WO2020026794A1 - Procédé de culture de microalgues hétérotrophes à l'aide d'un effluent de broyeur d'huile de palme (pome) et procédé de production de dha - Google Patents

Procédé de culture de microalgues hétérotrophes à l'aide d'un effluent de broyeur d'huile de palme (pome) et procédé de production de dha Download PDF

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WO2020026794A1
WO2020026794A1 PCT/JP2019/027981 JP2019027981W WO2020026794A1 WO 2020026794 A1 WO2020026794 A1 WO 2020026794A1 JP 2019027981 W JP2019027981 W JP 2019027981W WO 2020026794 A1 WO2020026794 A1 WO 2020026794A1
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pome
dha
culture
aurantiochytrium
culture method
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PCT/JP2019/027981
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Japanese (ja)
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清志 多田
信 渡邉
吉田　昌樹
順子 伊藤
敏秀 中島
ゴータマ マイケル
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ＭｏＢｉｏｌ株式会社
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Priority to MYPI2020001547A priority Critical patent/MY188244A/en
Priority to JP2019571086A priority patent/JP6709484B1/ja
Publication of WO2020026794A1 publication Critical patent/WO2020026794A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/12—Unicellular algae; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats

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  • the present invention provides a method for culturing heterotrophic microalgae and a method for producing DHA, using a palm oil mill effluent (Palm Oil Mill Effluent: POME) as a medium.
  • a palm oil mill effluent Palm Oil Mill Effluent: POME
  • palm oil factory effluent (hereinafter referred to as POME) is a major environmental and economic problem.
  • POME palm oil factory effluent
  • oxidation pond treatment ⁇ anaerobic treatment
  • aerobic treatment ⁇ aerobic treatment.
  • fermentation gas causes odor, damage to insects and pests, water pollution, and the like, and methane gas generated by anaerobic treatment is released to the atmosphere.
  • the global warming potential of methane is 21 times greater than that of carbon dioxide, which has a negative impact on global warming.
  • POME causes emission of warming substances and water pollution, causing widespread and serious environmental problems such as global warming, deterioration of living environment, and adverse effects on wildlife.
  • the palm oil production industry is a key industry in Indonesia, Malaysia, etc.
  • solving the environmental problems of POME is crucial for the sustainable development of the national society where the palm oil industries, such as Indonesia and Malaysia, are fostering. Is an important issue.
  • microalgae have been proliferated, and lipids or proteins, vitamins, and other valuable materials used as raw materials for fuels, feeds, health foods, and the like have been produced and stored in the cells of the microalgae. It is used for fuel and food.
  • production and storage of high value-added valuables such as ⁇ -3 polyunsaturated fatty acids have attracted attention as a component that can be immediately industrialized.
  • microalgae is also used in wastewater treatment such as purification of sewage and industrial wastewater (Patent Document 3) and exhaust gas treatment for absorbing and utilizing carbon dioxide (Patent Document 4).
  • wastewater treatment such as purification of sewage and industrial wastewater (Patent Document 3) and exhaust gas treatment for absorbing and utilizing carbon dioxide (Patent Document 4).
  • Patent Document 3 purification of sewage and industrial wastewater
  • Patent Document 4 exhaust gas treatment for absorbing and utilizing carbon dioxide
  • HABIB et. Al. “Growth and Nutritional Values of Moinamicrura Fed on Chlorella vulgaris Grown in Digested Palm Oil Mill Effluent”, Asian Fisheries Science, 16, 107-119, 2003 Putri et. Al. “Investigation of Microalgae for High Lipid Content using Palm Oil Mill Effluent (Pome) as Carbon Source, 2011 International Conference on Environment and Industrial Innovation, IPCBEE, 12,2011-89 Nwuche et. Al. “Use of Palm Oil Mill Effluent as Medium for Cultivation of Chlorella sorokiniana”, British Biotechnoloty Journal, 4 (3) :) 305-316, 2014 Nur et. Al.
