that point the field must be abandoned.
[005] Primary recovery generally results in an average recovery of only a fraction of the oil originally present in an oiled formation. Secondary recovery, for example, water flooding, usually recovers another 10% by the time it becomes uneconomic to continue. It is not uncommon for 60 to 70% of the oil originally in the formation to remain, even after the secondary recovery reaches the economic limit. In this situation, a third stage of oil recovery, called tertiary production, can be considered.
[006] In this tertiary stage, technically advanced methods are used to modify either the properties of the reservoir fluids or the characteristics of the reservoir rock. In general, the methods can be classified into four main categories: thermal methods, chemical methods, miscible or solvent injection, and microbial methods.
[007] Enhanced Petroleum Recovery by Microbial Action (MEOR) is a multidisciplinary field that incorporates, among others, geology, chemistry, microbiology, fluid mechanics, petroleum engineering, environmental engineering and chemical engineering. MEOR uses microorganisms and / or their metabolites to improve oil recovery. The microbial methods used in MEOR can address cleaning the well in order to remove mud and other debris that block the channels through which oil flows; stimulation from the well improves the flow of oil from the drainage area to the well; and improved water floods increase microbial activity by injecting selected microbes and sometimes nutrients.
[008] In MEOR, the proper nutrients and microbes, which preferably grow under the anaerobic conditions of the reservoir,
Petition 870190096868, of 9/27/2019, p. 10/127
3/50 are injected into the reservoir. Microbial by-products, which may include biotensives, biopolymers, acids, solvents, gases and enzymes, for example, can modify the properties of oil and the interactions between oil, water and the porous medium, alter the permeability of underground formations and, finally , increase mobility and oil recovery.
[009] Interest in microbial surfactants has steadily increased in recent years due to its diversity, ecological nature, the possibility of large-scale production, selectivity, performance under extreme conditions, and potential applications in environmental protection. The surfactants produced in a microbial way, that is, biotensoactives, reduce the interfacial tension between water and oil and, therefore, a lower hydrostatic pressure is necessary to move the liquid trapped in the pores to overcome the capillary effect. Second, biotensoatives contribute to the formation of micelles, providing a physical mechanism to mobilize oil in a mobile aqueous phase.
[0010] There is a continuing need for improved methods of oil recovery, specific methods that can be sustained for long periods of time. This includes the need for improved methods of enhanced oil recovery, such as methods that use, for example, microorganisms and / or their growth by-products. Biotensics increase the emulsification of hydrocarbons, have the potential to solubilize contaminants from hydrocarbons and increase their availability for microbial degradation. These compounds can also be used in enhanced oil recovery.
BRIEF SUMMARY OF THE INVENTION [0011] In certain embodiments, the present invention provides microbes, as well as substances, such as biotensive agents, solvents
Petition 870190096868, of 9/27/2019, p. 10/13
4/50 and / or enzymes, derived from these microbes and the fermentation broth in which they are produced. The present invention also provides methods of using these microbes and their by-products in improved oil production.
[0012] Specifically, the present invention provides economical and ecological approaches to improve oil recovery. Advantageously, these methods can be practiced over a wide temperature range, including 20 Ό to 70 Ό, and above.
[0013] In some embodiments, the present invention provides materials and methods to improve oil production, treating an oil production site, for example, a formation containing oil or an oil well, with microorganisms and / or their growth by-products. In one embodiment, the present invention may be useful for enhancing oil recovery from an oil well, by stimulating, for example, the flow of oil from the well while dissolving scale within the formation. [0014] In some embodiments, the present invention uses by-products of yeast growth, such as, for example, biotensives. Biotensics are useful in the oil and gas industry for their ability to improve oil recovery. Biotensics can modify the properties of oil and the interactions between oil, water and the porous media in which oil and gas originate, thereby increasing mobility and, consequently, oil recovery. Thus, the compositions and methods of the present invention can increase the recovery of crude oil and natural gas from formations containing oil and gas, dramatically reducing the surface and interfacial tension between substances in the formations and altering the wettability of the formations.
[0015] In one embodiment, the present invention provides products
Petition 870190096868, of 9/27/2019, p. 10/147
5/50 yeast fermentation to improve oil recovery from an oil-containing formation. In one embodiment, the yeast fermentation product is obtained through the cultivation of yeast producing bio-surfactants using processes that vary from small to large scale. The cultivation process can be, for example, submerged cultivation, solid state fermentation (SSF) and / or a combination thereof. In one embodiment, yeast products are grown using a simplified yeast fermentation technique, which reduces cultivation time by 50% and reduces carbon source supplementation.
[0016] The yeast fermentation product can be obtained by cultivating a yeast from the production of biochemicals, such as, for example, Pichia anomalous (Wickerhamomyces an o malus). The fermentation broth after 7 days of cultivation at 25 Ό to 30 Ό may contain the yeast cell suspension and, for example, 4 / g / L or more of glycolipid biotensoactives.
[0017] The yeast fermentation product can also be obtained by cultivating the biotensoactive yeast, Starmerella bombicola. The fermentation broth after 5 days of cultivation at 25 Ό may contain the yeast cell suspension and, for example, 150 / g / L or more of glycolipid biotensoactive.
[0018] The yeast fermentation product may comprise the fermentation broth, separated from the yeast cells. In one embodiment, biotensives or other growth by-products in the broth are further separated from the broth and purified.
[0019] In some embodiments, the present invention uses strains of bacteria and their by-products. Such by-products may include, for example, metabolites, polymers, biotensives, enzymes, organic acids and solvents. In certain modalities, bacteria are strains of Bacillus that thrive in environments with a high salt content,
Petition 870190096868, of 9/27/2019, p. 10/157
6/50 such as those often found at an oil extraction site. In certain embodiments, the bacteria are strains of Bacillus overproducing surfactants, which means that these strains are characterized by improved production of biotensives compared to naturally occurring Bacillus strains. In certain modalities, strains of Bacillus increased the production of enzymes. [0020] In one embodiment, the microorganisms are strains of Bacillus subtil is, Bacillus licheniformis and / or Bacillus amyloliquefaciens. In preferred embodiments, the bacteria are in the form of spores.
[0021] In some modalities, strains of Bacillus are able to thrive under conditions of low oxygen, thus facilitating growth under microaerophilic and anaerobic conditions. Under anaerobic conditions, nitrate salts can be added to replace oxygen as an electron acceptor to support microaerophilic and / or anaerobic respiration.
[0022] In one embodiment, strains of Bacillus subtilis are, for example, B. subtilis var. locuses of strains B1 and B2, which are effective producers of the amphiphilic lipopeptide surfactin.
[0023] In one embodiment, the present invention provides a method for enhancing oil recovery by applying one or more microorganisms capable of producing useful biochemical by-products at an oil production site, for example, a formation containing oil and / or oil well. The method optionally includes adding nutrients and / or other agents to the site. In certain embodiments, microorganisms are selected from strains of Bacillus, including, but not limited to, strains of Bacillus subtilis, Bacillus licheniformis and Bacillus amyloliquefaciens. In preferred embodiments, the bacteria are in the form of spores.
[0024] The method can also comprise adding a pro
Petition 870190096868, of 9/27/2019, p. 10/167
7/50 yeast fermentation duct, such as the fermentation broth resulting from the cultivation, for example, of Starmerella bombicola or Wickerhamomyces anomalus. In one embodiment, yeast cells can be removed from the yeast fermentation product and only the broth containing biotensoactives and other cellular exudates is applied. In one embodiment, the yeast fermentation product comprises biotensoatives that have been separated from the fermentation broth and purified.
[0025] The method can also comprise the application of microbes and / or by-products of microbial growth with one or more alkaline compounds. The alkaline compound can be, for example, ammonium hydroxide.
[0026] In some embodiments, the method may also comprise the application of microbes and / or by-products of microbial growth with one or more polymeric compounds. Polymeric compounds can be selected from biopolymers, such as, for example, hydrogels, polysaccharides, xanthan gum, guar gum and cellulose polymers.
[0027] In some modalities, the method may also include the application of microbes and / or by-products of microbial growth with one or more non-biological surfactants. Surfactants can be, for example, anionic, cationic, non-ionic or zwitterionic.
[0028] In one embodiment, microbes and / or by-products of microbial growth can be applied with one or more chelating agents to reduce, for example, dissolve, the scale that has accumulated within a formation containing oil. Chelating agents can be, for example, citric acid, EDTA, sodium citrate and / or a combination thereof.
[0029] In one embodiment, microorganisms can germinate
Petition 870190096868, of 9/27/2019, p. 10/177
8/50 and grow on site within a formation containing oil or oil well, and produce biotensoactive in it. Consequently, a high concentration of biotensoactive and micro-organisms producing biotensoactives in a treatment site (for example, an oil well) can be achieved easily and continuously.
[0030] In some embodiments, the present microbial products and methods in question can also be used for removing paraffin, liquefying solid asphaltene, and bioremediation of waters, soils and other sites contaminated with hydrocarbons. For such uses, the methods may further comprise the addition of solvents, such as isopropyl alcohol or ethanol, with the microbes and / or by-products of microbial growth.
[0031] In one embodiment, the present invention provides methods of producing a biotensoactive by cultivating a microbe strain of the present invention under conditions suitable for the growth and production of the biotensoactive; and purification of the biotensoactive. The present invention also provides methods for producing solvents, enzymes or other proteins by cultivating a strain of microbes of the present invention under conditions suitable for the growth and expression of solvent, enzyme or protein; and purifying the solvent, enzyme or other protein.
[0032] The microbial products of the present invention can be used in a variety of unique configurations due, for example, to the ability to efficiently provide fresh fermentation broth with active metabolites; a mixture of cells and fermentation broth; compositions with a high cell density; microbial-based products in the short term; and microbial products at remote sites.
[0033] Advantageously, the present invention can be used without releasing large amounts of inorganic compounds into the environment.
Petition 870190096868, of 9/27/2019, p. 10/187
9/50
In addition, the claimed compositions and methods use components that are biodegradable and toxicologically safe. Thus, the present invention can be used in the production of oil and gas (and other industries) as an ecological treatment.
DETAILED DESCRIPTION [0034] In certain embodiments, the present invention provides microbes, as well as substances, such as biotensoatives, solvents and / or enzymes, derived from these microbes and the fermentation broth in which they are produced. The present invention also provides methods of using these microbes and their by-products in improved oil production.
[0035] Specifically, the present invention provides economical and ecological approaches to improve oil recovery. Advantageously, these methods can be practiced over a wide temperature range, including 20 Ό to 70 Ό, and above.
[0036] In some embodiments, the present invention provides materials and methods for improving oil production by treating a petroleum production site with microorganisms and / or their growth by-products. In one embodiment, the present invention may be useful for enhancing oil recovery from an oil-containing formation or from an oil well, by stimulating, for example, the flow of oil from the formation or well, while dissolving the scale.
[0037] In some embodiments, the present invention uses by-products of yeast growth, such as, for example, biotensives. Biotensics are useful in the oil and gas industry for their ability to improve oil recovery. Biotensics can modify the properties of oil and the interactions between oil, water and the porous media in which oil and gas originate, thus increasing mobility and, consequently,
Petition 870190096868, of 9/27/2019, p. 10/197
10/50 feared, oil recovery. Thus, the compositions and methods of the present invention can increase the recovery of crude oil and natural gas from formations containing oil and gas, dramatically reducing both the surface tension and the interfacial tension between substances within the formations and changing the wettability of the formations.
