4-hydroxybenzoic acid composition

Abstract

To provide a 4-hydroxybenzoic acid composition with enhanced balance between environmental compatibility and color tone.SOLUTION: A 4-hydroxybenzoic acid composition includes at least 4-hydroxybenzoic acid (A) having an actually measured bioavailability of 80% to 100% inclusive, where the actually measured bioavailability is measured in accordance with ASTM D6866-20 and indicates the content ratio of radioactive carbon 14C derived from biomass resources with respect to the concentration of radioactive carbon in circulating carbon in the 1950s. The 4-hydroxybenzoic acid composition exhibits an absorbance of 0.015 or less at a wavelength of 425 nm when dissolved in ethanol at a concentration of 20 g / L.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to 4-hydroxybenzoic acid compositions. [Background technology]

[0002] Against the backdrop of global warming and the depletion of fossil fuel resources, the production of chemicals using renewable biomass resources as raw materials is attracting attention as a new industry aimed at realizing a low-carbon society.

[0003] 4-Hydroxybenzoic acid (4-HBA) is a useful chemical substance that is used as a raw material for liquid crystal polymers and as a synthetic raw material for the preservative paraben. Currently, 4-hydroxybenzoic acid is chemically produced using crude oil as a raw material. Chemical methods for producing 4-hydroxybenzoic acid include reacting phenol with potassium hydroxide and carbon dioxide under high pressure conditions.

[0004] However, these methods not only rely on fossil fuels for the raw material phenol, but also require strong alkali, carbon dioxide, and high temperature and pressure conditions in the reaction process, and generate harmful waste liquids and exhaust gases during the production process, placing a significant burden on the environment. In light of this background, there is a strong need to establish an environmentally friendly bioproduction technology for 4-hydroxybenzoic acid that saves energy and reduces hazardous wastewater.

[0005] For example, Non-Patent Documents 1 and 2 and Patent Documents 1 and 2 have revealed, using Escherichia coli, that 4-HBA is synthesized from chorismate, an intermediate in the shikimate pathway involved in the synthesis of aromatic amino acids, by chorismate-pyruvate lyase encoded by ubiC. [Prior art documents] [Patent documents]

[0006] [Non-Patent Document 1] J. Bacteriol., 174, 5309-5316 (1992) [Non-Patent Document 2] Microbiology, 140, 897-904 (1994) [Patent Document 1] US Patent No. 6030819 [Patent Document 2] U.S. Patent No. 6,114,157 Summary of the Invention [Problem to be solved by the invention]

[0007] According to the studies of the present inventors, it has become clear that there is room for improvement in terms of color tone of the above-mentioned biomass-derived 4-hydroxybenzoic acid.

[0008] The present invention has been made in view of the above circumstances, and provides a 4-hydroxybenzoic acid composition having an improved balance between environmental compatibility and color tone. [Means for solving the problem]

[0009] The present inventors have conducted extensive research to achieve the above object, and as a result have found that the balance between environmental compatibility and color tone of a 4-hydroxybenzoic acid composition can be improved by controlling the absorbance of light having a wavelength of 425 nm within a specific range, thereby completing the present invention.

[0010] [1] Radiocarbon from biomass resources determined based on the concentration of radiocarbon in circulating carbon in the 1950s, measured according to ASTM D6866-20 14 It contains at least 4-hydroxybenzoic acid (A) with a measured bio-based content rate of 80% or more and 100% or less of C, A 4-hydroxybenzoic acid composition having an absorbance of 0.015 or less at a wavelength of 425 nm when dissolved in ethanol at a concentration of 20 g / L. [2] In the 4-hydroxybenzoic acid composition described in [1] above, A 4-hydroxybenzoic acid composition, in which the content of the 4-hydroxybenzoic acid (A) is 95% by mass or more and 100% by mass or less, when the entire 4-hydroxybenzoic acid composition is taken as 100% by mass. [3] In the 4-hydroxybenzoic acid composition according to the above [1] or [2], A 4-hydroxybenzoic acid composition, in which the measured bio-content rate of the 4-hydroxybenzoic acid (A) is 95% or more and 100% or less. [4] In the 4-hydroxybenzoic acid composition according to any one of the above [1] to [3], A 4-hydroxybenzoic acid composition having an absorbance of 0.010 or less. [5] In the 4-hydroxybenzoic acid composition according to any one of the above [1] to [4], A 4-hydroxybenzoic acid composition having an absorbance of 0.001 or more. [6] In the 4-hydroxybenzoic acid composition according to any one of the above [1] to [5], The 4-hydroxybenzoic acid (A) is a 4-hydroxybenzoic acid composition containing at least a product of one or two transformants selected from transformant HBA-2 (accession number: NITE BP-01838) and transformant HBA-47 (accession number: NITE BP-01849). Effect of the Invention