  • the present invention provides a culture method capable of efficiently growing microalgae using POME discharged from a palm oil production process, and efficiently producing high value-added valuables from microalgae. It is an object to provide a production method.
  • the present inventors have conducted intensive studies in order to solve the above-mentioned problems.
  • using POME with or without an organic carbon source such as sugar and a mineral component as needed, ⁇ - such as DHA (docosahexaenoic acid).
  • ⁇ - such as DHA (docosahexaenoic acid).
  • the method for culturing heterotrophic microalgae according to the present invention for solving the above-mentioned problems is characterized by using POME containing squeezed juice obtained by squeezing out oil from palm fruit in the palm oil production process, using a high value-added component. Characterized by heterotrophic cultivation of microalgae that produce.
  • the method of culturing microalgae according to the present invention is characterized in that palm fruit is squeezed to supply POME containing pomace juice produced after palm oil production.
  • the present application provides the following inventions.
  • (1) A method for culturing heterotrophic microalgae using a medium containing POME.
  • (2) The culture method according to (1), wherein the heterotrophic microalgae is an algae that produces ⁇ -3 polyunsaturated fatty acids.
  • (3) The culture method according to (2), wherein the ⁇ -3 polyunsaturated fatty acid is DHA.
  • (4) The culture method according to any one of (1) to (3), wherein the algae is a strain of algae belonging to Thraustochytriales.
  • the culture method according to (4), wherein the Thraustochytriales is a species belonging to the genus Aurantiochytrium.
  • the medium further contains a sugar component.
  • the sugar component is one or more saccharides selected from glucose, galactose, fructose, maltose, sucrose, lactose, oligosaccharides, sugar alcohols such as glycerol, and the like.
  • the medium further includes a mineral component.
  • the mineral components include potassium sulfate, magnesium sulfate, iron sulfate, ammonium sulfate, or sulfates such as copper sulfate, nickel sulfate, and zinc sulfate; phosphates such as potassium phosphate; carbonates such as calcium carbonate; Chlorides such as cobalt, manganese chloride, sodium chloride and calcium chloride; alkali metal oxides; selenites and molybdates such as sodium molybdate and sodium selenite; halides such as potassium bromide or potassium iodide
  • DHA-producing algae such as Aurantiochytrium perimacinum 4W-1b strain can be well grown at a high growth rate not found in conventional microalgae, DHA production was achieved.
  • FIG. 1 shows the change over time of the algal cell concentration of Aurantiochytrium persiminanum 4W-1b when an artificial seawater-salt-supplemented medium adjusted so that the addition rate of the POME supernatant was 10 to 50% was used. (OD660).
  • FIG. 2 shows the total lipids of the Aurantiochytrium ⁇ Limasinum ⁇ 4W-1b strain when using a medium supplemented with artificial seawater salts adjusted so that the addition rate of the POME supernatant was 10 to 50% (left figure) and The production of DHA (right figure) is shown.
  • FIG. 1 shows the change over time of the algal cell concentration of Aurantiochytrium persiminanum 4W-1b when an artificial seawater-salt-supplemented medium adjusted so that the addition rate of the POME supernatant was 10 to 50% was used. (OD660).
  • FIG. 2 shows the total lipids of the Aurantiochytrium ⁇ Limasinum ⁇ 4W
  • FIG. 3 shows an Aurant in the case of using a medium in which glucose and artificial seawater salts were added to a POME supernatant (100%) and a diluent adjusted so that the addition ratio of the POME supernatant was 10 to 75%.
  • the change with time of the algal body concentration of the strain Okitorium Limasinum 4W-1b is shown (OD660).
  • FIG. 4 shows the results obtained by using POME supernatant (100%) and a medium obtained by adding glucose and artificial seawater salts to a diluent adjusted so that the addition rate of the POME supernatant becomes 10 to 75%.
  • 4 shows the correlation between the rate of addition of the fresh liquor and the amount of growth of the Aurantiochytrium limacinum 4W-1b strain.
  • FIG. 5 shows the Aurant in the case of using a medium in which glucose and artificial seawater salts were added to a POME supernatant (100%) and a diluent adjusted so that the addition rate of the POME supernatant was 10 to 75%.