[0038] In specific modalities, the methods and compositions described here use microorganisms to improve oil recovery. Microorganisms can improve the quantity and quality of oil recovered from reservoirs, including those considered mature. In addition, microorganisms can remove toxic substances from oil production sites.
[0039] In some embodiments, the present invention provides materials and methods for improving oil production by treating a petroleum production site with microorganisms and / or their growth by-products. In one embodiment, the present invention may be useful for enhancing oil recovery from a formation containing an oil or from an oil well, by stimulating, for example, the flow of oil from the well or formation.
[0040] In one embodiment, the present invention provides a method for enhancing oil recovery by applying one or more microorganisms capable of producing useful biochemical by-products at an oil production site, for example, a formation containing oil and / or oil well. The method optionally includes adding nutrients and / or other agents to the site. In certain embodiments, microorganisms are selected from strains of Bacillus, including, but not limited to, strains of Bacillus subtilis, Bacillus licheniformis and Bacillus amyloliquefaciens. In preferred embodiments, the bacteria are in the form of spores.
Petition 870190096868, of 9/27/2019, p. 10/20
11/50 [0041] In certain modalities, bacteria are strains of Bacillus that thrive in environments with a high salt content, such as those often found in an oil extraction site. In certain embodiments, the bacteria are strains of Bacillus that produce surfactants, which means that these strains are characterized by improved production of biotensoactive compared to naturally occurring Bacillus strains. In certain modalities, strains of Bacillus increased the production of enzymes.
[0042] The method may also comprise the addition of yeast fermentation products, such as the fermentation broth resulting from the cultivation of biochemical-producing yeasts, for example, Starmerella bombicola or Wickerhamomyces anomalus. In one embodiment, the yeast fermentation product comprises purified biotensoatives produced by these yeasts.
[0043] The method can also comprise the application of microbes with one or more alkaline compounds, polymers, surfactants and / or chelating agents.
Selected Definitions [0044] As used herein, reference to a microbial-based composition means a composition that comprises components that have been produced as a result of the growth of microorganisms or other cell cultures. Thus, the microbial-based composition can comprise the microbes themselves and / or by-products of microbial growth. The microbes can be in the vegetative state, in the form of spores, in the mycelial form, in any other form of propagule, or in a mixture of them. The microbes can be planktonic or they can be in the form of biofilm, or a mixture of both. Growth by-products can be, for example, metabolites (for example, biotensives), cell membrane components, expressed proteins, and / or other ce components
Petition 870190096868, of 9/27/2019, p. 10/21
12/50 lulars. The microbes can be intact or lysed. The cells may be absent or may be present, for example, in a concentration of 1 x 10 ⁴ , 1 x 10 ⁵ , 1 x 10 ⁶ , 1 x 10 ⁷ , 1 x 10 ⁸ , 1 x 10 ⁹ , 1 x 10 ¹⁰ or 1 x 10 ¹¹ or more cells or propagules per milliliter of the composition. As used here, a propagule is any portion of a microorganism from which a new and / or mature organism can grow, including, but not limited to, cells, conidia, cysts, spores (eg, reproductive spores, endospores and exospores), mycelia, sprouts and seeds.
[0045] The present invention also provides products based on microbes, which are products that must be applied in practice to achieve the desired result. The microbial product may simply be the microbial composition harvested from the microbial culture process. Alternatively, the microbial product may comprise other ingredients that have been added. Such additional ingredients may include, for example, stabilizers, buffers, appropriate vehicles, such as water, saline or any other appropriate vehicle, nutrients added to support additional microbial growth, non-nutritive growth enhancers, such as plant hormones and / or agents that facilitate the tracking of microbes and / or composition in the environment in which it is applied. The microbial product can also comprise mixtures of microbial compositions. The microbial product can also comprise one or more components of a microbial composition that have been processed in some way, such as, but not limited to, filtration, centrifugation, lysis, drying, purification and the like.
[0046] As used here, harvested refers to the removal of part or all of the microbial-based composition from a growth vessel.
Petition 870190096868, of 9/27/2019, p. 10/22
13/50 [0047] In some embodiments, the microbes used in accordance with the present invention are super-producers of surfactant. For example, the strain can produce at least 0.1 to 10 / g / L, for example, 0.5 to 1 / g / L of surfactant. For example, bacteria produce at least 10%, 25%, 50%, 100%, 2 times, 5 times, 7.5 times, 10 times, 12 times, 15 times or more compared to other microbial strains recovering Petroleum. Specifically, Bacillus subtilis ATCC 39307 is used here as a reference strain.
[0048] As used herein, an isolated or purified nucleic acid molecule, polynucleotide, polypeptide, protein or organic compound, such as a small molecule (for example, those described below), is substantially free of other compounds, such as cellular material , with which it is associated in nature. The reference to isolate in the context of a microorganism strain means that the strain is removed from the environment in which it exists in nature. Thus, the isolated strain can exist as, for example, a biologically pure culture, or as spores (or other forms of the strain) in association with a vehicle. A purified or isolated polynucleotide (ribonucleic acid (RNA) or deoxyribonucleic acid (DNA)) is free of the genes or sequences that flank it in its natural state. A purified or isolated polypeptide is free of the naturally occurring amino acids or sequences that flank it.
[0049] In certain embodiments, the purified compounds are at least 60% by weight (dry weight) of the compound of interest. Preferably, the preparation is at least 75%, more preferably at least 90% and most preferably at least 99%, by weight of the compound of interest. For example, a purified compound is one that has at least 90%, 91%, 92%, 93%, 94%, 95%, 98%, 99% or 100% (by weight) of the desired compound by weight. Purity is measured by any appropriate standard method, for example,
Petition 870190096868, of 9/27/2019, p. 10/23
14/50 by column chromatography, thin layer chromatography, or high performance liquid chromatography (HPLC) analysis.
[0050] A metabolite refers to any substance produced by metabolism or necessary to participate in a specific metabolic process. A metabolite can be an organic compound that is a starting material (for example, glucose), an intermediate (for example, acetyl-CoA) or an end product (for example, n-butanol) of metabolism. Examples of metabolites may include, but are not limited to, enzymes, toxins, acids, solvents, alcohols, proteins, carbohydrates, vitamins, minerals, microelements, amino acids, polymers and surfactants.
[0051] Modular means changing (increasing or decreasing). Such changes are detected by methods known in the standard art, such as those described herein.
[0052] The ranges provided here are understood as shortcuts to all values within the range. For example, a range from 1 to 50 is understood to include any number, combination of numbers or sub-range of the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 , 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 25, 26, 27, 28, 29, 30, 31,32, 33, 34, 35, 36, 37 , 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50, as well as all intermediate decimal values between the whole numbers mentioned above, such as, for example, 1.1 , 1,2,
1.3, 1.4, 1.5, 1.6, 1.7, 1.8 and 1.9. With respect to the sub-bands, nested sub-bands that extend from any end point of the range are specifically considered. For example, a nested subrange of an exemplary range from 1 to 50 can comprise 1 to 10, 1 to 20, 1 to 30 and 1 to 40 in one direction or 50 to 40, 50 to 30, 50 to 20 and 50 to 10 in the other direction.
[0053] By reduction means a negative change of at least 1%, 5%, 10%, 25%, 50%, 75% or 100%.
Petition 870190096868, of 9/27/2019, p. 10/24
15/50 [0054] By reference is meant a standard or control condition.
[0055] By salt tolerant is meant a microbial strain capable of growing in a sodium chloride concentration of fifteen (15) percent or more. In a specific modality, salt tolerant refers to the ability to grow by 150 / g / L or more of NaCI.
[0056] As used here, a biofilm is a complex aggregate of microorganisms, such as bacteria, in which cells adhere to one another. The cells in biofilms are physiologically distinct from planktonic cells in the same organism, which are unique cells that can float or swim in a liquid medium.
[0057] As used here, surfactant refers to a compound that reduces the surface tension (or interfacial tension) between two liquids or between a liquid and a solid. Surfactants act as detergents, agents, wetting agents, emulsifiers, foaming agents and / or dispersants. A surfactant produced by microorganisms is called a bio-surfactant.
[0058] As used in this document, oil production refers to any and all operations involved in the extraction of hydrocarbons, such as crude oil or natural gas from a formation, through its eventual processing and use by consumers. Oil production may include, but is not limited to, drilling, pumping, recovery, transmission, processing, refining, transportation and storage of hydrocarbons.
[0059] An oil production site refers to any environment or structure, naturally occurring or artificial, in which one or more aspects of the recovery of hydrocarbons, oil and / or natural gas occurs, including, among others, underground formations , formations containing oil and gas, and wells.
[0060] As used here, inlay refers to accumulations outside
Petition 870190096868, of 9/27/2019, p. 10/25
16/50, for example, by deposits of precipitated mineral salts, which may arise as a result, for example, of changes in the pressure, composition and / or temperature of crude oil. Fouling may result from precipitates, for example, barium sulfate, calcium carbonate, strontium sulfate, calcium sulfate, sodium chloride, silicon dioxide, iron sulfide, iron oxides, iron carbonate, silicates, phosphates and oxides or any one of several insoluble or slightly water-soluble compounds.
[0061] As used here, improving oil recovery includes improving oil and hydrocarbon recovery and means increasing the amount of hydrocarbons produced and / or increasing the rate at which they are produced, for example, by stimulating the flow of oil from from the well.
[0062] The transitional term comprising, which is synonymous with including or containing, is inclusive or open and does not exclude additional elements not mentioned or steps in the method. On the other hand, the transition phrase consisting of excludes any element, step or ingredient not specified in the claim. The transition phrase essentially consists of limiting the scope of a claim to the specified materials or steps and to those that do not materially affect the basic and new features of the claimed invention.
[0063] Unless it is specifically stated or obvious from the context, as used here, the term is either understood as inclusive. Unless it is specifically stated or obvious in the context, as used here, the terms one, one and o, a are understood to be singular or plural.
[0064] Unless it is specifically stated or obvious from the context, as used here, the term about is understood to be within a normal tolerance range in the art, for example, within 2 standard deviations from the mean. About can be understood
Petition 870190096868, of 9/27/2019, p. 10/26
17/50 given as within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05 % or 0.01% of the declared value.
[0065] The citation of a listing of chemical groups in any definition of a variable cited here includes definitions of that variable as any single group or combination of groups listed. The citation of a modality for a variable or aspect mentioned here includes that modality as any single modality or in combination with any other modalities or parts thereof.
[0066] Any compositions or methods provided herein can be combined with one or more of any of the other compositions and methods provided here.
[0067] Other features and advantages of the invention will be evident from the description below of the preferred modalities of the same and the claims. All references cited herein are incorporated by reference.
By-products of Microbial Growth in accordance with the Present Invention [0068] In preferred embodiments, a composition is provided to enhance and / or improve the recovery of oil from an oil production site, where the composition comprises one or more micro - growth organisms and / or by-products. In specific modalities, microorganisms are capable of producing, and are used to produce, one or more biotensives.
[0069] Biotensive agents are a structurally diverse group of active surface substances produced by microorganisms. Biotensics are biodegradable and can be produced easily and inexpensively using organisms selected on renewable substrates. Most of the biotensoative-producing organisms produce biotensoatives in response to the presence of a hydro source
Petition 870190096868, of 9/27/2019, p. 10/277
18/50 carbide (for example, oils, sugar, glycerol, etc.) in the culture media. Other components of the media, such as iron concentration, can also significantly affect the production of biotensoactive.
[0070] All biotensives are amphiphilic. They consist of two parts: a polar (hydrophilic) portion and a non-polar (hydrophobic) portion. Due to their amphiphilic structure, biotensoatives increase the surface area of hydrophobic substances insoluble in water, increase the water bioavailability of such substances, and alter the properties of bacterial cell surfaces.