[0011] According to the present invention, it is possible to provide a 4-hydroxybenzoic acid composition having an improved balance between environmental compatibility and color tone. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] Hereinafter, an embodiment of the present invention will be described in detail.

[0013] [4-Hydroxybenzoic acid composition] The 4-hydroxybenzoic acid composition of the present embodiment has a radioactive carbon content derived from biomass resources, which is calculated based on the radioactive carbon concentration in circulating carbon in the 1950s measured according to ASTM D6866-20. 14 It contains at least 4-hydroxybenzoic acid (A) with a measured bio-content rate, which indicates the proportion of C, of ​​80% or more and 100% or less, and when dissolved in ethanol at a concentration of 20 g / L, the absorbance at a wavelength of 425 nm is 0.015 or less.

[0014] In such a 4-hydroxybenzoic acid composition, the measured bio-based content indicates the amount of plant-derived carbon contained in 4-hydroxybenzoic acid (A), and the larger this value, the more the 4-hydroxybenzoic acid composition is composed of naturally derived raw materials. Therefore, the higher the measured bio-based content, the better the environmental compatibility from the viewpoint of carbon neutrality. In addition, when the absorbance is 0.015 or less, a 4-hydroxybenzoic acid composition with a more suitable color tone can be obtained even when using raw materials derived from plants.

[0015] Such a 4-hydroxybenzoic acid composition can be produced by the production method described below, particularly by (i) using raw materials derived from specific plants as raw material compounds, (ii) removing impurities by adding an adsorbent to the reaction solution, and (iii) washing with pure water upon recovery.

[0016] In the 4-hydroxybenzoic acid (A) in the 4-hydroxybenzoic acid composition of this embodiment, the radioactive carbon derived from biomass resources is determined based on the concentration of radioactive carbon in circulating carbon in the 1950s, as measured based on ASTM D6866-20. 14 The measured bio-based content rate, which indicates the proportion of C, is 80% or more and 100% or less, preferably 85% or more and 100% or less, more preferably 90% or more and 100% or less, and even more preferably 95% or more and 100% or less. If the measured bio-based content rate is within the above range, environmental compatibility will be better from the perspective of carbon neutrality.

[0017] At this time, the radioactive carbon in the total carbon atoms of 4-hydroxybenzoic acid (A) in the 4-hydroxybenzoic acid composition 14 The actual biobased ratio, which indicates the proportion of C, is calculated based on the biobased concentration test standard ASTM D6866-20 established by the American Society for Testing and Materials (ASTM). 14 The C content was measured, and the results were compared with the radioactive carbon in the circulating carbon as of 1950. 14 The concentration of C is calculated from the concentration of C. In addition, this test standard specifies several analytical methods, among which analytical method B (AMS method) is preferably used. For analytical method B, an analytical device such as an accelerator mass spectrometer Pelletron AMS manufactured by NEC Corporation is used.

[0018] In the 4-hydroxybenzoic acid composition of this embodiment, when dissolved in ethanol at a concentration of 20 g / L, the upper limit of the absorbance at a wavelength of 425 nm is 0.015 or less, preferably 0.013 or less, more preferably 0.010 or less, and even more preferably 0.006 or less. By having the absorbance at a wavelength of 425 nm be equal to or less than the upper limit, the color tone of the 4-hydroxybenzoic acid composition of this embodiment becomes more suitable. In addition, the lower limit of the absorbance at a wavelength of 425 nm when dissolved in ethanol at a concentration of 20 g / L is not particularly limited, but is, for example, 0.000 or more, and from the viewpoint of environmental compatibility and production efficiency, is preferably 0.001 or more, and may be 0.003 or more. In this case, the absorbance of 0.000 includes the case where the absorbance at a wavelength of 425 nm is below the lower detection limit and cannot be detected by quantification.