  • 2 shows total lipid and DHA production of the strain Ochtorium limacinum 4W-1b.
  • FIG. 6 shows glucose and 0.00%, 0.38%, 0.75%, 1.50%, 3.00% (wt%) red sea salt added to POME supernatant (100%).
  • 5 shows the time-dependent change (OD660) and the DHA concentration of the algal body concentration of the Aurantiochytrium limacinum 4W-1b strain when the used culture medium was used.
  • FIG. 7 shows the production of total lipids and DHA of various strains of the genus Aurantiochytrium using a medium in which glucose and artificial seawater salts were added to the POME supernatant (100%).
  • palm oil production processes include steam treatment for inactivating palm fruit clusters (Fresh Fruit Bunch: FFB), separation of palm fruit and palm empty clusters (Empty Fruit Bunch: EFB), and fruit parts from palm fruit ( It goes through a pretreatment process of separation of mesoca) and seed part.
  • FFB Frsh Fruit Bunch
  • EFB Easy Fruit Bunch
  • the process proceeds to the extraction and separation and purification of crude palm oil from the fruit, producing high-purity palm oil.
  • POME in the present invention means used steam water, a mixed effluent of oil juice and unrecovered oil, and / or unrecovered oil obtained by centrifuging the above-mentioned mixed liquid.
  • POME is rich in carbohydrates, organic acids, vitamins, amino acids, peptides, proteins, minerals, etc., derived from raw materials.
  • the POME used for culturing the algae of the present invention may be before or after the oxidation pond treatment, the anaerobic treatment, or the aerobic treatment. In some embodiments, it is preferable to use POME before performing these oxidation pond treatment, anaerobic treatment, and aerobic treatment.
  • @POME contains various solids derived from raw materials.
  • the POME may or may not contain a solid, and is not limited.
  • clear POME obtained by a known means such as centrifugation or filtration may be used for the medium.
  • POME is sterilized by known means such as filtration sterilization, autoclave sterilization, boiling sterilization, and radiation sterilization, but may be sterilized by known means such as sodium hypochlorite and ozone treatment.
  • the content of POME in the medium of the heterotrophic microalgae is not limited, but is, for example, a concentration of 1%, at least 10%, at least 25%, at least 50%, 75%, 100% (% by volume). May be contained in the culture solution.
  • the medium may be adjusted to have a composition suitable for culturing DHA-producing algae.
  • the additive include saccharides, organic acids, inorganic acids, organic bases, inorganic bases, vitamins, amino acids, peptides, proteins, and minerals (including natural seawater, artificial seawater, and the like).
  • it may be preferable to add sugars and / or minerals.
  • it may be preferable not to add minerals. If no mineral is added, there are advantages in that metal corrosion of the apparatus can be prevented, wastewater treatment and discharge after culturing are easy, and the cost of the culture medium is kept low.
  • the pH can be appropriately adjusted by adding an appropriate acid or base.
  • the suitable pH of the medium depends on the type of DHA-producing algae to be cultured, and may be adjusted to, for example, pH 6 to pH 7.
  • the mineral component can be, but is not limited to, chemically defined inorganic salts, such as alkali and alkaline earth metal salts, and salts of other metals.
  • inorganic salts include sulfates such as potassium sulfate, magnesium sulfate, iron sulfate, ammonium sulfate, or copper sulfate, nickel sulfate, and zinc sulfate; phosphates such as potassium phosphate; carbonates such as calcium carbonate; Chlorides such as cobalt chloride, manganese chloride, sodium chloride and calcium chloride; alkali metal oxides; selenites and molybdates such as sodium molybdate and sodium selenite; halogens such as potassium bromide or potassium iodide Or an artificial seawater salt such as Red @ Sea @ Salt (Red Sea Salt).
  • the salts may be added in a total amount of 0.01 to 5.0, 5 to 40, 5 to 35, 10 to 30, or 10 to 25% (g / medium L).
  • the salts are added in a total amount of 16.7 ⁇ (16.7 g / L of medium).
