Microbial protein particle

Abstract

To provide microbial protein particles that are excellent as food raw materials or aquatic feed raw materials.SOLUTION: The present invention provides microbial protein particles with a brightness (L* value) of 75 to 97.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to bacterial protein particles. [Background technology]

[0002] To help meet the growing need for food and feed, sustainable and renewable sources of amino acids and / or proteins are needed. There is also a need to reduce the emission of carbon dioxide and other greenhouse gases (GHGs) into the atmosphere, and to reduce water consumption and global energy consumption based on the use of coal, oil, and natural gas in food and feed production systems. For this reason, technology to produce bacterial proteins using hydrogen-oxidizing bacteria using carbon dioxide as a raw material is being researched.

[0003] For example, Patent Documents 1 and 2 disclose techniques for producing proteins using carbon dioxide as a raw material and hydrogen-oxidizing bacteria.

[0004] Patent Document 2 also discloses that the color of the obtained biomass slurry is brown, and after cell disruption by ultrasonic treatment, it becomes cream colored. [Prior art documents] [Patent documents]

[0005] [Patent Document 1] Special Publication No. 2020-526230 [Patent Document 2] Special Publication No. 2020-506708 Summary of the Invention [Problem to be solved by the invention]

[0006] However, as described in Patent Document 2, biomass products obtained using hydrogen-oxidizing bacteria are colored brown to cream, and when such biomass products are used as food ingredients or as raw materials for food production by mixing them with other food raw materials to produce food, there is a problem that the color of the entire food is impaired.

[0007] In addition, even if a biomass product deteriorates or discolors due to the effects of ultraviolet rays, oxygen in the air, mold, etc., it is difficult to notice such deterioration because the biomass product is colored.

[0008] The present invention has been made in consideration of such problems, and aims to provide bacterial protein particles which, when added to food, do not mar the appearance of the food and which allow easy detection of deterioration or denaturation. [Means for solving the problem]

[0009] The bacterial protein particles of the present invention have a brightness (L * The value is 75 to 97.

[0010] In addition, the bacterial protein particles of the present invention have a brightness (L * It is preferable that the value is 77 to 91.

[0011] The bacterial protein particles of the present invention preferably have an average particle size (D50) of 0.5 to 5.0 μm.

[0012] The bacterial cell protein particle of the present invention may contain alkanoic acid and / or a polymer of alkanoic acid. The alkanoic acid is preferably 3-hydroxybutyric acid. If the microorganism has the ability to synthesize alkanoic acid and / or a polymer of alkanoic acid and accumulate it in the microorganism, the particle composed of the microorganism becomes the bacterial cell protein particle of the present invention, and the bacterial cell protein particle is a particle or an aggregate thereof formed by encapsulating the alkanoic acid and / or a polymer of alkanoic acid in the cell membrane of the microorganism.

[0013] The bacterial protein particle of the present invention may be an ingredient of food or aquatic feed. In this case, the bacterial protein particle preferably contains alkanoic acid and / or alkanoic acid polymer derived from a microorganism. In addition, the alkanoic acid and / or alkanoic acid polymer is preferably encapsulated in the cell membrane of the microorganism.

[0014] Food ingredients comprising the bacterial protein particles of the present invention can be utilized, for example, in dairy products, dairy alternative products, meat products, meat alternative and / or imitation meat products, bakery products, confectionery, fortified foods, health and protein bars, protein powders, sports and / or energy drinks, and / or protein shakes and / or smoothies.

[0015] The aquatic feed containing the bacterial cell protein particles of the present invention can be used, for example, for fish or seafood farming. Since the bacterial cell protein particles of the present invention are not soluble in water, the aquatic feed containing the bacterial cell protein particles of the present invention has little impact on the environment and is preferable as a method for feeding aquatic organisms.

[0016] The bacterial protein particles of the present invention may have one peak in the particle size distribution curve. Since the bacterial protein particles of the present invention have one peak in the particle size distribution curve, the bacterial protein particles disperse quickly when added to water to obtain a protein dispersion. Furthermore, when the protein dispersion is stirred, a smooth solution with little viscosity is obtained. Since the particles can be well mixed with other feed ingredients or food ingredients, the particles are also suitable as food or aquatic feed for fish or seafood farming. [Brief description of the drawings]

[0017] [Figure 1] FIG. 1 is a TEM photograph (magnification 20,000 times) of hydrogen-oxidizing bacteria after main culture in an autotrophic medium, which is a bacterial protein particle of the embodiment. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0018] The bacterial protein particles of the present invention will be specifically described below. However, the present invention is not limited to the following configurations, and can be appropriately modified and applied within the scope of the present invention. Note that the present invention also includes a combination of two or more of the individual preferred configurations of the present invention described below.