[0071] Biotensive agents accumulate at the interfaces, thus reducing interfacial tension and leading to the formation of aggregated micellular structures in solution. The ability of biotensoactives to form pores and destabilize biological membranes allows their use as antibacterial, antifungal and hemolytic agents. Combined with the characteristics of low toxicity and biodegradability, biotensoactives are advantageous for use in the oil and gas industry in several applications. These applications include, but are not limited to, improving crude oil recovery; reduced oil viscosity; paraffin removal from rods, pipes, liners and pumps; corrosion prevention of oil equipment; fracturing fluids; reducing the concentration of HES in the extracted crude oil; as well as cleaning tanks, flow lines and pipes.
[0072] Safe and effective microbial biotensoatives reduce surface and interfacial tension between molecules of liquids, solids and gases. As discussed here, this activity can be highly beneficial in the context of oil recovery.
[0073] The bio-surfactants produced in accordance with the present invention can be used for purposes other than recovery
Petition 870190096868, of 9/27/2019, p. 10/28
19/50 of oil, including, for example, cleaning pipes, reactors and other machines or surfaces.
[0074] Biotensive agents include low molecular weight glycolipids (GLs), lipopeptides (LPs), flavolipids (FLs), phospholipids and high molecular weight polymers such as lipoproteins, lipopolysaccharide - protein complexes, and polysaccharide - protein complexes fatty acids. The hydrocarbon chain of a fatty acid acts like the common lipophilic portion of a biotensoactive molecule, while the hydrophilic portion is formed by ester or alcohol groups of neutral lipids, by the carboxylate group of fatty acids or amino acids (or peptides), organic acid in the case of flavolipids, or, in the case of glycolipids, by the carbohydrate.
[0075] In one embodiment, the microbial biotensoatives according to the present invention include glycolipids, such as ramnolipids (RLP), soforolipids (SLP), trehalose lipids or mannosileritritol lipids (MEL).
[0076] In one embodiment, the microbial biotensoactive is a lipopeptide, such as, for example, surfactin or iturine A.
[0077] Microbial biotensoatives are produced by a variety of microorganisms, such as bacteria, fungi and yeasts. Examples of biotensive-producing microorganisms include species of Pseudomonas (P. aeruginosa, P. putida, P. floresens, P. fragi, P. syringae); Pseudozyma (P. aphidis) Flavobacterium spp .; Pichia spp. (P. anomalous, P. lynferdii, P. guilliermondii, P. sydowiorum), Bacillus spp. (B. subtilis, B. amyloliquefaciens, B. pumillus, B. cereus,
B. licheniformis); Wickerhamomyces spp. (W. anomalus), Starmerella spp. (S. good beak there), Candida spp. (C. albicans, C. rugosa, C. tropical is,
C. lipolytica, C. torulopsis); Rhodococcus spp .; Arthrobacter spp .; Campylobacter spp .; Cornybacterium spp., And so on. Biotensive agents can be obtained by fermentation processes known
Petition 870190096868, of 9/27/2019, p. 10/29
20/50 acids in the technique.
Growth of Microbes According to the Present Invention [0078] The present invention uses methods for the cultivation of microorganisms and the production of microbial metabolites and / or other by-products of microbial growth. The present invention further uses cultivation processes that are suitable for the cultivation of microorganisms and the production of microbial metabolites on a desired scale. Microbial culture systems would typically use submerged culture fermentation; however, surface culture and hybrid systems can also be used. As used here, fermentation refers to the growth of cells under controlled conditions. Growth can be aerobic or anaerobic.
[0079] In one embodiment, the present invention provides materials and methods for the production of biomass (for example, viable cellular material), extracellular metabolites (for example, small molecules and proteins excreted), residual nutrients and / or intracellular components (for example, enzymes and other proteins).
[0080] The microbial growth vessel used in accordance with the present invention can be any fermentor or cultivation reactor for industrial use. In one embodiment, the vessel may have functional controls / sensors or may be connected to functional controls / sensors to measure important factors in the cultivation process, such as pH, oxygen, pressure, temperature, agitator shaft power, humidity, viscosity and / or microbial density and / or concentration of metabolites.
[0081] In an additional embodiment, the vessel may also be able to monitor the growth of microorganisms within the vessel (for example, measuring the number of cells and growth phases). Alternatively, a daily sample can be taken from the vessel and subjected to enumeration by known techniques, such as the technique
Petition 870190096868, of 9/27/2019, p. 10/30
21/50 dilution in plates. Plaque dilution is a simple technique used to estimate the number of bacteria in a sample. The technique can also provide an index by which different environments or treatments can be compared.
[0082] In one embodiment, the method includes supplementing the crop with a nitrogen source. The nitrogen source can be, for example, potassium nitrate, ammonium nitrate, ammonium sulfate, ammonium phosphate, ammonia, urea and / or ammonium chloride. These nitrogen sources can be used independently or in a combination of two or more.
[0083] The cultivation method can provide oxygenation for the growing culture. One method uses slow air movement to remove air containing low oxygen concentration and introduce oxygenated air. The oxygenated air can be ambient air supplemented daily by means of mechanisms, including impellers for mechanical agitation of the liquid, and air sprinklers to supply gas bubbles to the liquid to dissolve oxygen in the liquid.
[0084] The method can also include supplementing the cultivation with a carbon source. The carbon source is typically a carbohydrate, such as glucose, sucrose, lactose, fructose, trehalose, mannose, mannitol and / or maltose; organic acids such as acetic acid, fumaric acid, citric acid, propionic acid, malic acid, malonic acid and / or pyruvic acid; alcohols such as ethanol, propanol, butanol, pentanol, hexanol, isobutanol and / or glycerol; fats and oils such as soybean oil, rice bran oil, olive oil, canola oil, coconut oil, corn oil, sesame oil and / or linseed oil. These carbon sources can be used independently or in a combination of two or more.
[0085] In one embodiment, growth factors and trace nutrients for microorganisms are included in the medium. This is private
Petition 870190096868, of 27/09/2019, p. 10/317
22/50 preferentially in the cultivation of microbes that are unable to produce all the vitamins they need. Inorganic nutrients, including trace elements such as iron, zinc, copper, manganese, molybdenum and / or cobalt can also be included in the medium. In addition, sources of vitamins, essential amino acids and microelements can be included, for example, in the form of flour or meals, such as corn flour, or in the form of extracts, such as yeast extract, potato extract, meat extract beef, soy extract, banana peel extract, and the like, or in purified forms. Amino acids, such as, for example, those useful for protein biosynthesis, can also be included, for example, L-alanine.
[0086] In one embodiment, inorganic salts can also be included. Usable inorganic salts can be potassium dihydrogen phosphate, dipotassium hydrogen phosphate, disodium hydrogen phosphate, magnesium sulphate, magnesium chloride, iron sulphate, iron chloride, manganese sulphate, manganese chloride, zinc sulphate, lead chloride, sulphate of lead copper, calcium chloride, calcium carbonate, and / or sodium carbonate. These inorganic salts can be used independently or in a combination of two or more.
[0087] In some embodiments, the method for cultivation may further comprise the addition of additional acids and / or antimicrobials in the liquid medium before and / or during the cultivation process. Antimicrobial agents or antibiotics can be used to protect the culture from contamination. In addition, anti-foaming agents can also be added to prevent foam formation and / or build-up when the gas is produced during cultivation.
[0088] The pH of the mixture must be suitable for the microorganism of interest. Buffers and pH regulators, such as carbonates and phosphates, can be used to stabilize the pH near
Petition 870190096868, of 27/09/2019, p. 10/32
23/50 a preferential value. When metal ions are present in high concentrations, it may be necessary to use a chelating agent in the liquid medium.
[0089] The method and equipment for the cultivation of microorganisms and the production of microbial by-products can be carried out in batches, almost continuous or continuous processes.
[0090] Microbes can be grown in planktonic form or as biofilm. In the case of biofilm, the pot may have a substrate inside which the microbes can be grown in the biofilm state. The system may also have, for example, the ability to apply stimuli (such as shear stress) that encourage and / or improve biofilm growth characteristics.
[0091] In one embodiment, the method for cultivating microorganisms is carried out at about 5 Ό to about 1 00 Ό, preferably 15 Ό to 60 Q, more preferably, 25 Ό to 50 Ό. In another embodiment, cultivation can be carried out continuously at a constant temperature. In another modality, cultivation may be subject to temperature changes.
[0092] In one embodiment, the equipment used in the cultivation method and process is sterile. The cultivation equipment, such as the reactor / vessel, can be separated, but connected to a sterilization unit, for example, an autoclave. The cultivation equipment may also have a sterilization unit that sterilizes on site before starting inoculation. The air can be sterilized by methods known in the art. For example, ambient air can pass through at least one filter before being introduced into the vessel. In other embodiments, the medium can be pasteurized or, optionally, no added heat, in which the use of low water activity and low pH can be exploited to control bacterial growth.
[0093] The biomass content of the fermentation broth can be, for
Petition 870190096868, of 27/09/2019, p. 10/33
24/50 example, from 5 / g / L to 180 / g / L or more. In one embodiment, the solids content of the broth is 10 / g / L to 150 / g / L.
[0094] In one embodiment, the present invention further provides a method for the production of microbial metabolites such as ethanol, lactic acid, beta-glucan, proteins, peptides, metabolic intermediates, polyunsaturated fatty acid, and lipids. The content of metabolites produced by the method can be, for example, at least 20%, 30%, 40%, 50%, 60%, 70%, 80% or 90%.
[0095] The by-product of microbial growth produced by microorganisms of interest can be retained in the microorganisms or secreted in the liquid medium. In another embodiment, the method for the production of microbial growth by-product may also comprise stages of concentration and purification of the microbial growth by-product of interest. In an additional embodiment, the liquid medium may contain compounds that stabilize the activity of the microbial growth by-product.
[0096] In one embodiment, the entire microbial culture composition is removed after completion of the culture (for example, after, for example, reaching a desired cell density, or density of a specified metabolite in the broth). In this batch procedure, a completely new batch is started after the first batch has been harvested.
[0097] In another mode, only a part of the fermentation product is removed at any time. In this modality, the biomass with viable cells remains in the pot as an inoculant for a new cultivation lot. The composition that is removed may be a cellless broth or may contain cells. In this way, an almost continuous system is created.
[0098] Advantageously, the method does not require complicated equipment or high energy consumption. The microorganisms of interest can be grown on a small or large scale on the site and use
Petition 870190096868, of 27/09/2019, p. 10/34
25/50, even though they are still mixed with their means. Likewise, microbial metabolites can also be produced in large quantities at the required site.
[0099] Advantageously, microbial-based products can be produced at remote sites. Microbe growth facilities can operate outside the grid using, for example, solar, wind and / or hydroelectric power.
[00100] The microorganisms useful according to the systems and methods of the present invention can be, for example, bacteria, yeasts and / or fungi. These microorganisms can be natural or genetically modified microorganisms. For example, microorganisms can be transformed with specific genes to display specific characteristics. Microorganisms can also be mutants of a desired strain. As used here, mutant means a strain, genetic variant or subtype of a reference microorganism, in which the mutant has one or more genetic variations (for example, a point mutation, missense mutation, non-sense mutation, deletion, duplication, mutation of frame shift, or repetition expansion) compared to the reference microorganism. The procedures for producing mutants are well known in the microbiological art. For example, UV mutagenesis and nitrosoguanidine are widely used for this purpose.
[00101] In one embodiment, microorganisms are bacteria, including Gram-positive and Gram-negative bacteria. Bacteria can be bacteria forming endospores or exospores. The bacteria can be, for example, Agrobacterium radiobacter, Alcanivora borkumensis, Azobacter (A. vinelandii, A. chroococcum), Azospirillum brasil iens is, Bacillus (for example, B. subtil is, B. licheniformis, B. firmus, B. laterosporus, B. megaterium, B. amyloliquifaciens), Clostridium
Petition 870190096868, of 27/09/2019, p. 10/35
26/50 (C. butyricum, C. tyrobutyricum, C. acetobutyricum, Clostridium NIPER 7 and C. beijerinckii), LactoBacillus fermentum, Norcardia sp., Pseudomonas (P. chlororaphis subsp. Aureofaciens (Kluyverum), aeruginosa), Rhizobium rubrum. Sphingomonas paucimobilis, Ralston ia eulropha, Serratia marcescens and / or Tsukamurella sp.
[00102] In preferred modalities, the microorganism is a strain of Bacillus selected from the species B. subtilis, B. amyloliquefaciens and B. licheniformis. Even more preferably, the Bacillus strain is in the form of spores.