  • no additional mineral components eg, salts such as chlorides
  • the medium does not contain additional mineral components other than those originally contained in POME or the like.
  • the concentration of minerals in such a medium may be, for example, a total of 6.0%, 5.0%, 4.0% in terms of NaCl amount. ⁇ , 3.0 ⁇ , 2.0 ⁇ , less than 1.0 ⁇ (g / medium L), total less than 3.0 ⁇ , 2.0 ⁇ , 1.0 ⁇ (g / medium L) in terms of Cl amount Can be
  • the inclusion of saccharides in the culture along with POME may promote the growth of the heterotrophic microalgae.
  • Saccharides are carbon sources of heterotrophic microalgae, but if the POME stock solution does not contain sufficient saccharides available for microalgae, or if it is desired to promote the growth of the heterotrophic microalgae
  • a saccharide capable of utilizing microalgae in any form may be added to the culture solution.
  • saccharide examples include, but are not limited to, one or more saccharides selected from glucose, galactose, fructose, maltose, sucrose, lactose, oligosaccharides, sugar alcohols such as glycerol, and the like.
  • the saccharide may be added in a total amount of 5 to 40, 10 to 30, and 15 to 25 (g / medium L).
  • the saccharide is added in a total amount of 20% (20 g / medium L).
  • the microalgae used in the present invention may be an algae that produces ⁇ -3 polyunsaturated fatty acids such as DHA (hereinafter referred to as DHA-producing algae).
  • DHA-producing algae include organisms belonging to Thraustochytriales, such as Aurantiochytrium, Schizochytrium, Thraustochytrium, Parietichytrium, and Parietichytrium.
  • Organisms belonging to the genus Ulkenia or organisms belonging to the genus Oblongichytrium are exemplified.
  • Examples of the organisms belonging to the genus Aurantiochytrium include Aurantiochytrium limacinum, Aurantiochytrium mangrovei, and the like.
  • Examples of the organism belonging to the genus Schizochytorium include Schizochytrium aggregatum.
  • strains such as Schizochytrium sp. Maku-1 can be used.
  • Examples of the organism belonging to the genus Thraustochytrium include Thraustochytrium aureum, Thraustochytrium pachydermum, and Thraustochytrium aggregatum.
  • Organisms belonging to the genus Parietichytrium include, for example, Parietichytrium sarkarianum.
  • strains such as Parietichytrium sp. 6F-10b can be used.
  • Examples of the organism belonging to the genus Ulkenia include Ulkenia visurgensis and Ulkenia profunda. For example, strains such as Ulkenia sp. Yonez 6-9 can be used.
  • Examples of the organism belonging to the genus Oblongichytrium include Oblongichytrium multirudimentale, Oblongichytrium minutum, and the like. For example, strains such as Oblongichytrium sp. H9 can be used.
  • algae of the genus Aurantiochytrium such as Aurantiochytrium Limasinum 4W-1b, Aurantiochytrium Limasinum SR-21, and Aurantiochytrium Limasinum NIES3737, and the like.
  • Limasinum strains and Aurantiochytrium mangrovey strains such as Aurantiochytrium mangrovey strain 18W-13a can be used.
  • the cultivation of algae producing DHA or the like in the present invention is carried out by inoculating the algae in a medium containing POME adjusted as described above and culturing them according to a standard method.
  • the culture conditions depend on the type of algae producing DHA or the like to be cultured, and the temperature is 5 to 40 ° C, preferably 10 to 35 ° C, particularly preferably 20 to 25 ° C, more preferably 25 ° C ⁇ 1 ° C.
  • Culture is performed for 1 to 10 days, preferably 3 to 7 days, for example, 4 to 5 days, and the culture can be performed by aeration-agitation culture, shaking culture, or static culture.
  • the production algae such as DHA used in the present invention may be cultured in a culture device having an appropriate cell culture means.