[0019] The bacterial protein particles of the present invention have a brightness (L * The value is 75 to 97.

[0020] The term "cell protein particle" as used in the present invention refers to a particle or aggregate containing a protein contained in the cell of a microorganism. For example, a single-cell microorganism such as yeast or bacteria capable of producing a target protein is used, and in this case, the protein contained in the cell protein particle of the present invention may be a protein called a single cell protein (abbreviated as SCP). The cell may be dried as it is and used as feed or food, or may be mixed with feed or food, but the cell may be crushed to extract the protein, and the extract may be made into a particulate form to form the cell protein particle of the present invention. That is, the cell protein of the cell protein particle of the present invention may be a cell protein (crude protein) of a cell extract of each cell. For example, the cell protein particle of the present invention may contain one or more of a single-cell protein, a cell lysate, a protein isolate, a protein extract, a protein hydrolysate, a free amino acid, a peptide, an oligopeptide, or a combination thereof, which are processed from the microorganism and / or isolated or purified from the microorganism.

[0021] In the present invention, as described above, the bacterial protein particles are L * a * b * In the color system, L * The value is 75 to 97. * The value is L * a * b *It is an index that represents the lightness (brightness) of a color in the color space of a color system, and is expressed on a scale from 0 to 100, with 0 being black and 100 being white, and the higher the number, the brighter the color.

[0022] The bacterial protein particles of the present invention are not brown to cream colored particles as in the prior art, but have a lightness (L) of 75 to 97. * Since the bacterial protein particles of the present invention have a color value (L * Since the color of the bacterial cell is close to white, any deterioration in quality due to ultraviolet light, oxygen in the air, mold, etc. can be visually confirmed by the change in color, making it easy to check for deterioration or denaturation of bacterial protein particles.

[0023] In the present invention, the brightness (L * The color difference value) can be measured, for example, using a colorimeter used in the examples.

[0024] The bacterial protein particles of the present invention have a brightness (L * The lightness (L value) must be 75 or more. * If the lightness (L value) of the bacterial protein particles of the present invention is less than 75, the color and / or appearance of the food may be impaired when the food is mixed with other food ingredients, and discoloration due to deterioration may not be visible. * The brightness value must be 97 or less. If one attempts to adjust the brightness of bacterial protein particles to exceed 97, the protein must be denatured, resulting in the loss of the original functions and efficacy of the protein.

[0025] The bacterial cell protein particles of one embodiment of the present invention have an average particle size (D50) of 0.5 to 5.0 μm. In one embodiment, the bacterial cell protein particles of the present invention are bacterial cell protein particles in which alkanoic acid and / or alkanoic acid polymers are encapsulated by a cell membrane containing protein.

[0026] Fig. 1 is a TEM photograph (magnification 20,000 times) of the bacterial protein particles of the example. Fig. 1 shows that alkanoic acid and / or alkanoic acid polymers, which appear as white lumps in the photograph, are encapsulated in the cell membrane of the hydrogen-oxidizing bacteria.

[0027] The average particle size (D50) of bacterial cell protein particles can be measured, for example, after disintegrating aggregates of bacterial cell protein particles under the following conditions. In this specification, the "average particle size (D50)" refers to the particle size at an integrated value of 50% in the volume-based particle size distribution determined by a laser diffraction / scattering method. Measurement equipment: Particle size measurement equipment using laser diffraction and scattering method (e.g. MT3300EX manufactured by Microtrack Bell Co., Ltd.) Measurement range: 0.02~2000μm Light source: Semiconductor laser with wavelength of 780 nm, output of 3 mW Disintegration conditions: Add an appropriate amount of bacterial protein particles to water, stir with a spoon, and disintegrate by ultrasonic treatment for 5 minutes.

[0028] When the average particle size (D50) of the particles constituting the bacterial cell protein particles is 0.5 to 5.0 μm, the particles disperse quickly when the bacterial cell protein particles are added to a solution such as water to obtain a protein dispersion. Furthermore, when the protein dispersion is stirred, a smooth solution with little viscosity is obtained.

[0029] The bacterial protein particles of the present invention can be used in edible products such as dairy products, dairy substitute products, meat products, meat substitute and / or imitation meat products, bakery products, confectionery, fortified foods, health and protein bars, protein powders, sports and / or energy drinks, and / or protein shakes and / or smoothies, and can therefore be food ingredients for human or animal consumption. For example, dairy products include ready-to-drink dairy beverages, concentrated milk, evaporated milk, sweetened concentrated milk, milk powder, yogurt, fresh cheese, cheese, ice cream, and dairy spreads such as spreadable fresh cheese, cottage cheese, quark, crème fraîche, clotted cream, and cream cheese. In addition, the bacterial protein particles can be easily mixed uniformly with various dairy products, meat products, seasonings, extenders, fish meal, vegetable raw materials (soybean oil cake, corn gluten meal, etc.), animal raw materials (chicken meal, etc.), and grains (wheat), to produce foods and feeds. In addition, it is preferable that the particle size distribution curve of the bacterial cell protein particles of the present invention has only one peak. This is because it is believed that the water dispersibility of the particles will be higher. For example, the bacterial cell protein particles of the present invention may be contained in the above-mentioned foods and feeds at about 0.1 to 95% (w / w).