[0019] In the 4-hydroxybenzoic acid composition of this embodiment, the content of the 4-hydroxybenzoic acid (A) is preferably 95% by mass or more and 100% by mass or less, more preferably 96% by mass or more and 100% by mass or less, even more preferably 97% by mass or more and 100% by mass or less, even more preferably 98% by mass or more and 100% by mass or less, and even more preferably 99% by mass or more and 100% by mass or less, when the entire 4-hydroxybenzoic acid composition is taken as 100% by mass. By having the content of the 4-hydroxybenzoic acid (A) within the above numerical range, the environmental compatibility is improved from the viewpoint of carbon neutrality, and the color tone of the 4-hydroxybenzoic acid composition of this embodiment becomes more suitable. When the content of the 4-hydroxybenzoic acid (A) is 100% by mass, the content of the other components described below is 0.000% by mass, including the case where the content of the other components is below the detection limit and cannot be detected by quantitative analysis.

[0020] In the 4-hydroxybenzoic acid composition of this embodiment, the 4-hydroxybenzoic acid (A) preferably contains at least one or two products of transformants selected from transformant HBA-2 (accession number: NITE BP-01838) and transformant HBA-47 (accession number: NITE BP-01849). When the 4-hydroxybenzoic acid (A) contains at least the above-mentioned products, the environmental compatibility becomes better from the viewpoint of carbon neutrality, and the color tone of the 4-hydroxybenzoic acid composition of this embodiment becomes more suitable.

[0021] In the 4-hydroxybenzoic acid composition of this embodiment, the 4-hydroxybenzoic acid (A) contains at least the product of one or more transformants selected from transformant HBA-2 and transformant HBA-47, and thus the color tone is more favorable. This is presumably because the presence of trace amounts of components that may adversely affect the color tone is kept relatively low. Here, the extremely large number of components derived from the transformant are present in extremely small amounts, and the contents of the extremely small components are below the detection limit and cannot be analyzed. Therefore, in the 4-hydroxybenzoic acid composition of the present embodiment, the embodiment in which 4-hydroxybenzoic acid (A) contains at least one or more products of transformants selected from transformant HBA-2 and transformant HBA-47 is considered to be impractical and impossible to directly identify the characteristic portion of this embodiment based on the structure or properties of the product, and therefore the embodiment is considered to be impractical and impossible.

[0022] (Other Ingredients) The 4-hydroxybenzoic acid composition of the present embodiment may contain components other than those described above, provided that the balance between environmental compatibility and color tone is not impaired. The other components include one or more compounds selected from the group consisting of the following compounds: Monosaccharides such as fructose, mannose, arabinose, xylose, galactose, etc.; disaccharides such as cellobiose, sucrose, lactose, maltose, trehalose, cellobiose, xylobiose, etc.; saccharides such as dextrin or soluble starch, etc. Sugar alcohols such as mannitol, sorbitol, xylitol, and glycerin; organic acids such as acetic acid, citric acid, lactic acid, fumaric acid, maleic acid, and gluconic acid; alcohols such as ethanol and propanol; and hydrocarbons such as normal paraffin. Inorganic or organic ammonium compounds such as ammonium chloride, ammonium sulfate, ammonium nitrate, and ammonium acetate; urea, aqueous ammonia, sodium nitrate, potassium nitrate, corn steep liquor, meat extract, peptone, NZ-amine, protein hydrolysates, and nitrogen-containing organic compounds such as amino acids. Examples of such salts include monopotassium phosphate, dipotassium phosphate, magnesium sulfate, sodium chloride, ferrous sulfate, ferrous nitrate, manganese sulfate, zinc sulfate, cobalt sulfate, and calcium carbonate.