[00103] In certain embodiments, the present invention uses strains of Bacillus subtilis with enhanced production of biotensoactive in comparison with naturally occurring Bacillus subtilis, as well as in comparison with other microbes used in oil recovery. In certain modalities, strains of Bacillus subtilis increased the production of biopolymers, solvents and / or enzymes. Such Bacillus subtilis have been called members of the B series, including, among others, B1, B2 and B3.
[00104] In one embodiment, the microorganism is B. subtilis var. B1 or B2 locuses, which are effective producers, for example, of surfactin and other biotensives, as well as biopolymers. This specification incorporates by reference International Publication No. WO 2017/044953 A1 insofar as it is consistent with the teachings described herein.
[00105] A culture of the microbe B. subtilis B1 was deposited at the American Type Culture Collection (ATCC), 10801 University Blvd., Manassas, Va. 20110-2209 USA. The deposit received accession number ATCC No. PTA-123459 by the depositary and was deposited on August 30, 2016.
[00106] The vegetative cells of the B1 strain of Bacillus subtilis are stems from 0.7 to 0.9 pm wide by 1.6 to 3.3 pm long and
Petition 870190096868, of 27/09/2019, p. 10/36
27/50 occur in isolation. It is mobile, Gram-positive and produces biopolymers in nutrient agar and potato dextrose agar. It also produces ellipsoidal spores centrally or paracentrally in non-swollen sporangia. The size of mature spores is 0.8 to 1.0 pm wide by 1.6 to 1.9 pm long. The agar colonies are cream / beige, elevated, mucous, circular, whole, smooth, shiny and 3.0 to 7.0 mm in diameter after 16 hours at 40 Ό on the nutrient agar plate. It is an optional aerobic with a growth temperature range of 25 a to 55 Ό, with an ideal growth temperature of 35 Ό. It hydrolyzes starch, is positive in the Voges-Proskauer test, can use citrate and can grow with 15% NaCI.
[00107] In certain modalities, strains of Bacillus subtilis are tolerant to salt. Tolerance to salt can be in relation to any one or more of a variety of salts. For example, the salt can be a monovalent salt, such as a sodium or potassium salt, for example, NaCI or KCI, or a divalent salt such as a magnesium or calcium salt, for example, MgCh or CaCh, or a salt trivalent. Given the geographical sites to be treated, zinc, bromine, iron or lithium salts are present in the composition or on the site. In preferred embodiments, the bacteria described here are tolerant to NaCI, as well as other of the salts mentioned above, and are therefore widely useful for oil recovery.
[00108] In some modalities, strains of Bacillus subtilis are able to thrive under conditions of low oxygen concentration. In some modalities, the Bacillus subtilis strain is grown under microaerophilic or anaerobic conditions. Under microaerophilic and / or anaerobic conditions, nitrate salts can be added to replace oxygen as an electron acceptor to support anaerobic respiration.
[00109] Bacillus subtilis B series of strains produces more bioten
Petition 870190096868, of 27/09/2019, p. 37/107
28/50 soactive compared to reference strains of Bacillus subtilis. In addition, strains of Bacillus subtilis survive better under conditions of high salt concentration and anaerobic conditions than other known strains. Strains are also able to grow in anaerobic conditions. Strains in the Bacillus subtilis B series can also be used to produce enzymes that degrade or metabolize oil or other petroleum products.
[00110] In one embodiment, the methods in question may use yeast or fungus fermentation products. Yeast and fungi species suitable for use according to the present invention include, for example, Candida, Saccharomyces (S. cerevisiae, S. boulardii sequela, S. torula), Issatchenkia, Kluyveromyces, Pich ia, Wickerhamomyces (for example, l / IZ. anomalus), Starmerella (for example, S. bombicola), Rhodotorula (for example, R. glutinous and R. graminus), Mycorrhiza, Mortierella, Phycomyces, Blakeslea, Thraustochytrium, Phythium, Entomophthora, Aureobasidium pullulans, Pseudozy , Aspergillus and / or Rhizopus spp.
[00111] In one embodiment, yeast is a killer yeast. As used herein, killer yeast means a yeast strain characterized by its secretion of toxic proteins or glycoproteins, to which the strain itself is immune. Exotoxins secreted by killer yeasts are capable of killing other strains of yeasts, fungi or bacteria. For example, microorganisms that can be controlled by killer yeasts include Fusarium and other filamentous fungi. Such yeasts may include, but are not limited to, Wickerhamomyces (for example, W. anomalus), Pichia (for example, P. anomalous, P. guielliermondii, P. occidentalis, P. kudriavzevii), Hanse nu Ia, Saccharomyces, Hanseniaspora (e.g., H. uvarum), Ustilago (e.g., U. maydis), Debaryomyces hansenii, Candida, Cryptococcus, Kluyveromyces, Torulopsis, Williopsis, ZygoPetition 870190096868, from 27/09/2019, p. 38/107
29/50 saccharomyces (for example, Z. bailii) and others.
[00112] In one embodiment, the yeast fermentation product can be obtained via cultivation of a biochemical-producing yeast, such as, for example, anomalous Pichia (Wickerhamomyces anomalus). Wickerhamomyces anomalus is often associated with the production of food and grains and is an effective producer of various solvents, enzymes, toxins, as well as biotensoactive glycolipids, such as SLP. The fermentation broth after 7 days of cultivation at 25 O to 30 O may contain the yeast cell suspension and, for example, 4 / g / L or more of glycolipid biotensoactives.
[00113] In one embodiment, the yeast fermentation product can also be obtained through the cultivation of the yeast producing a biotensoactive, Starmerella bombicola. This species is an effective producer of glycolipid biotensoatives, such as SLP. The fermentation broth after 5 days of cultivation at 25 ° C may contain the yeast cell suspension and, for example, 150 / g / L or more of glycolipid biotensoactive.
[00114] In one embodiment, the yeast fermentation product may comprise the fermentation broth, separated from the yeast cells. In one embodiment, biotensives or other growth by-products in the broth are further separated from the broth and purified.
[00115] Other microbial strains, including, for example, other fungal strains capable of accumulating significant amounts, for example, of glycolipid biotopes or lipopeptides or other metabolites, can be used in accordance with the present invention. Other useful metabolites according to the present invention include manoprotein, beta-glucan and others that have bioemulsifying properties and surface / interfacial tension reduction properties. Preparation of Microbial Products
Petition 870190096868, of 27/09/2019, p. 10/39
[00116] A microbial product of the present invention is simply the fermentation broth that contains the microorganism and / or microbial metabolites produced by the microorganism and / or any residual nutrients. The fermentation product can be used directly without extraction or purification. If desired, extraction and purification can be easily achieved using standard extraction methods or techniques known to those skilled in the art.
[00117] The microorganisms in the product based on microbes can be in an active or inactive form. Microbe-based products can be used without further stabilization, preservation and storage. Advantageously, the direct use of these microbial-based products preserves a high viability of microorganisms, reduces the possibility of contamination from foreign agents and undesirable microorganisms, and maintains the activity of microbial growth by-products.
[00118] The microbes and / or broth resulting from microbial growth can be removed from the growth vessel and transferred, for example, via tubing for immediate use.
[00119] Advantageously, according to the present invention, the microbial product can comprise broth in which the microbes were grown. The product can be, for example, at least by weight, 1%, 5%, 10%, 25%, 50%, 75% or 100% of broth. The amount of biomass in the product, by weight, can be, for example, between 0% and 100%, including all percentages between them.
[00120] In other modalities, the composition (microbes, broth, or microbes and broth) can be placed in containers of appropriate size, taking into account, for example, the intended use, the application method considered, the size of the fermentation, and any mode of transport from the growing facility
Petition 870190096868, of 27/09/2019, p. 10/40
31/50 microbial growth to the site of use. Thus, the containers in which the microbial-based composition is placed can be, for example, from 1 gallon to 1,000 gallons or more. In certain embodiments, the containers are 2 gallons, 5 gallons, 25 gallons, or larger.
[00121] After harvesting the microbial-based composition from the growth vessels, other components can be added as the harvested product is placed in containers and / or piped (or transported for use). Additives can be, for example, buffers, vehicles, other microbial compositions produced in the same or another facility, viscosity modifiers, preservatives, nutrients for microbial growth, tracking agents, pesticides and other specific ingredients for use intended.
[00122] Up to, for example, 50% by weight or more of additives can be added, as needed, for specific applications, such as, for example, to vary VOC levels, increase the penetration of the mixture, decrease the viscosity of the mixture, as couplers for insoluble in the mixture, and to provide solvents. All additives must have a flash point greater than 37.7 Ό (100 ° F), preferably greater than 655 Ό (150 ° F) and more preferably 90.5 CC TCC (195 ° F) to achieve a final product flash point greater than 93.3 Ό (200 ° F).
[00123] Optionally, the product can be stored before use. The storage time is preferably short. Thus, the storage time can be less than 60 days, 45 days, 30 days, 20 days, 15 days, 10 days, 7 days, 5 days, 3 days, 2 days, 1 day or 12 hours. In a preferred embodiment, if living cells are present in the product, the product is stored at a low temperature such as, for example, less than 20 Ό, 15 Ό, 10 Ό or 5 Ό. On the other hand, a bio-surfactant composition can typically be filed under Application 870190096868, of 27/09/2019, pg. 41/107
32/50 stored at room temperature.
Local Production of Microbe-Based Products [00124] In preferred embodiments of the present invention, a microbial growth plant produces fresh high-density microorganisms and / or microbial growth by-products of interest on a desired scale. The microbial growth facility can be located at or near the application site. The facility produces compositions based on high density microbes in batch, quasi-continuous or continuous cultivation.
[00125] The present invention uses cultivation processes ranging from small (for example, laboratory environment) to large (for example, industrial environment) scales. Such cultivation processes include, but are not limited to, submerged cultivation / fermentation, solid state fermentation (SSF), and combination thereof.
[00126] The microbial growth facilities of the present invention produce fresh microbial compositions, comprising the microbes themselves, microbial metabolites and / or other components of the broth in which the microbes are grown. If desired, the compositions can have a high density of vegetative cells or a mixture of vegetative cells, spores, mycelia, conidia or other microbial propagules.
[00127] Advantageously, the microbial products in question can be adapted for use in a specified location. In one embodiment, the microbial growth facility is located at, or close to, a site where microbial products will be used. For example, the microbial growth facility may be less than 300, 250, 200, 150, 100, 75, 50, 25, 15, 10, 5, 3 or 1 mile from the location of use.
[00128] As the product based on microbes is generated locally, on the site or close to the application site, without resorting to the processes
Petition 870190096868, of 27/09/2019, p. 42/107
33/50 stabilization, preservation, storage and transport of microorganisms from conventional microbial production, a much higher density of living microorganisms can be generated. Therefore, a smaller volume of the microbial-based product is required for use in site application. In addition, this allows for higher density microbial applications, when necessary, to achieve the desired effectiveness.
[00129] Advantageously, this allows for a small-scale bioreactor (for example, smaller fermentation tank and smaller supplies of starting material, nutrients, pH control agents and anti-foaming agents, etc.), which makes the system efficient and facilitates product portability. The local generation of the product based on microbes also facilitates the inclusion of the growth broth in the product, thus eliminating the need to stabilize the cells or separate them from their culture broth. The broth may contain agents produced during fermentation that are particularly suitable for local use [00130] Robust, high-density microbial cultures produced locally are more effective in the field than those that have undergone cell stabilization or are in the supply chain some time ago. The microbial products of the present invention are particularly advantageous compared to traditional products in which cells, spores, mycelia, conidia or other microbial propagules have been separated from the metabolites and nutrients present in the fermentation growth medium. The reduced transport times allow the production and delivery of fresh batches of microbes and / or their metabolites in the time and volume required by local demand.
[00131] Advantageously, these microbial growth facilities provide a solution to the current problem of having
Petition 870190096868, of 27/09/2019, p. 43/107
34/50 large industrial producers whose product quality suffers from delays in upstream processing, bottlenecks in the supply chain, inadequate storage, and other contingencies that inhibit the timely delivery and application, for example, of a viable high-end product cell count and the associated broth and metabolites in which the cells were originally grown.