[0030] The bacterial cell protein particle of the present invention is preferably one in which an organic compound is encapsulated by a cell membrane. In this case, the protein contained in the bacterial cell protein particle of the present invention may be a protein contained in the cell membrane. In addition, the protein contained in the bacterial cell protein particle may be a protein as an organic compound encapsulated by a cell membrane. The amino acid sequence and three-dimensional structure constituting the protein contained in the bacterial cell protein particle are not particularly limited.

[0031] The organic compound is preferably one synthesized by a microorganism, and the organic compound synthesized by a microorganism is preferably at least one selected from the group consisting of alkanoic acid and / or polymers of alkanoic acid, proteins, amino acids, sugars, and ketones.

[0032] Examples of microorganisms that can be used to produce such bacterial cell protein particles include microorganisms that have the ability to synthesize organic compounds and accumulate them in their cells. The cell membrane of the microorganism contains protein, and in such a microorganism, the microorganism itself can become a bacterial cell protein particle in which an organic compound is encapsulated in the cell membrane containing protein.

[0033] Microorganisms from which bacterial protein particles are derived include autotrophic microorganisms that can use carbon dioxide (CO2) as their main carbon source, such as hydrogen-oxidizing bacteria, which are microorganisms that use CO2 and hydrogen to synthesize organic compounds and accumulate them within the bacterial cell. For example, hydrogen-oxidizing bacteria may be, but are not limited to, those belonging to the genera Alcaligenes, Aquaspirillum, Arthrobacter, Azospirillum, Bacillus, Bradyrhizobium, Burkholderia, Calderobacterium, Cupriavidus, Derxia, Flavobacterium, Frankia, Helicobacter, Hydrogenobacter, Hydrogenomonas, Hydrogenovibrio, Mycobacterium, Microcyclus, Norcadia, Paracoccus, Pseudmonas, Pyrinomonas, Streptomyces, Ralstonia, Renobacter, Rhizobium, Rhodococcus, Variovorax, and Xantobacter. In addition, the microorganism may be a mutant selected for a specific characteristic, a naturally occurring strain, or a genetically engineered variant of a naturally occurring strain. The microorganism from which the bacterial protein of the present invention is derived may be a so-called non-GMO. Preferably, the microorganism from which the bacterial protein of the present invention is derived may be a generally recognized as safe (GRAS) microorganism. The microorganism from which the bacterial protein particles are derived may be one or more microorganisms. Aerobic or anaerobic bacteria may be used as the hydrogen-oxidizing bacteria.

[0034] Hydrogen oxidizing bacteria that are aerobic bacteria include Acidvorax facilis, Achromobacter xylosoxidans, Alcaligenes latus, Cupriavidus necator, Ancylobacter aquaticus, Aquaspirillum autotrophicum, Azospirillum lipoferum, Bradyrhizobium japonicum, Derxia gummosa, Hydrogenophaga palleronii, Hydrogenophaga flava, Hydrogenophaga pseudoflava, Paracoccus denitrificans, Variovorax paradoxus, Xanthobacter autotrophicus, tusciae, Bacillus These microorganisms use CO2, H2, and O2 to synthesize organic compounds (proteins, 3-hydroxybutyric acid, and its aggregate polyhydroxybutyric acid (PHB), etc.) and accumulate them in the bacterial cells.

[0035] Anaerobic hydrogen-oxidizing bacteria include Acetobacterium woodii DSM 1030, Acetohalobium arabaticum DSM 5501, Carboxydothermus hydrogenoformans Z-2901, Clostridium aceticum DSM 1496, Clostridium autoethanogenum DSM 10061, Clostridium carboxidivorans P7, Clostridium ljungdahlii DSM 13528, Clostridium scatologenes ATCC 25755, Clostridium sticklandii DSM 519, Eubacterium limosum KIST612, Eubacterium limosum SA11, Moorella thermoacetica ATCC 39073, Moorella thermoacetica DSM 521, Moorella thermoacetica DSM 2955, Clostridium difficile 630, Clostridium difficile CD196, Clostridium difficile M120, Clostridium difficile 630, Clostridium difficile 630 Deltaerm, Thermoacetogenium phaeum DSM 12270, Thermoanaerobacter kivui LKT-1, Treponema primitia ZAS-2. These microorganisms use CO2, CO and H2 to synthesize organic compounds and accumulate them in the bacterial cell.