[0023] In the 4-hydroxybenzoic acid composition of the present embodiment, the content of the other components is preferably 5% by mass or less, more preferably 4% by mass or less, even more preferably 3% by mass or less, even more preferably 2% by mass or less, and even more preferably 1% by mass or less. By having the content of the other components be equal to or less than the upper limit, the color tone of the 4-hydroxybenzoic acid composition of the present embodiment becomes more suitable. The lower limit of the content of the other components is not particularly limited, but is, for example, 0.000% by mass or more. In this case, 0.000% by mass includes the case where the content of the other components is below the lower detection limit and cannot be detected by quantitative analysis. In addition, when the 4-hydroxybenzoic acid composition of the present embodiment contains a plurality of the other components described above, the total content of the other components described above may be equal to or less than the upper limit value and equal to or more than the lower limit value.

[0024] Methods for quantifying the content of such 4-hydroxybenzoic acid (A) and other components include, for example, high performance liquid chromatography using a high performance liquid chromatograph equipped with a UV detector (manufactured by Hitachi High-Tech Science Corporation, Chromaster (registered trademark)), ion chromatography, gas chromatography mass spectrometry (GC / MS), inductively coupled plasma mass spectrometry (ICP-MS), and the like.

[0025] [Method for producing 4-hydroxybenzoic acid composition] Next, a method for producing the 4-hydroxybenzoic acid composition of the present embodiment will be described.

[0026] The method for producing the 4-hydroxybenzoic acid composition of the present embodiment preferably includes the following steps (A) to (C). (A) A step of reacting the transformant with at least one raw material compound selected from the group consisting of sugars, compounds which the transformant can metabolize to produce chorismic acid, or chorismic acid and salts thereof in a reaction solution containing the transformant. (B) A step of removing impurities from the reaction solution. (C) A step of recovering the 4-hydroxybenzoic acid composition in the reaction solution.

[0027] [(A) reacting the transformant with at least one raw material compound selected from the group consisting of saccharides, compounds which the transformant can metabolize to produce chorismic acid, or chorismic acid and salts thereof in a reaction solution containing the transformant] The method for producing a 4-hydroxybenzoic acid composition of this embodiment first involves a step of reacting (A) a transformant with at least one raw material compound selected from the group consisting of sugars, compounds that the transformant can metabolize to produce chorismic acid, or chorismic acid and salts thereof, in a reaction solution containing the transformant. Each compound used in this step and the reaction conditions are described in detail below.

[0028] (Transformant) In the method for producing a 4-hydroxybenzoic acid composition of the present embodiment, any transformant can be used as long as it is capable of producing 4-hydroxybenzoic acid from sugars through metabolism. Examples of such transformants include the coryneform bacteria transformant HBA-2 (accession number: NITE BP-01838) described in Japanese Patent No. 6327653 and the coryneform bacteria transformant HBA-47 (accession number: NITE BP-01849) described in Japanese Patent No. 6327654.

[0029] Prior to the reaction, it is preferable to culture the transformant under aerobic conditions. The culture temperature and culture time are not particularly limited as long as the transformant can be cultured and a required amount or more of the transformant can be obtained. The culture temperature is, for example, 25°C or higher and 38°C or lower. The culture time is, for example, 12 hours or higher and 48 hours or lower.

[0030] (Culture medium) The medium used for aerobic cultivation of the transformant prior to the reaction may be a natural medium or a synthetic medium containing a carbon source, a nitrogen source, inorganic salts, and other nutrients. As carbon sources, sugars (monosaccharides such as glucose, fructose, mannose, xylose, arabinose, and galactose; disaccharides such as sucrose, maltose, lactose, cellobiose, xylobiose, and trehalose; polysaccharides such as starch; molasses, etc.), sugar alcohols such as mannitol, sorbitol, xylitol, and glycerin; organic acids such as acetic acid, citric acid, lactic acid, fumaric acid, maleic acid, and gluconic acid; alcohols such as ethanol and propanol; and hydrocarbons such as normal paraffin can also be used. The carbon source may be used alone or in combination of two or more. The concentration of the carbon source in the medium may vary depending on the compound used, but may be, for example, 0.1 w / v% to 20 w / v%.