  • Cell culturing means means means having all functions for culturing cells, for example, a culturing tank, and the culturing tank includes a stirring device, a vibration device, a temperature control device, a pH control device, It may have one or more devices selected from a degree measuring device, a light control device, a device for measuring the concentration of a specific gas such as O 2 and CO 2 and a pressure measuring device.
  • the culture tank may be the same tank as the concentration / separation tank, or may be a separate tank from the concentration / separation tank. When it is a separate tank from the concentration / separation tank, the tank may be connected by an appropriate means, for example, a flow path or the like.
  • the present invention further provides a DHA production method, wherein DHA-producing algae cultured by the above culture method using a medium containing POME are collected, and DHA is extracted from the collected DHA-producing algae.
  • the DHA production method of the present invention is characterized in that DHA or the like-producing algae is produced by culturing DHA or the like-producing algae using a medium containing POME.
  • DDHA produced by the DHA-producing algae of the present invention can be extracted and analyzed by methods known to those skilled in the art.
  • the DHA-producing algae can be recovered by an existing method such as centrifugation or filtration.
  • the recovered DHA-producing algae pellets are dried by freeze-drying or drying by heating or the like, or the DHA-producing algal culture is concentrated, and then the undried wet alga is used in the DHA extraction step. Is also good.
  • DDHA can be extracted from the obtained dried algal cells or wet algal cells.
  • the extraction method is not particularly limited, and can be performed by a known method such as extraction with an organic solvent, squeezing, or supercritical extraction.
  • organic solvent used in the organic solvent extraction method include hexane, acetone, chloroform, methanol, ethanol, diethyl ether and the like, and may be used alone or as a mixture of two or more liquid solvents such as a polar solvent and a nonpolar solvent. A liquid can also be used.
  • Algae may be crushed before extraction, but is not limited to, chemical crushing with alkali or acid, homogenizer or ultrasonic wave, mechanical crushing with a bead mill, biological crushing with enzymes, etc., performed by known methods. You may.
  • the obtained extract may be concentrated and purified by a method known to those skilled in the art, for example, column chromatography using silica gel or acidic clay, high-performance liquid chromatography, liquid-liquid distribution, urea addition method, and the like. May be concentrated and purified using a known method.
  • the DHA-producing algae may contain at least 5%, at least 15%, at least 20%, at least 25%, at least 30% (% by weight) of DHA based on its dry weight.
  • Example 1 Growth of Aurantiochytrium Limasinum 4W-1b strain, production of total lipids and DHA (seed algae) in a medium supplemented with artificial seawater adjusted to have an addition rate of POME supernatant liquid of 10 to 50% )
  • Aurantiochytrium limacinum 4W-1b strain belonging to Thraustochytriales provided by the University of Tsukuba in a GTY medium (containing 1 g of 1/2 diluted seawater with 20 g of glucose, 5 g of yeast extract, and 10 g of tryptone) in the following medium for 4 days It was cultured and used as a seed algae.
  • the high-temperature POME provided from a factory in Riau, Indonesia in April 2018 was quickly cooled in a refrigerator and stored for 3 days, and then centrifuged at 5000 rpm for 15 minutes. Liquid).
  • a medium containing 16.7 g / L of red sea salt (artificial seawater salts) was prepared so that the addition rate of the POME supernatant was 10%, 25%, and 50% (volume%).
  • FIG. 1 shows the time course of the algal body concentration of Aurantiochytrium Limasinum 4W-1b cultured in 10%, 25%, and 50% series culture media of the POME supernatant (OD660).
  • the strain grew in all addition mediums, and the higher the POME concentration (addition rate), the higher the growth of the strain.
  • FIG. 2 shows the total lipid and DHA production after 48 hours of culture. Lipid and DHA were produced in all the additive mediums, and the higher the POME concentration (addition rate), the higher the lipid and DHA production of the strain.
  • Table 1 shows the amount of growth (g / L), algal productivity (mg / L / day), lipid content (%), Table summarizing lipid amount (mg / L), lipid productivity (mg / L / day), DHA content (%), DHA production amount (mg / L), and DHA productivity (mg / L / day) Is shown.