[0036] The organic compounds synthesized by these anaerobic bacteria and accumulated within the bacteria include one or more of acetic acid, ethanol, acetate, 1-butanol, butyrate, 2,3-butanediol, butadiene, methyl ethyl ketone (2-butanone), ethylene, acetone, isopropanol, lipids, 3-hydroxypropionate (3-HP), terpenes, isoprene, fatty acids, 2-butanol, 1,2-propanediol, 1-propanol, 1-hexanol, 1-octanol, chorismate-derived products, 3-hydroxybutyrate, 1,3-butanediol, 1,4-butanediol, 2-hydroxyisobutyrate or 2-hydroxyisobutyric acid, isobutylene, adipic acid, 1,3-hexanediol, 3-methyl-2-butanol, 2-buten-1-ol, isovalerate, isoamyl alcohol, and monoethylene glycol.

[0037] The bacterial protein particles are preferably those in which alkanoic acid and / or a polymer of alkanoic acid is encapsulated in a cell membrane containing protein.

[0038] In the present specification, the alkanoic acid includes alkanoic acids that do not have a substituent other than a carboxy group (acetic acid, propanoic acid, butanoic acid, etc.), and alkanoic acids that have a substituent (hydroxyalkanoic acid, aminoalkanoic acid, etc.). In particular, hydroxyalkanoic acid is preferable, and 3-hydroxybutyric acid is preferable. As a polymer of alkanoic acid, polyhydroxyalkanoic acid is preferable, and as a polyhydroxyalkanoic acid, polyhydroxybutyric acid (PHB), which is an aggregate of 3-hydroxybutyric acid, is preferable.

[0039] Such bacterial protein particles in which alkanoic acid and / or polymers of alkanoic acid are encapsulated by cell membranes can be used, for example, for aquatic feed. That is, the bacterial protein particles of the present invention can be suitably used as single-cell feed. Since alkanoic acid and / or polymers of alkanoic acid are easily soluble in water, they may dissolve and disperse in water before being eaten by cultured organisms (e.g., fish, shrimp, etc.), resulting in a decrease in feed efficiency. As a result, an increase in the amount of feed and a decrease in water quality due to dissolved organic compounds may become a problem. However, in the present invention, since alkanoic acid and / or polymers of alkanoic acid are encapsulated by cell membranes containing proteins and are hardly dissolved in water, excellent feed efficiency can be obtained and the impact on the environment is thought to be small. In addition, alkanoic acid and / or polymers of alkanoic acid are known to activate the action of denitrifying bacteria in water, and since they reduce and purify nitrates generated from fish and shellfish excrement, it may be particularly advantageous to use them as aquatic feed for closed circulation aquaculture systems.

[0040] Therefore, the present invention relates to an aquatic feed containing bacterial cell protein particles. Even if additional nutritional components (carbohydrates, amino acids, taurine, fatty acids, etc.), vitamins, minerals, phosphorus, calcium, antioxidants, etc. are mixed into the aquatic feed in consideration of the health, growth, etc. of the cultured organism, the aquatic feed can be uniformly dispersed. Aquatic feed with excellent growth efficiency can be produced. The bacterial cell protein particles of the present invention can be used as feed for any aquatic organism, and are preferably used as feed for fish, invertebrates such as octopus and squid, shellfish, shrimp, and other seafood. Therefore, the present invention relates to a method for feeding aquatic organisms, which is characterized by feeding the aquatic organisms with aquatic feed containing the bacterial cell protein particles of the present invention.

[0041] When bacterial protein particles form aggregates, the average particle size (D50) of the aggregates can be measured under the following conditions. Measurement equipment: Particle size measurement equipment using laser diffraction and scattering method (e.g. MT3200 manufactured by Microtrack Bell Co., Ltd.) Measurement range: 0.2 to 1400 μm Light source: Semiconductor laser with wavelength of 780 nm, output of 3 mW

[0042] The bacterial protein particles of the present invention can be obtained, for example, by culturing a microorganism by the following method.

[0043] (Preparation of heterotrophic medium) Nutrient components used in heterotrophic media include Bacto yeast extract, tryptone, beef extract, and the like.