[0031] As the nitrogen source, inorganic or organic ammonium compounds such as ammonium chloride, ammonium sulfate, ammonium nitrate, and ammonium acetate, urea, aqueous ammonia, sodium nitrate, and potassium nitrate can be used. In addition, nitrogen-containing organic compounds such as corn steep liquor, meat extract, peptone, NZ-amine, protein hydrolysates, and amino acids can also be used. The nitrogen source can be used alone or in combination of two or more. The concentration of the nitrogen source in the medium varies depending on the nitrogen compound used, but may be, for example, 0.1 w / v% to 10 w / v%. Examples of inorganic salts include potassium dihydrogen phosphate, potassium dihydrogen phosphate, magnesium sulfate, sodium chloride, ferrous sulfate, ferrous nitrate, manganese sulfate, zinc sulfate, cobalt sulfate, calcium carbonate, etc. The inorganic salts can be used alone or in combination of two or more. The concentration of the inorganic salts in the medium varies depending on the inorganic salts used, but may be, for example, 0.01 w / v% to 1 w / v%.

[0032] Examples of the nutritional substances include meat extract, peptone, polypeptone, yeast extract, dry yeast, corn steep liquor, skim milk powder, hydrolyzed skim soybeans with hydrochloric acid, extracts of animals, plants or microbial cells, and decomposition products thereof, etc. The concentration of the nutritional substance in the medium varies depending on the nutritional substance used, but may be, for example, 0.1 w / v% to 10 w / v%. Furthermore, vitamins can be added as necessary, such as biotin, thiamine (vitamin B1), pyridoxine (vitamin B6), pantothenic acid, inositol, and nicotinic acid. The pH of the medium is preferably 6 or more and 8 or less.

[0033] Specific preferred media include A medium [Inui, M. et al., Metabolic analysis of Corynebacterium glutamicum during lactate and succinate productions under oxygen deprivation conditions. J. Mol. Microbiol. Biotechnol. 7:182-196 (2004)], BT medium [Omumasaba, CA et al., Corynebacterium glutamicum glyceraldehyde-3-phosphate dehydrogenase isoforms with opposite, ATP-dependent regulation. J. Mol. Microbiol. Biotechnol. 8:91-103 (2004)], etc. In these media, the sugar concentration may be set within the above range.

[0034] (Raw material compound) The raw material compound is preferably a compound that can be incorporated into cells of the transformant, and more preferably a compound that is easily utilized industrially, for example, a compound that is abundant in plants. Among the above raw material compounds, the saccharides are preferably saccharides capable of producing glucose by the metabolism of the transformant, and more preferably glucose. Such saccharides include oligosaccharides and polysaccharides having glucose units. The oligosaccharides and polysaccharides include monosaccharides such as fructose, mannose, arabinose, xylose, and galactose; disaccharides such as cellobiose, sucrose, lactose, maltose, trehalose, cellobiose, and xylobiose; and polysaccharides such as dextrin or soluble starch. In addition, the use of glucose is preferable from the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition of the present embodiment. Among the above-mentioned raw material compounds, examples of compounds that can be metabolized by the transformant to produce chorismic acid include quinic acid and shikimic acid. Furthermore, for example, raw materials derived from plants can be used as raw materials containing these raw material compounds. Examples of raw materials derived from plants include molasses, inedible agricultural waste such as straw (rice straw, barley straw, wheat straw, rye straw, oat straw, etc.), bagasse, corn stover, etc., energy crops such as switchgrass, napier grass, and miscanthus, wood chips, waste paper, etc., which are saccharified with saccharifying enzymes and the like to produce saccharified liquids containing multiple sugars such as glucose. Among these, glucose, chorismic acid, quinic acid, and shikimic acid are preferred as raw material compounds, and it is preferable to use raw materials of plant origin as raw materials containing these.

[0035] (Reaction solution) The reaction liquid may be a natural or synthetic reaction liquid containing a carbon source, a nitrogen source, inorganic salts, and the like. The carbon source may be the above-described raw material compound or molasses or saccharified liquid containing the raw material compound. In addition to sugars, other carbon sources that may be used include sugar alcohols such as mannitol, sorbitol, xylitol, and glycerin; organic acids such as acetic acid, citric acid, lactic acid, fumaric acid, maleic acid, and gluconic acid; alcohols such as ethanol and propanol; and hydrocarbons such as normal paraffin. The carbon source can be used alone or in combination of two or more. The concentration of the raw material compound in the reaction solution is preferably 1 w / v% or more and 20 w / v% or less, more preferably 2 w / v% or more and 10 w / v% or less, and even more preferably 2 w / v% or more and 5 w / v% or less. The concentration of the total carbon source including the raw material compounds may be 2 w / v % or more and 5 w / v % or less.