[00132] Microbial growth facilities offer manufacturing versatility due to the ability to adapt products based on microbes to enhance synergies with destination geographies. Advantageously, in preferred modalities, the systems of the present invention take advantage of the power of naturally occurring local microorganisms and their metabolic by-products to improve oil production. Local microbes can be identified based on, for example, salt tolerance or the ability to grow at high temperatures.
[00133] The cultivation time for individual pots can be, for example, from 1 to 7 days or more. The crop product can be harvested in several different ways.
[00134] Local production and delivery within, for example, 24 hours of fermentation results in pure high cell density compositions and substantially lower shipping costs. Given the prospects for rapid advances in the development of more effective and powerful microbial inoculants, consumers will benefit greatly from this ability to rapidly supply microbial products.
Enhanced Oil Recovery Methods [00135] In some embodiments, the present invention provides materials and methods for improving oil production, treating an oil production site, for example, a formation containing oil or an oil well, with micro -organisms and / or their
Petition 870190096868, of 27/09/2019, p. 44/107
35/50 growth by-products. In one embodiment, the present invention may be useful for improving oil recovery from an oil well, by stimulating, for example, the flow of oil from the well while dissolving the scale inside the formation. [00136] As used here, applying a composition or product refers to putting it in contact with a target or site so that the composition or product can have an effect on that target or site. The effect may be due, for example, to microbial growth and / or the action of a biotensoactive or other growth by-product. For example, microbial-based compositions or products can be injected into oil wells and / or into the pipe, liner, annulus, pumps, tanks, etc. associated with oil production sites and oil-containing formations.
[00137] In one embodiment, the present invention provides a method for enhancing oil recovery by applying one or more microorganisms capable of producing useful biochemical by-products in an oil production site, for example, a formation containing oil and / or oil well. The method optionally includes adding nutrients and / or other agents to the site. In preferential modalities, the microorganism is a species of bacteria producing biotensoactive.
[00138] In certain embodiments, microorganisms are selected from strains of Bacillus, including, but not limited to strains of Bacillus subtilis, Bacillus licheniformis and Bacillus amyloliquefaciens. In preferred embodiments, the bacteria are in the form of spores.
[00139] In one embodiment, the method also includes adding nutrients and / or germination enhancers to promote germination and microbial growth. For example, nutrients such as sources of carbon, nitrogen, magnesium, phosphorus and protein can
Petition 870190096868, of 27/09/2019, p. 45/107
36/50 be added. Germination enhancers such as L-alanine and manganese can also be added.
[00140] The method may also comprise the addition of a yeast fermentation product, such as the fermentation broth resulting from the cultivation, for example, of Starmerella bombicola or Wickerhamomyces anomalus. In one embodiment, yeast is a yeast that produces biotensoactive. In one embodiment, the fermentation broth comprises the by-products of yeast growth, such as, for example, biotensoactive glycolipids and other metabolites.
[00141] In one embodiment, yeast cells can be removed from the yeast fermentation product and only the broth containing biotensoactives and other metabolites is applied. In one embodiment, the yeast fermentation product comprises biotensoatives that have been separated from the fermentation broth and purified.
[00142] In certain embodiments, the yeast fermentation products of the present invention have advantages over, for example, only biotensives, including one or more of the following: high concentrations of manoprotein as a part of the outer surface of a yeast cell wall; the presence of beta-glucan in the yeast cell walls; and the presence of biotensoactives and other metabolites (for example, lactic acid, ethanol, ethyl acetate, etc.) in the culture.
[00143] The method can also comprise the application of microbes and / or by-products of microbial growth with one or more alkaline compounds. The alkaline compound can be, for example, ammonium hydroxide.
[00144] In some embodiments, the method may also comprise the application of microbes and / or by-products of microbial growth with one or more polymeric compounds. The compounds
Petition 870190096868, of 27/09/2019, p. 46/107
37/50 polymers can be selected from biopolymers, such as, for example, hydrogels, polysaccharides, xanthan gum, guar gum and cellulose polymers.
[00145] In some modalities, the method may also include the application of microbes and / or by-products of microbial growth with one or more non-biological surfactants. Surfactants can be, for example, anionic, cationic, non-ionic or zwitterionic.
[00146] In one embodiment, microbes and / or by-products of microbial growth can be applied with one or more chelating agents to reduce, for example, dissolve, the scale that has accumulated in the formation containing oil. Chelating agents can be, for example, citric acid, EDTA and / or sodium citrate.
[00147] In some modalities, the microbial products and methods in question can still be used for removing paraffin, liquefying solid asphaltene, and bioremediation of waters, soils and other sites contaminated with hydrocarbons.
[00148] In one embodiment, products based on microbes are applied to a working well, including the surrounding formation. In this mode, the product can be poured or injected on the side of the liner (posterior lines) of a well and allowing it to mix with the fluid that is already in the well. When sufficient fluid is present, the composition can then optionally be circulated by, for example, a pump for 24 to 72 hours, preferably 48 to 72 hours. Before circulation, the composition can be left to stand for 8 to 24 hours, for example. The curing time, circulation time and dosage depend on the depth and size of the well. A basic initial dosage can be, but is not limited to, 20 gallons of composition, and at least about 5 gallons of composition per well, periodically, for example, fortnightly, menPetição 870190096868, from 27/09/2019, pg. 47/107
38/50 sally, bimonthly.
[00149] In one embodiment, microorganisms can germinate and grow on the spot and produce biotensoactives on the oil production site. Consequently, a high concentration of biotensoactive and micro-organisms producing biotensoactives in a treatment site (for example, an oil well) can be achieved easily and continuously.
[00150] In one embodiment, it is desirable to introduce the composition, through perforations in the coating, in the formation containing surrounding oil. The composition can be forced into the surrounding formation by applied pressure or, if the composition is allowed to settle to the bottom of the coating, the composition can infiltrate the formation without additional pressure. The composition permeates the formation, dissolving blocks in the formation to provide a more efficient recovery of oil and gas.
[00151] In additional embodiments, the composition of the present invention can be applied directly to the equipment. For example, before placing rods and linings in gas and / or oil wells, these parts can be sprayed with or embedded in the composition. The parts can also be dipped in tanks filled with the composition.
[00152] The composition can be introduced by means of injection pumps in oil wells or offshore gas to improve oil recovery. To treat the lines, from 1-500 gallons to 1000 barrels, 10,000 barrels or more, for example, the composition can be applied to the composition at an injection rate, for example, 1 to 20 gallons per minute, or 1 to 20 barrels per minute.
[00153] The treatment in question can be effective in a variety of different geological formations. For example, the present invention can be useful in formations as deep as about
Petition 870190096868, of 27/09/2019, p. 48/107
39/50
7,000 feet or more deep, and as shallow as about 1,500 feet or more shallow. In addition, the invention can be useful in formations with a range of porosity and / or permeability, for example, from about 0.1% to about 20% or more. The invention can also be useful in formations having a wide range of temperatures, pH and salinity.
[00154] In one embodiment, enhanced oil recovery is achieved through selective clogging, in which the fluid flow through the reservoir is moved from the high permeability zones of the reservoir to zones of moderate or low permeability. The sweeping efficiency can be increased, for example, by forcing the injected water to pass through oil areas previously bypassed the reservoir. Changes in the flow pattern can be achieved by an increase in microbial cell mass within the reservoir, for example, injecting microorganisms together with nutrients. The injected nutrient and microbes preferentially flow to the high permeability zones of the reservoir and, as a result of cell growth, biomass selectively seals these zones to a greater extent than the moderate or low permeability zones. In one embodiment, microbes are injected as spores and germinate while they are inside the reservoir.
Enhanced Oil Recovery Using the Alkaline-Surfactant-Polymer (ASP) Method [00155] In one embodiment, methods are provided to enhance oil recovery, in which a microbial product of the present invention is applied to an oil production site in combination with one or more alkaline compounds, polymers, surfactants, or combinations thereof.
[00156] In floods of surfactants, reducing the interfacial tension between oil and displaced water and also the interfacial tension
Petition 870190096868, of 27/09/2019, p. 49/107
40/50 between the oil and rock interfaces, the residual oil can be moved and recovered.
[00157] In the caustic flood, the reaction of the alkaline compounds with the organic acids in the oil forms natural surfactants in the place that decrease the oil-water interfacial tension.
[00158] In addition to the surfactant and alkaline floods, polymers are used to increase the viscosity of displacement water to improve the efficiency of oil sweep.
[00159] ASP flooding is a combination process in which alkalis, surfactants and polymers are injected. ASP involves the injection of a solution containing polymer, alkali and surfactant into an impoverished or matured oil field, with the goal of achieving optimal chemistry in large injection volumes at minimal cost. The alkali-surfactant mixture forms an emulsion with the oil, which is then swept and displaced from the reservoir using a polymer unit. The ASP flooding improves the efficiency of microscopic displacement, reducing the interfacial tension (IFT) between water and oil by adding a surfactant to water, while combining the mobility of oil and water through the addition of polymer. Alkali is also added to the water to reduce the adsorption of the surfactant on the pore walls and to control local salinity to ensure minimum IFT and change the wettability of the rock. Use of Microbes with Surfactants in Oil Recovery [00160] In certain embodiments, the oil recovery methods described here use one or more microbes and / or microbial growth by-products (for example, biotensives), combined with other compositions. In one embodiment, the other compositions are non-biological surfactants.
[00161] A surfactant molecule (Active Surface Agent) has two functional groups, that is, a hydrophilic group (soluble in
Petition 870190096868, of 27/09/2019, p. 50/107
41/50 water) or polar and a hydrophobic (oil-soluble) or non-polar group. The hydrophobic group is usually a long chain of hydrocarbons (C8-C18), which may or may not be branched, while the hydrophilic group is formed by portions such as carboxylates, sulfates, sulfonates (anionic), alcohols, polyoxyethylene (non-ionic) chains and quaternary ammonium salts (cationic).
[00162] Surfactants work in ASP floods to decrease the interfacial tension (IFT) between oil and brine, to help mobilize and contribute to the formation of oil banks. The IFT reduction reduces capillary forces and allows the oil bank to flow more freely without re-locking. The selection of a suitable surfactant for EOR purposes is based on the ability to reduce IFT between brine and brine, thermal stability, tolerance to salinity and hardness of brine, brine solubility, phase behavior parameters, adsorption test under conditions static and dynamic and displacement studies under reservoir conditions.
[00163] Surfactants according to the described methods include, among others: anionic surfactants, ammonium lauryl sulfate, sodium lauryl sulfate (also called SDS, sodium dodecyl sulfate), alkyl ether sulfates, sodium laureth sulfate (also known such as sodium lauryl ether sulfate (SLES)), myreth sodium sulfate; docusates, sodium dioctyl sulfosuccinate, perfluorooctane sulfonate (PFOS), perfluorobutanesulfonate, linear alkyl benzene sulfonates (LABs), alkyl aryl ether phosphates, alkyl ether phosphate; carboxylates, alkyl carboxylates (soaps), sodium stearate, sodium lauroyl sarcosinate, carboxylate-based fluorostensives, perfluorononanoate, perfluorooctanoate; cationic surfactants, pH-dependent primary, secondary or tertiary amines, octenidine dihydrochloride, permanently charged quaternary ammonium cations, alkyl trimethyl salts
Petition 870190096868, of 27/09/2019, p. 51/107
42/50 mononium, cetyl trimethylammonium bromide (CTAB) (also known as hexadecyl trimethyl ammonium bromide), cetyl trimethylammonium chloride (CTAC), cetyl pyridinium chloride (CPC), benzalkonium chloride (BAC), benzethonium chloride (BZT ), 5-bromo-5-nitro-1,3-dioxane, dimethyldioctadecylammonium chloride, cetrimony bromide, dioctadecyldiomethylammonium bromide (DODAB); zwitterionic (amphoteric) surfactants, CHAPS sultaines (3 - [(3-Colamidopropyl) dimethylammonium] -1propanesulfonate), cocamidopropyl hydroxisultaine, betaines, cocamidopropyl betaine, phosphatidyl serine, phosphatidylethanolamine, phosphatidylethylamine, long-term, phosphatidylethylate cetyl alcohol, stearyl alcohol, keto stearyl alcohol, oleyl alcohol, polyoxyethylene glycol alkyl ethers (Brij): CH3- (CH2) 10-16- (O-C2H4) 1-25-OH (octaethylene glycol monododecyl ether, pentaethylene glycol monododecyl ether) , polyoxypropylene glycol alkyl ethers: CH3- (CH2) 10-16- (O-C3H6) 1-25OH, glycoside alkyl ethers: CH3- (CH2) 10-16- (O-glycoside) 1-3OH (decyl glycoside, lauryl glycoside, octyl glycoside), polyoxyethylene glycol octylphenol ethers: C8H17- (C6H4) - (O-C2H4) 1-25-OH (Triton X100), polyoxyethylene glycol alkylphenol ethers: C9H19- (C6H4) - (OC2H4) 1-25-OH (nonoxynol-9), glycerol alkyl esters (glyceryl laurate), polyoxyethylene glycol sorbitan alkyl esters (polysorbate) , sorbitan alkyl esters (spans), cocamide MEA, cocamide DEA, dodecyldimethylamine oxide, copolymers of polyethylene glycol and polypropylene glycol (poloxamers) and polyethoxylated tallow amine (POEA).