[0044] (Preparation of autotrophic medium) Ingredients that may be included in the autotrophic medium include nitrogen sources such as ammonia, ammonium (e.g., ammonium chloride (NH4Cl), ammonium sulfate ((NH4)2SO4), ferric ammonium citrate), nitrates (e.g., potassium nitrate (KNO3)), urea, or organic nitrogen sources; phosphates (e.g., sodium dihydrogen phosphate (NaH2PO4), sodium hydrogen phosphate (Na2HPO4), potassium dihydrogen phosphate (KH2PO4), phosphoric acid (H3PO4), potassium dithiophosphate (K3PS2O2), potassium orthophosphate (K3PO4), phosphoric acid (H3PO4), potassium dithiophosphate (K3PS2O2), potassium orthophosphate (K3PO4), phosphoric acid (H3PO4), potassium dithiophosphate (K3PS2O2), potassium dihydrogenphosphate (K3PO4 ... dipotassium hydrogen (K2HPO4); sulfates (e.g., ammonium sulfate ((NH4)2SO4), magnesium sulfate (heptahydrate (MgSO4·7H2O)); yeast extract; chelated iron; potassium salts, potassium iodide (KI), potassium bromide (KBr); and other inorganic salts, minerals, and micronutrients (e.g., sodium chloride (NaCl), or magnesium chloride (MgCl2), calcium chloride (CaCl2) or calcium carbonate (CaCO3), manganese sulfate heptahydrate (MnSO4·7H2O) or manganese chloride (MnCl2), iron(III) chloride hexahydrate hydrate (FeCl3·6H2O), ferrous sulfate heptahydrate (FeSO4·7H2O) or ferrous chloride tetrahydrate (FeCl2·4H2O), sodium bicarbonate (NaHCO3) or sodium carbonate (Na2CO3), zinc sulfate (ZnSO4) or zinc chloride (ZnCl2), ammonium molybdate (NH4MoO4) or sodium molybdate dihydrate (Na2MoO4·2H2O), cuprous sulfate (CuSO4) or copper chloride dihydrate (CuCl2·2H2O), cobalt chloride hexahydrate (CoCl2·6H2O), aluminum chloride hexahydrate (AlCl3·6H2O), lithium chloride (LiCl), boric acid (H3BO3), nickel(II) chloride hexahydrate (NiCl2·6H2O), tin chloride monohydrate (SnCl2·H2O), barium chloride dihydrate (BaCl2·2H2O), copper selenate pentahydrate (CuSeO4·5H2O) or one or more inorganic salts of sodium selenite (Na2SeO3), sodium metavanadate (NaVO3), and chromium salts. For example, the inclusion of the above-mentioned micronutrients can promote the growth and proliferation of hydrogen-oxidizing bacteria.

[0045] Suitable inorganic salts which may be included in the autotrophic medium include ammonium chloride, sodium hydrogen phosphate, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, sodium bicarbonate, ferric ammonium citrate, iron(III) chloride hexahydrate, ammonium chloride, calcium chloride, nickel(II) chloride hexahydrate, and the like.

[0046] The average particle size (D50) of bacterial protein particles can be adjusted by adjusting the nitrogen content of the autotrophic medium. Specifically, increasing the concentration of ammonium salts, such as ammonium sulfate, ammonium citrate (ammonium tricitrate), and ammonium chloride, increases the average particle size of bacterial protein particles, and conversely, decreasing the concentration decreases the average particle size of bacterial protein particles. This is presumably because there is a correlation between the amount of protein produced and the nitrogen concentration in the medium.

[0047] The nitrogen content can be adjusted within the range of 0.01 g / L to 10 g / L in terms of N.

[0048] The above-mentioned components are used to carry out cultivation as described below, and somatic protein particles can be prepared from the cultured microorganism. The seed bacteria is inoculated into the above-mentioned autotrophic medium and initial culture is carried out. The initial culture of microorganisms is harvested by centrifugation. The collected microorganisms are inoculated into the above-mentioned autotrophic medium and cultured. The cultured microorganisms are separated and collected by a technique such as centrifugation. The collected microorganisms are dried to obtain bacterial protein particles.

[0049] The drying can be carried out by any drying method such as heat drying, freeze drying, or spray drying.

[0050] As the spray dryer, for example, a disk-type device (a device in which droplets are dropped onto a rotating disk, and the liquid blown off the disk is dried and granulated by contact with hot air) can be preferably used. When a spray dryer is used, the drying conditions can be, for example, the following conditions. Rotation speed: 5,000~50,000rpm Raw liquid processing capacity: 0.1-80kg / h Inlet temperature: 50~300℃ Cyclone differential pressure: 0.10~10kPa

[0051] Alternatively, the heat drying may be performed in air at, for example, 100°C for a predetermined time in a dryer. The size of the bacterial protein particles can be adjusted by adjusting the drying conditions. Alternatively, the bacterial protein particles may be obtained by putting the bacterial protein particles in water and dispersing them by shaking.

[0052] As a freeze-drying method, for example, a method in which the mixture is frozen to −35° C. in a freezer, then reduced in pressure to 50 Pa and dried for 36 hours can be used.

[0053] The initial culture and the main culture may use the same medium or different media. For example, a heterotrophic medium may be used for the initial culture and an autotrophic medium may be used for the subsequent main culture. Alternatively, an autotrophic medium may be used for both the initial culture and the main culture, or a heterotrophic medium may be used for both the initial culture and the main culture.