  • the productivity is a value obtained by converting the amount of algal bodies, the amount of total lipids, and the amount of DHA after 48 hours of growth per day.
  • Each of the lipid content and the DHA content indicates a ratio to the algal body content (% by weight).
  • the algal productivity and lipid productivity were 1760 mg / L / day and 395 mg, respectively. / L / day.
  • Table 2 shows a comparison of the algal growth and productivity and the total lipid production and productivity between the prior art in which other microalgae were cultured using POME and the present invention.
  • algal cell productivity of the conventional microalgae is 2.9 to 1,040 (mg / L / day), whereas algal cell productivity of the present invention is 50% POME and no glucose added.
  • the total lipid productivity of the microalgae of the prior art is 0.63 (mg / L / day) to 100.9 (mg / L / day), whereas in the present invention, Alanthiochitrium @ Limasinum @ 4W-
  • the algal productivity when 1b was used was as high as 3.9 to 626 times even when a medium without 50% POME and glucose was used.
  • Example 2 Aurantiochi in the case of using a medium in which glucose and artificial seawater salts are added to a POME supernatant (100%) and a diluent adjusted so that the addition ratio of the POME supernatant is 10 to 75% Propagation of thorium limacinum 4W-1b strain, production of total lipids and DHA Using the same POME and seed algae as in Example 1, 20 g / L glucose and 16 g of POME supernatant (100%) obtained by the same method. A 100% POME medium was prepared by adding 7 g / L of Red Sea salt.
  • the POME supernatant was diluted with distilled water so that the addition ratio of the supernatant became 10%, 25%, 50%, and 75% (% by volume), and 16.7 g / L of red sea salt and 20 g / L of glucose were used. A medium adjusted to contain the same was also prepared. These were defined as a POME addition series. Then, in the same manner as in Example 1, the growth of Aurantiochytrium Limasinum 4W-1b was evaluated by OD, and the dry weight of the algal cells was measured to calculate the amount of algal cells per liter of the culture solution. Then, the total lipid and the amount of DHA were measured.
  • FIG. 3 shows the change over time in the algal body concentration of the algae strain when the prepared POME addition series was used as a medium (OD660).
  • OD660 a medium
  • FIG. 4 shows the relationship between the POME addition rate and the amount of proliferation.
  • FIG. 5 shows the total lipid and DHA production after 48 hours of culture.
  • lipids and DHA were produced in all the addition mediums, and the higher the addition rate of POME, the higher the production amount of lipids and DHA, indicating the highest production in the medium containing 100% POME.
  • Table 3 shows the growth amount (g / L), algal cell productivity (mg / L / day), lipid content (%), and total lipid content after the strain was grown for 48 hours in each addition rate medium of Example 2.
  • Table summarizing lipid amount (mg / L), lipid productivity (mg / L / day), DHA content (%), DHA production amount (mg / L), and DHA productivity (mg / L / day) Is shown. The method of obtaining each numerical value and the unit are the same as in Table 1.
  • Example 2 Compared to Example 1 performed without glucose, the strain showed clearly higher growth in each of the 10%, 25%, and 50% POME-added strains to which glucose was added in Example 2. As shown in Table 3, the algal cell productivity was 2,815 mg / L / day, which was twice as high as that of the 50% added series with glucose added, compared to the 50% added series with glucose added in Table 1. The lipid productivity was 2.8 times higher at 1,110 mg / L / day, and the DHA productivity was 7.7 times higher at 228.5 mg / L / day.
  • the algal cell productivity of the strain when glucose was added at 100% POME was 4,150 mg / L / day, and the lipid productivity was 1,675 mg / L / day, which was higher than that of the prior art.
  • the values were 4 times to 1430 times and 16.6 times to 2660 times higher, respectively.
  • Example 3 Time-dependent change in growth and DHA concentration of Aurantiochytrium limacinum 4W-1b strain in a medium in which glucose and artificial seawater at each concentration were added to POME supernatant (100%) Same as Examples 1 and 2 Using POME and seed algae, 20 g / L glucose and 0.00%, 0.38%, 0.75%, 1.50%, 3% were added to the POME supernatant (100%) obtained in the same manner. 0.000% (wt%) red sea salt was added.