[00164] Anionic surfactants contain anionic functional groups in your head, such as sulfate, sulfonate, phosphate and carboxylates. Prominent alkyl sulfates include ammonium lauryl sulfate, sodium lauryl sulfate (also called SDS, sodium dodecyl sulfate) and alkyl ether sulfates related to sodium laureth sulfate, also known as sodium lauryl ether sulfate (SLES) and myreth sulfate sodium. Carboxylates are the most common surfactants and comprise
Petition 870190096868, of 27/09/2019, p. 52/107
43/50 include alkyl carboxylates (soaps), such as sodium stearate.
[00165] Surfactants with cationic head groups include: pH-dependent primary, secondary or tertiary amines; octenidine dihydrochloride; permanently charged quaternary ammonium cations, such as alkyl trimethylammonium salts: cetyl trimethylammonium bromide (CTAB) also known as hexadecyl trimethyl ammonium bromide, cetyl trimethylammonium chloride (CTAC); cetyl pyridinium chloride (CPC); benzalkonium chloride (BAC); benzethonium chloride (BZT); 5-bromo-5-nitro-1,3-dioxane; dimethyldioctadecylammonium chloride; cetrimony bromide; and dioctadecyldi-methylammonium bromide (DODAB).
[00166] Zwitterionic (amphoteric) surfactants have cationic and anionic centers linked to the same molecule. The cationic part is based on primary, secondary or tertiary amines or quaternary ammonium cations. The anionic part can be more variable and can include sulfonates. The most common biological zwitterionic surfactants have a phosphate anion with an amine or ammonium, such as phosphatidylserine phospholipids, phosphatidylethanolamine, phosphatidylcholine and sphingomyelin.
[00167] A surfactant with an uncharged hydrophilic part, for example, ethoxylate, is non-ionic. Many long-chain alcohols exhibit some properties of surfactants.
Use of Microbes with Polymers in Oil Recovery [00168] The present invention provides methods of enhanced oil recovery using one or more microbes and / or microbial growth by-products, combined with one or more polymeric compounds. The polymeric compounds used to recover oil in combination with the microbes of the present invention include, but are not limited to: hydrogels, acrylic acid, acrylamide, polyacrylamide (PAM), hydrolyzed polyacrylamide (HPAM), polysaccharí
Petition 870190096868, of 27/09/2019, p. 53/107
44/50 deo, xanthan gum, guar gum, and cellulose polymer. In preferred embodiments, the polymer is a biopolymer selected from, for example, hydrogels, xanthan gum, guar gum, cellulose polymers, polysaccharides, and others.
[00169] The associative water-soluble polymer is a relatively new class of polymers that has recently been introduced for applications in oil fields. These polymers consist of a long chain hydrophilic skeleton, with a small number of hydrophobic groups located either randomly along the chain or at the ends of the chain. When these polymers are dissolved in water, hydrophobic groups are added to minimize their exposure to water. The incorporated groups are associated due to intramolecular hydrophobic interactions and intermolecular hydrophobic interactions. The functional groups of this polymer are less sensitive to brine salinity compared to polyacrylamide, whose viscosity decreases dramatically with increasing salinity.
[00170] Polymer flooding may involve adding polymer to the water from a water flood to decrease its mobility. The polymers increase the viscosity of the aqueous phase as well as reduce the permeability to water due to mechanical entrapment, consequently resulting in a more favorable rate of mobility. With a more viscous phase, the collected oil bank can be moved more easily through the reservoir and eventually to the production well.
[00171] Polymers according to these modalities can also be removed and / or degraded using the microbial-based composition of the present invention, once their function in the well is no longer needed.
[00172] In one embodiment, the present invention provides a method
Petition 870190096868, of 27/09/2019, p. 54/107
45/50 all to improve the recovery of hydrocarbons from a fracturing well by applying a microbial-based composition to one drilling site that comprises one or more strains of microorganisms. In certain embodiments, polymers have accumulated within the well after they have performed their desired function within the well.
[00173] The microbes of the microbial-based composition and / or their growth by-products can rapidly digest polymers such as polylactic acid (PLA); thus, the method improves the ability to recover hydrocarbon resources by reducing the accumulation of PLA and other resins within fractures and in the wells of fracturing wells, once their usefulness has been exhausted. The method optionally includes the addition of nutrients and / or other agents to the site in order to promote microbial growth. The method may also include the addition of polymer degradation enzymes to the site in order to improve polymer degradation.
[00174] In one embodiment, the present invention provides methods of recovering polymeric substances that remain in the wells, including fracturing wells, after their usefulness has been exhausted. For example, biotensives produced by the methods and microorganisms of the present invention can reduce the interfacial tension of fluids used to elevate polymeric substances, such as PAM gel friction reducers. In another modality, biotensoatives can be used to cleave the PAM gel prior to elevation. Use of Microbes with Alkaline Compounds in Oil Recovery [00175] The present invention provides improved oil recovery methods using one or more microbes and / or microbial growth by-products, combined with one or more alkaline compounds. Alkaline compounds used to recover petroleum
Petition 870190096868, of 27/09/2019, p. 55/107
46/50 oil in combination with the microbes of the present invention include, but are not limited to, ammonium hydroxide.
[00176] Alkali is a basic ionic salt of an element of alkali metal or alkaline earth metal. The use of alkalis in a chemical flood offers several benefits, including the promotion of crude oil emulsification, the increase in the ionic strength of the aqueous phase leading to the regulation of the behavior of the injected surfactant phase, and the reduction of IFT to very low values in the presence of surfactant. [00177] Alkali can also reduce costs by limiting the amount of surfactant needed in two ways. First, the alkali reduces the adsorption of the surfactant, increasing the density of the negative charge on the rock surface, making it preferably moist. Second, alkali reacts with acids in crude oils to produce soaps on the spot, which in turn expands the ideal range of salinity. The soap generated creates a microemulsion phase that can coexist with oil and water, extending the three-phase region (or ultra low IFT region).
[00178] The selection of alkalis is guided by the type of formation, type of clay and divalent cations. Common alkaline agents include sodium hydroxide (NaOH or lye), sodium carbonate (NazCCh or sodium carbonate), sodium bicarbonate (NaHCOs) and sodium metaborate (NaBOz). Sodium hydroxide solutions have been reported to interact strongly with sandstone at an elevated temperature (85 Ό (185 ° F)), resulting in weight loss and increased sandstone porosity. The caustic consumption resulting from the dissolution of NaOH from silicate minerals can be a significant and harmful factor during application in the field. Anionic surfactants showed much less adsorption in the presence of NazCOs compared to NaOH. Hydroxide is not a potential determining ion for carbonate surfaces. Calcium and other divalent cations can precipitate
Petition 870190096868, of 27/09/2019, p. 56/107
47/50 alkali, such as NazCCb, unless soft brine is used. This is a limitation of NazCCb. The use of NaBÜ2 as a substitute for NazCCh has been reported. This alkali provided pH values of about 11 to 1% by weight of alkaline concentration and generated soap for acidic crude oils. Another great advantage of NaBOz (sodium metaborate) species is their tolerance to divalent cations. In carbonate reservoirs, sodium metaborate is used in place of other alkalis. If the reservoir contains clays, NaHCOs are preferred. NazCOs are the most commonly used alkali because they are inexpensive and transport better in porous media.
[00179] The preferred oil formations for alkaline floods are sandstone reservoirs instead of carbonate formations that contain anhydrite (calcium sulfate) (CaSO4) or plaster (dehydrated calcium sulfate) (CaSO4.2H2O), which can consume large quantities amounts of alkaline chemicals. In addition, in carbonate reservoirs, precipitation of calcium carbonate (CaCOs) or calcium hydroxide (Ca (OH) 2) occurs when Na2COs or NaOH is added. Carbonate reservoirs also contain brine with a higher concentration of divalents and can cause precipitation. To overcome this problem, the suggested NaHCOs and sodium sulfate (Na2SO4) are used. NaHCOs have a much lower concentration of carbonate ions, and additional sulfate ions can decrease the concentration of calcium ions in the solution. These chemicals are also consumed by clays, minerals or silica, and the higher the temperature of the reservoir, the greater the consumption of alkalis. Another common problem during caustic flooding is the formation of scale in the production wells. During alkaline flooding, the injection sequence usually includes: (1) a prewash to condition the reservoir prior to the injection of the primary sludge, (2) primary sludge (alkaline chemicals), (3) polymer as a mobility buffer
Petition 870190096868, of 27/09/2019, p. 57/107
48/50 to move the primary mud. The alkaline flood can be modified as the processes AP (alkali - polymer), AS (alkali - surfactant) and alkali - surfactant - polymer (ASP). The soap produced from the reaction between the acidic components of a crude oil and the injected alkali is the main mechanism of oil recovery in alkaline floods.
Use of Microbes with Chelating Agents [00180] In some embodiments, microorganisms and / or their growth by-products can be applied with a chelator or chelating agent. Advantageously, the use of chelating agents helps to improve the recovery of oil by dissolving the incrustations within a formation. Fouling can block the pores and other flow paths of an oil-containing formation, slowing and / or blocking the flow of oil from the formation.
[00181] As used here, chelating or chelating agent means an active agent capable of removing a metal ion from a system forming a complex, so that the metal ion, for example, cannot readily participate or catalyze formation of oxygen radicals.
[00182] Examples of chelating agents suitable for the present invention include, but are not limited to, dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropanesulfonic acid (DMPS), alpha lipoic acid (ALA), thiamine-tetrahydrofurfuryl disulfide (TTFD), penicillamine, ethylene diaminetetraacetic acid (EDTA), sodium acetate, sodium citrate, and citric acid.
[00183] In preferred embodiments, the chelating agent is sodium citrate, citric acid, EDTA or a combination thereof.
EXAMPLES [00184] A greater understanding of the present invention and its many advantages can be obtained from the following examples,
Petition 870190096868, of 27/09/2019, p. 58/107
49/50 dos by way of illustration. The following examples are illustrative of some of the methods, applications, modalities and variants of the present invention. They are not to be considered as limiting the invention. Numerous changes and modifications can be made with respect to the invention.
EXAMPLE 1 - PRODUCTION OF BACILLUS SUBTILIS [00185] The fermentation of Bacillus subtilis var. locuses can be carried out in a 500 L reactor with 350 L of a nutrient medium containing (/ g / L):
Glucose 18 Powder molasses 2 Sucrose 1 KH2PO4 0.5 Na ₂ HPO ₄ -7H ₂ O 2.1 KCI 0.1 MgSO4 0.5 CaCI ₂ 0.05 Urea 2.5 NH4CI 1.24 Yeast extract 2 Corn Peptone 0.5 TekNova trace element (mL) 1
[00186] The cultivation temperature is 40 O, the pH stabilization is 6.8 to 7.0, and the DO stabilization is 20 to 30% (the oxygen concentration in the air is considered 100%). The duration of cultivation is 24 to 36 hours, or until at least 95% of the bacteria reach sporulation. The final concentration of the bacterial culture is not less than 1 χ 10 ⁹ CFU / ml. The amount of culture produced by a single fermentation cycle allows the production of more than 2,000 barrels of final treatment formulation containing 10 ⁶ CFU of this strain of Bacillus.
EXAMPLE 2 - FERMENTATION OF STARMERELLA BOMBICOLA FOR THE PRODUCTION OF BIOTENSOACTIVES
Petition 870190096868, of 27/09/2019, p. 59/107
50/50 [00187] The fermenter is an autoclavable stainless steel vessel designed specifically for the cultivation of yeasts and the production of biotensives. The fermenter is equipped with a microprocessor and an impeller, in addition to dissolved oxygen, pH, temperature and foam probes. It has an integrated control station with a colorful touch-sensitive interface, built-in pumps for an improved broth mix, gas flow controllers, and DO level / pH / foam controllers. The working volume of the 550-gallon reactor is 500 gallons.
[00188] The nutrient medium contains sources of carbon, protein, nitrogen and unsaturated oil or fatty acids. The one-day-old Starmerella bombicola culture (60-70L) is used to inoculate the reactor. The initial pH of the culture is 5.0-6.0 until microbial growth occurs and the pH begins to decrease. The duration of cultivation and collection of the finished product continues for 5 days at 25 Ό and the pH is stabilized at 3.5. The final content of soforolipid can reach at least 40% of the workload per cycle, or 150 / g / L or higher.