[0054] When culturing hydrogen oxidizing bacteria, CO2 and H2 are blown into the medium during the culture. If the hydrogen oxidizing bacteria are aerobic bacteria, it is preferable to further blow O2 into the medium during the culture.

[0055] The culture conditions in the initial culture and the main culture are not particularly limited, and the culture conditions for ordinary microorganisms can be used. For example, the molar concentration ratio of each gas of H2, O2, and CO2 blown into the medium is preferably H2:O2:CO2=4-90:2-10:2-20. The substrate gases of H2, O2, and CO2 supplied are not required to be 100% pure of the three-component system, and may contain gases and impurities other than hydrogen, carbon dioxide, and oxygen. For example, in the culture tank, the hydrogen concentration may be within the range of 80-99%, preferably 90-95%, the oxygen concentration may be within the range of 2-8%, preferably 4-6%, and the carbon dioxide concentration may be within the range of 1-15%, preferably 1-10%.

[0056] The flow rates of the gases introduced into the culture tank during culture can be, for example, as follows. CO2:10~1000mL / min H2:500~2000mL / min O2:10~1000mL / min When O2 is introduced as air, the flow rate of the air should be adjusted so that the flow rate of O2 falls within the above range.

[0057] The substrate gas may be hydrogen exhaust gas containing by-product hydrogen and the like discharged from a factory, combustion exhaust gas containing CO2 and the like discharged from a factory, or air. When CO2 is collected from exhaust gas, it can be concentrated and used. As a method for concentration, the pressure swing adsorption method (PSA method) in which CO2 is adsorbed on zeolite or the like under pressure and concentrated, and the CO2 is released by reducing the pressure, or the so-called amine method in which CO2 is absorbed in an amine solution and concentrated, and the CO2 is released by heating the solution, can be used. In addition, H2 can be reused by separating only H2 using a gas separation membrane and refluxing it to the culture tank.

[0058] The temperature during the culture is not particularly limited, and may be set to a temperature suitable for the culture of the microorganism used, for example, 25 to 35° C. The culture time is also not particularly limited, and may be, for example, 30 to 80 hours.

[0059] For example, the culture solution may be provided with pH buffering ability by adding sodium hydrogen phosphate (Na2HPO4) and sodium dihydrogen phosphate (KH2PO4), or sodium bicarbonate (NaHCO3). As the culture proceeds, the pH of the culture solution may become lower (biased toward the acidic side) due to the dissolution of the products of the microorganisms in the culture solution or due to the dissolved carbon dioxide, so in that case it is preferable to add an alkaline component to adjust the pH. This can prevent inhibition of the growth and proliferation of the microorganisms. As the alkaline component, for example, ammonia water or the like can be used, and ammonia is thought to be able to replenish the nitrogen source in the culture solution consumed by the microorganisms. The pH is preferably adjusted to around 6.50 to 6.75.

[0060] In addition, the microorganisms recovered from the main culture may be used as seed culture, or the microorganisms recovered from the main culture may be used in place of the microorganisms that have undergone initial culture to perform the main culture.

[0061] After the cultivation is completed, the microorganisms removed from the culture medium are decolorized and washed using water and an organic solvent, thereby adjusting the brightness of the bacterial protein particles. The decolorization and washing method is preferably a water washing step followed by decolorization and washing using an organic solvent. The organic solvents that can be used include ethanol, methanol, propanol, acetone, dimethyl sulfoxide (DMSO), butanediol, hexane, chloroform, and dichloromethane, but ethanol, which is less toxic to living organisms, is preferable, especially for food and feed applications.

[0062] In addition, in Patent Document 2, an alkaline aqueous solution is used for decolorization washing, but this is inappropriate from the viewpoint of brightness adjustment, because the alkaline aqueous solution not only breaks down proteins, reducing the protein content, but also discolors the product (to a brightness of about 65), deteriorating its appearance. EXAMPLES

[0063] EXAMPLES Hereinafter, examples that more specifically disclose the present invention will be described, however, the present invention is not limited to these examples.

[0064] <Production of bacterial protein particles> Example 1 (Preparation of autotrophic medium) An autotrophic medium having the following composition was prepared: Sodium hydrogen phosphate 3.57g / L Potassium dihydrogen phosphate 1.50g / L Magnesium sulfate 0.2g / L Iron(III) chloride hexahydrate 0.005g / L Hydrochloric acid 0.1mL / L Ammonium chloride 1.6g / L Calcium chloride 0.01g / L Nickel(II) chloride aqueous solution 0.0002g / L The above was made up to 1L with distilled water. The nitrogen content is 0.42 g / L in terms of N.