  • the medium was cultured under the same culture conditions (500 mL Erlenmeyer flask (200 mL of medium charged), 25 ° C., reciprocal shaking culture (100 strokes / min)), and OD was measured.
  • the growth of Aurantiochytrium perimacinum 4W-1b was evaluated, and the dry weight of algal cells was measured to calculate the amount of algal cells per liter of culture solution, and the DHA concentration was measured.
  • FIG. 6 shows the time course of the algal body concentration (OD660) and the DHA concentration after growing for 48 hours.
  • the algal body concentration was not significantly affected by the concentration of the added mineral component, but the DHA concentration was higher when the added mineral component concentration was lower (0.00%, 0.38%, 0.75% added) (1. (50%, 3.00% added).
  • the concentration of the added mineral component the better the DHA production (mg / L).
  • the salt concentration of the POME supernatant (100%) before the addition of the mineral component and glucose was measured and found to be 2230 (mg / L) in terms of Cl (chloride ion). This corresponds to 12.5% by weight of seawater, which corresponds to about 0.375% by weight in terms of NaCl.
  • Cl chloride ion
  • Example 4 Culture test using various strains of Aurantiochytrium sp. POME supernatant obtained by the same method as in Examples 1 to 3 except that POME provided from a factory in North Sumatra, Indonesia was used. 20 g / L glucose and 16.7 g / L red sea salt were added to the liquid (100%) to prepare a 100% POME medium, which was then sterilized and cooled.
  • FIG. 7 shows the total lipid amount (mg / L) and the DHA concentration (mg / L) of various strains of the genus Aurantiochytrium.
  • Table 5 shows the growth amount (g / L), algal cell productivity (mg / L / day), lipid content (%), and total lipid amount (mg) after the various strains were grown for a certain period of time in Example 4.
  • / L lipid productivity
  • DHA content %
  • DHA production amount mg / L
  • DHA productivity mg / L / day
  • Example 5 Culture test using various strains of other DHA-producing algae Cultured under the same conditions as in Example 4 except for using various strains of Thraustochytriales and other DHA-producing algae as shown in Table 6 below. Was performed to measure the productivity of algal cells, total lipids, and DHA.
  • Table 7 shows the results. Table 7 shows algal productivity (mg / L / day), lipid productivity (mg / L / day), and DHA productivity (mg / L / day) after growing various strains in Example 5. Is a table summarizing. The definition of each value is the same as in Table 1. In addition, only the productivity was described because the culturing time differs depending on each strain. From these results, it is understood that DHA-producing algae such as Thraustochytriales generally show extremely excellent algal productivity, total lipid and DHA productivity when the medium of the present invention is used.

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  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

La présente invention aborde le problème consistant à fournir un procédé de culture permettant de faire proliférer efficacement des microalgues à l'aide d'un POME déchargé à partir d'un procédé de fabrication d'huile de palme et un procédé de production efficace d'un matériau de valeur ajoutée à valeur élevée à partir des microalgues. Des microalgues hétérotrophes capables de produire un acide gras insaturé polyvalent ω-3 tel que le DHA (acide docosahexaénoïque) sont cultivées à l'aide d'un POME. Ainsi, ces microalgues hétérotrophes sont efficacement proliférées et le DHA est produit à une concentration élevée.
PCT/JP2019/027981 2018-08-01 2019-07-16 Procédé de culture de microalgues hétérotrophes à l'aide d'un effluent de broyeur d'huile de palme (pome) et procédé de production de dha WO2020026794A1 (fr)

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MYPI2020001547A MY188244A (en) 2018-08-01 2019-07-16 Method for culturing heterotrophic microalgae using palm oil mill effluent (pome) and method for producing dha
JP2019571086A JP6709484B1 (ja) 2018-08-01 2019-07-16 パームオイル工場排出液（ｐｏｍｅ）をつかった従属栄養性微細藻類の培養方法及びｄｈａ製造方法

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