权利要求:
Claims (35)
[1]
1. Method to improve the amount of recoverable oil from an oil-containing formation, characterized by the fact that it comprises applying a biotensoactive-producing bacteria and a growth by-product to the formation, in which the bacteria is inactivated.
[2]
2. Method, according to claim 1, characterized by the fact that the bacterium is a strain of Bacillus in the form of inactivated spores.
[3]
3. Method, according to claim 2, characterized by the fact that the Bacillus strain is a strain of B. subtilis, B. amyloliquefaciens and / or B. licheniformis.
[4]
4. Method according to claim 2, characterized by the fact that the Bacillus strain is the B. subtilis B1, B2 or B3 strain.
[5]
5. Method according to claim 1, characterized by the fact that the growth by-product is a biotopeactive lipopeptide selected from surfactin and iturine A.
[6]
6. Method, according to claim 1, characterized by the fact that it also comprises administering one or more alkaline compounds to the formation.
[7]
7. Method according to claim 6, characterized by the fact that the alkaline compound is ammonium hydroxide.
[8]
8. Method, according to claim 1, characterized by the fact that it also comprises administering one or more polymeric compounds to the formation.
[9]
9. Method according to claim 8, characterized by the fact that one or more polymeric compounds are selected from xanthan gum, guar gum, hydrogels, polysaccharides, and cellulose polymers.
Petition 870190096868, of 27/09/2019, p. 67/107
2/4
[10]
10. Method, according to claim 1, characterized by the fact that it also comprises administering one or more non-biological co-surfactants to the formation.
[11]
11. Method according to claim 1, characterized by the fact that it further comprises applying a yeast fermentation product which comprises broth resulting from the fermentation of a yeast, in which the yeast fermentation product does not comprise yeast cells.
[12]
12. Method according to claim 11, characterized by the fact that the yeast is selected from Wickerhamomyces anomalus, Pichia guilliermondii and Starmerella bombicola.
[13]
13. Method according to claim 11, characterized in that the yeast fermentation product comprises a glycolipid biotensoactive.
[14]
14. Method, according to claim 13, characterized by the fact that the biotensoactive is a soforolipid.
[15]
15. Method according to claim 11, characterized in that the yeast fermentation product comprises a biotensoactive phospholipid.
[16]
16. Method according to claim 1, characterized by the fact that one or more chelating agents are applied to the formation.
[17]
17. Method according to claim 16, characterized in that it comprises using one or more of the following chelating agents: citric acid, sodium citrate, and EDTA.
[18]
18. Method, according to claim 1, characterized by the fact that it improves oil recovery and also dissolves scale deposits present in the formation.
[19]
19. Method according to claim 1, characterized by the fact that it further comprises applying a solvent to the formation.
Petition 870190096868, of 27/09/2019, p. 68/107
3/4
[20]
20. Method according to claim 19, characterized by the fact that the solvent is isopropyl alcohol or ethanol.
[21]
21. Method according to claim 11, characterized by the fact that the yeast fermentation product is produced at a location 300 miles or less from the formation.
[22]
22. Method, according to claim 1, characterized by the fact that it also comprises applying one or more purified biotensoatives to the formation.
[23]
23. Method according to claim 22, characterized by the fact that the one or more purified biotensoatives are glycolipids and / or phospholipids.
[24]
24. Composition, characterized by the fact that it comprises:
a yeast fermentation product comprising broth resulting from the fermentation of a yeast an alkaline compound; and a chelating agent.
[25]
25. Composition according to claim 24, characterized by the fact that it comprises a yeast selected from Wickerhamomyces anomalus, Pichia guilliermondii and Starmerella bombicola.
[26]
26. Composition according to claim 24, characterized in that the yeast fermentation product does not comprise yeast cells.
[27]
27. Composition according to claim 24, characterized by the fact that the alkaline compound is ammonium hydroxide.
[28]
28. Composition according to claim 24, characterized in that it comprises one or more of the following chelating agents: citric acid, sodium citrate, and EDTA.
[29]
29. Composition according to claim 24, character
Petition 870190096868, of 27/09/2019, p. 69/107
4/4 characterized by the fact that it still comprises a by-product of microbial growth.
[30]
30. Composition according to claim 29, characterized by the fact that the microbial growth by-product is a biotensoactive selected from glycolipids, lipopeptides and phospholipids.
[31]
31. Composition according to claim 24, characterized in that it further comprises a solvent.
[32]
32. Composition according to claim 31, characterized by the fact that the solvent is isopropyl alcohol or ethanol.
[33]
33. Composition, characterized by the fact that it comprises:
Water;
a surfactant, isopropyl alcohol, ammonium hydroxide, and a mixture of EDTA, sodium citrate and citric acid.
[34]
34. Composition according to claim 33, characterized by the fact that the surfactant is a glycolipid produced by a microorganism.
[35]
35. Method for improving the amount of oil recoverable from an oil-containing formation, characterized by the fact that it comprises applying a composition as defined in claim 24 to the formation.