[0065] (1) The hydrogen-oxidizing bacterium Cupriavidus necator (H16 strain) was grown in the above autotrophic medium.

[0066] (2) The hydrogen-oxidizing bacteria Cupriavidus necator (H16 strain) grown in the above autotrophic medium was added to the OD 600 The OD was inoculated to be 0.1. 600 (Absorbance at a wavelength of 600 nm) was measured using a commercially available spectrophotometer (BioSpec-mini spectrophotometer, manufactured by Shimadzu Corporation).

[0067] (3) The autotrophic medium inoculated in (2) was placed in a culture tank, and CO2 gas was bubbled into the culture tank at 24 mL / min, H2 gas at 1700 mL / min, and air (gas containing O2) at 80 mL / min in terms of oxygen per 5 L of culture solution, and hydrogen-oxidizing bacteria were cultured for 12 hours while stirring at 500 rpm with a stirring blade. The culture temperature was 30°C.

[0068] (4) After the cultivation in (3), the hydrogen-oxidizing bacteria were decolorized and washed. The decolorization and washing process was carried out according to the following procedure. (First water washing step): After cultivation, 200 mL of culture liquid was removed from the culture tank and centrifuged at 10,000 g, 5° C., and 5 min to separate the supernatant from the solid portion. After discarding the supernatant, 200 mL of water was added to the solid portion to disperse the hydrogen-oxidizing bacteria, which was then centrifuged (10,000 g, 5° C., 5 min) to separate the supernatant from the solid portion, after which the supernatant was discarded. This first water washing step was performed twice. (Ethanol washing step): 10 mL of ethanol was added to the hydrogen-oxidizing bacteria that had been through the first water washing step twice, and the hydrogen-oxidizing bacteria were dispersed and then centrifuged (10,000 g, 5° C., 5 min) to separate the supernatant and solid portion, and the supernatant was discarded. In Example 1, this ethanol washing step was carried out once. (Water washing step after ethanol washing step (second water washing step)): 200 mL of water was added to the hydrogen oxidizing bacteria that had been subjected to the above ethanol washing step to disperse the hydrogen oxidizing bacteria, which were then centrifuged (10,000 g, 5° C., 5 min) to separate the supernatant and solid portion, and the supernatant was discarded. This second water washing step after the ethanol washing step was performed once. That is, the decolorizing and washing step in Example 1 was carried out by carrying out the first water washing step twice, followed by one ethanol washing step, and finally carrying out one second water washing step.

[0069] (5) 100 g of the slurry-like solid portion after the decolorization and washing process was frozen in a freezer to -35°C. Then, using a freeze dryer (Tokyo Rikakikai Co., Ltd., product name FDU-1200), the pressure was reduced to 50 Pa and freeze-dried for 36 hours.

[0070] (6) Then, 20 g of the dried solid portion was crushed three times for 10 seconds using a mixer (T-fal Mill, model number BL13C5JP) on high mode to obtain the bacterial cell protein particles of Example 1.

[0071] Example 2 The bacterial protein particles of Example 2 were obtained under the same conditions as in Example 1, except that the ethanol washing step in the decolorizing washing step was performed four times.

[0072] Example 3 The bacterial protein particles of Example 2 were obtained under the same conditions as in Example 1, except that the ethanol washing step in the decolorizing washing step was performed three times.

[0073] Comparative Example 1 After culturing the hydrogen-oxidizing bacteria in the same manner as in Example 1, the cultured hydrogen-oxidizing bacteria was subjected to only a water washing step. That is, the first water washing step of the decolorizing washing step in Example 1 was carried out once, but the subsequent ethanol washing step and the second water washing step after the ethanol washing step were not carried out.

[0074] 100 g of the slurry-like solid portion after the washing process was placed on a drying pad (560 x 413 x 21 mm, slurry height 2 mm or less), placed in a constant temperature dryer (forced convection dryer OF-600V, manufactured by AS ONE Corporation), and dried in air at 100°C for 5 hours to obtain bacterial protein particles of Comparative Example 1.

[0075] Comparative Example 2 Similar to Comparative Example 1, the first water washing step was carried out three times.

[0076] Comparative Example 3 The procedure was the same as in Comparative Example 1, but in the first washing step, washing was performed once using a 0.1 mM aqueous sodium hydroxide solution instead of water.

[0077] <Lightness(L * Measurement of (value) The brightness (L * values) were measured. Measuring device: Colorimeter (manufactured by Nippon Denshoku Industries Co., Ltd., product name: ZE6000) Observation light source: C light source halogen lamp 12V 20W Observation conditions: 2° field of view Method: SCE (specular reflection light elimination) method Measurement item: L * a * b * L according to color system * value Measurement procedure: About 80% of the powder of bacterial protein particles of Examples 1 to 3 and Comparative Examples 1 to 3 was placed in a glass petri dish for powder samples, and the dish was placed in a measurement holder with the lid on and tapped 30 times to make the powder densely packed on the bottom of the glass petri dish for powder samples. The measurement holder was placed in a color difference meter and light was irradiated from the bottom to measure the L. * a * b * L according to color system * The values ​​were measured.