类似技术:

---

公开号 | 公开日 | 专利标题

---

BR112019020271A2|2020-04-22|microbial products and their uses to enhance oil recovery

---

US20200179999A1|2020-06-11|Enhanced microbial production of biosurfactants and other products, and uses thereof

---

CN110291173B|2021-10-22|Microbial products and their use for bioremediation, removal of paraffins and other contaminants in oil and natural gas production processing equipment

---

US10947444B2|2021-03-16|Materials and methods for reducing viscosity of oil

---

CN108373912A|2018-08-07|A kind of salt-resisting high-temperature-resisting foam discharging agent and preparation method thereof

---

CN105441351B|2018-08-10|One plant of thermophilic production surfactant oil degradation bacteria and its application

---

US20200362228A1|2020-11-19|Multifunctional Composition for Enhanced Oil Recovery, Improved Oil Quality and Prevention of Corrosion

---

US11279965B2|2022-03-22|Symbiotic fermentation of Acinetobacter and Bacillus and applications thereof

---

US20200224238A1|2020-07-16|Symbiotic Fermentation of Acinetobacter and Bacillus and Applications Thereof

---

US20200291289A1|2020-09-17|Materials and Methods for Recovering Oil from Oil Sands

---

WO2019140440A1|2019-07-18|Reactors for modified solid-state fermentation

---

WO2018148265A2|2018-08-16|Materials and methods for reducing viscosity of oil

---

US10844276B2|2020-11-24|Composition and methods for microbial enhanced digestion of polymers in fracking wells

---

US20200157408A1|2020-05-21|Two-Step Process for Microbial Enhanced Oil Recovery

---

CA3125459A1|2020-07-09|Materials and methods for extended reduction of heavy crude oil viscosity

---

Mehta et al.2009|Microbial enhanced oil recovery

---

WO2020264071A1|2020-12-30|Multi-functional compositions comprising concentrated acids for enhanced oil and gas recovery

---

CN112877049A|2021-06-01|Microbial foam drainage agent and application thereof in gas well exploitation

同族专利:

---

公开号 | 公开日

---

CA3058761A1|2018-10-18|

---

CL2019002863A1|2020-04-03|

---

AU2018251675A2|2019-10-31|

---

EA201992274A1|2020-02-19|

---

EP3609975A4|2021-01-13|

---

CN110506095A|2019-11-26|

---

CO2019010273A2|2019-10-09|

---

US20200071600A1|2020-03-05|

---

MX2019012055A|2019-11-11|

---

WO2018191172A1|2018-10-18|

---

AU2018251675A1|2019-10-10|

---

EP3609975A1|2020-02-19|

引用文献:

---

公开号 | 申请日 | 公开日 | 申请人 | 专利标题

---

---

US4522261A|1983-04-05|1985-06-11|The Board Of Regents For The University Of Oklahoma|Biosurfactant and enhanced oil recovery|

---

WO2008086221A2|2007-01-04|2008-07-17|Advanced Biocatalytics Corporation|Enhanced oil recovery compositions comprising proteins and surfactants and methods of using the same|

---

CA2692658A1|2007-07-12|2009-01-15|Glori Oil Limited|Process for enhanced oil recovery using a microbial consortium|

---

WO2012116230A1|2011-02-25|2012-08-30|Geo Fossil Fuels, Llc|Alkaline microbial enhanced oil recovery|

---

AU2014318639B2|2013-09-12|2018-07-05|Geo Fossil Fuels, Llc|Microbial enhanced oil recovery method|

---

US10576519B2|2015-09-10|2020-03-03|Locus Oil Ip Company, Llc|Enhanced microbial production of biosurfactants and other products, and uses thereof|CN109897621A|2019-03-25|2019-06-18|大庆华理生物技术有限公司|A kind of binary biology oil displacement agent and its application|

---

CN111763240A|2019-04-02|2020-10-13|中国石油天然气股份有限公司|Sodium lauroyl sarcosinate-diglucoside peptide compound and composite oil displacement agent|

---

CA3144947A1|2019-06-26|2020-12-30|Locus Oil Ip Company, Llc|Multi-functional compositions comprising concentrated acids for enhanced oil and gas recovery|

---

CA3144950A1|2019-06-26|2020-12-30|Locus Oil Ip Company, Llc|Compositions and methods for paraffin liquefaction and enhanced oil recovery using concentrated acids|

---

IT201900012309A1|2019-07-18|2021-01-18|Soil Remediation G P T Sagl|COMPOSITION AND METHOD FOR THE REMOVAL AND RECOVERY OF HYDROCARBONS FROM A SOLID PHASE|

---

CN110566168A|2019-08-28|2019-12-13|北京大学|method for activating endogenous microorganism to enhance oil recovery or pollution treatment by injecting Dietzia|

---

WO2021064131A1|2019-10-03|2021-04-08|Clariant International Ltd|Biopolymers for enhanced hydrocarbon recovery|

---

CN110699408A|2019-10-21|2020-01-17|天津大学|Method for improving surfactant yield by mixed fermentation|

---

CN112708406A|2019-10-24|2021-04-27|中国石油化工股份有限公司|Biochemical composite blocking remover and preparation method and application thereof|

---

CN112745855A|2019-10-29|2021-05-04|中国石油化工股份有限公司|Synergist in microbial remediation process of petroleum-polluted soil and preparation method thereof|

---

CN111608623A|2020-04-27|2020-09-01|夏文杰|Biological nano preparation applied to oil and gas resource exploitation|

---

CN111394284B|2020-04-28|2021-10-15|河北萌帮水溶肥料股份有限公司|Serratia marcescens MB21 and application thereof|

---

CN112159649A|2020-08-07|2021-01-01|长江大学|Biochemical viscosity reducer for thickened oil and preparation method thereof|

法律状态:
2021-10-19| B350| Update of information on the portal [chapter 15.35 patent gazette]|

优先权:

---

申请号 | 申请日 | 专利标题

---

US201762483425P| true| 2017-04-09|2017-04-09|

---

PCT/US2018/026724|WO2018191172A1|2017-04-09|2018-04-09|Microbial products and uses thereof to improve oil recovery|

---