[0078] <Appearance evaluation of foods containing bacterial protein particles> Yogurt (manufactured by Meiji Co., Ltd., product name "Bulgaria Yogurt LB81 Plain") 30 g was mixed with 1 g of each of the bacterial protein particles of Examples 1 to 3 and Comparative Examples 1 to 3 to prepare yogurt containing added bacterial protein particles.

[0079] A sensory evaluation of each prepared yogurt was carried out by five experienced panelists. The appearance of the yogurt containing added bacterial protein particles was visually observed for "yogurt-likeness" and rated on a 5-point scale according to the following criteria. The average score of each panelist's evaluation was also calculated. 5 points: Particularly good, comparable to yogurt. 4 points: Good. 3 points: Generally no discomfort. 2 points: Somewhat uncomfortable. 1 point: There is a strong sense of discomfort.

[0080] The luminosity (L * The results of the evaluation of the appearance of the foods containing each bacterial protein particle are summarized in Table 1.

[0081] [Table 1]

[0082] The bacterial cell protein particles of Examples 1 to 3 have a higher brightness (L * The color was close to white.

[0083] <Measurement of average particle size (D50)> The bacterial protein particles of Example 1 were added to water, stirred with a spoon, and disintegrated by ultrasonic treatment for 5 minutes to obtain a dispersion of bacterial protein particles. The obtained dispersion was placed in a particle size measuring device (MT3300EX manufactured by Microtrack Bell Co., Ltd.) and the average particle size (D50) of the bacterial protein particles of Example 1 was measured by the measurement method described in the specification.

[0084] The average particle size (D50) of the bacterial cell protein particles of Example 1 was 1.6 μm, and the particle size distribution curve had one peak. As described above, since the average particle size of the bacterial cell protein particles of this example is relatively small, and the particle size distribution curve had one peak, it is considered that the bacterial cell protein particles of this example have excellent water dispersibility.

[0085] Furthermore, since the bacterial cell protein particles of the examples are hydrophilic 3-hydroxybutyric acid polymers encapsulated in cell membranes containing proteins, it is assumed that the particles are unlikely to form hydrogen bonds with each other, which is also the reason for their excellent dispersibility. It was also found that the average particle size of the bacterial cell protein particles increases by increasing the oxygen concentration supplied to the medium and the nitrogen content of the medium (specifically, the ammonium chloride concentration) (data not shown).

[0086] As explained above, the bacterial protein particles of the present invention have a brightness of 75 to 97, which is close to white, and therefore do not impair the color or appearance of food or feed even when mixed with other food or feed ingredients, making them excellent as food and feed ingredients. Therefore, they are easy to mix with other food or feed ingredients for cooking and feed processing, and are suitable for food and feed applications. Food products include food products, dairy products, dairy product substitutes, meat products, meat product substitutes and / or artificial meat products, bakery products, confectioneries (cookies, cakes, etc.), cereals, etc. In addition, feeds include feeds for aquaculture and feeds for livestock.

[0087] In addition, the bacterial protein particles of the present invention have a brightness of 75 to 97, which is close to white, so discoloration due to deterioration in quality caused by ultraviolet rays, oxygen in the air, mold, etc. is easily visible, making it easy to control the quality as a food or feed ingredient.

[0088] Furthermore, the bacterial protein particles of the present invention are easily dispersible in water and the viscosity of the resulting dispersion is not high. This means that it is easy to uniformly mix the bacterial protein particles with substances other than the bacterial protein particles, such as seasonings, bulking agents, fish meal, vegetable raw materials (soybean meal, corn gluten meal, etc.), animal raw materials (chicken meal, etc.) and grains (wheat), thereby offering the advantage of facilitating the production of foods and feeds.

Claims

1. Lightness (L * Bacterial protein particles having a molecular weight (Mw) of 75 to 97.

2. Lightness (L * 2. The bacterial protein particle according to claim 1, wherein the molecular weight (Mg) is 77 to 91.

3. 2. The bacterial protein particles according to claim 1, having an average particle size (D50) of 0.5 to 5.0 μm.

4. The bacterial cell protein particle according to claim 1 , which contains an alkanoic acid and / or a polymer of an alkanoic acid.

5. The bacterial protein particle according to claim 4, wherein the alkanoic acid is 3-hydroxybutyric acid.

6. A food material comprising the bacterial protein particles according to claim 1.

7. 2. A raw material for aquatic feed comprising the bacterial protein particle according to claim 1.

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