[0036] As the nitrogen source, inorganic or organic ammonium compounds such as ammonium chloride, ammonium sulfate, ammonium nitrate, and ammonium acetate, urea, aqueous ammonia, sodium nitrate, and potassium nitrate can be used. Nitrogen-containing organic compounds such as corn steep liquor, meat extract, peptone, NZ-amine, protein hydrolysates, and amino acids can also be used. The nitrogen source can be used alone or in combination of two or more. The concentration of the nitrogen source in the reaction solution varies depending on the nitrogen compound used, but may be, for example, 0.1 w / v% to 10 w / v%. Examples of inorganic salts include potassium dihydrogen phosphate, potassium dihydrogen phosphate, magnesium sulfate, sodium chloride, ferrous sulfate, ferrous nitrate, manganese sulfate, zinc sulfate, cobalt sulfate, calcium carbonate, etc. The inorganic salts can be used alone or in combination of two or more. The concentration of the inorganic salt in the reaction solution varies depending on the inorganic salt used, but may be, for example, 0.01 w / v% to 1 w / v%.

[0037] Examples of the nutritional substances include meat extract, peptone, polypeptone, yeast extract, dry yeast, corn steep liquor, skim milk powder, hydrolyzed skim soybeans with hydrochloric acid, extracts of animals, plants or microbial cells, and decomposition products thereof, etc. The concentration of the nutritional substance in the medium varies depending on the nutritional substance used, but may be, for example, 0.1 w / v% to 10 w / v%. Furthermore, vitamins can be added as necessary, such as biotin, thiamine (vitamin B1), pyridoxine (vitamin B6), pantothenic acid, inositol, and nicotinic acid. The pH of the reaction solution is preferably 6 or more and 8 or less.

[0038] Specific preferred reaction solutions include the above-mentioned A medium and BT medium, etc. In these reaction solutions, the sugar concentration may be set within the above range.

[0039] (Reaction conditions) From the viewpoint of production efficiency of the 4-hydroxybenzoic acid composition, the reaction temperature, i.e., the survival temperature of the transformant, is preferably 20° C. or higher, more preferably 23° C. or higher, and even more preferably 25° C. or higher. From the same viewpoint, the survival temperature is preferably 50° C. or lower, more preferably 48° C. or lower, and even more preferably 47° C. or lower. From the viewpoint of production efficiency of the 4-hydroxybenzoic acid composition, the reaction time is preferably 7 days or less, more preferably 5 days or less, and even more preferably 3 days or less. The reaction time is not particularly limited as long as the reaction proceeds sufficiently, but is, for example, 1 day or more. The culture may be any of a batch system, a fed-batch system, and a continuous system, among which the batch system is preferred from the viewpoint of production efficiency of the 4-hydroxybenzoic acid composition. The reaction may be carried out under aerobic conditions or reducing conditions. The productivity of the transformant itself for the 4-hydroxybenzoic acid composition is higher under aerobic conditions. However, since the transformant grows under aerobic conditions, the raw material compounds are consumed for the growth, and the production efficiency of the 4-hydroxybenzoic acid composition is accordingly reduced. Therefore, it is preferable to carry out the reaction under aerobic conditions where the transformant does not grow. In this specification, "not growing" includes substantially no growth or almost no growth. For example, the growth of the transformant can be avoided or suppressed by using a reaction solution that is deficient or limited in one or more of compounds essential for the growth of microorganisms, such as vitamins such as biotin and thiamine, and nitrogen sources.

[0040] Moreover, under reducing conditions, the transformant does not substantially grow, and therefore the raw material compounds are not consumed for growth, thereby increasing the production efficiency of the 4-hydroxybenzoic acid composition. The reduction conditions are determined by the redox potential of the reaction solution, which is preferably from -500 mV to -200 mV, more preferably from -500 mV to -150 mV. The reduction state of the reaction solution can be easily estimated using a resazurin indicator (in a reduced state, the indicator discolors from blue to colorless), but can be measured more accurately using an oxidation-reduction potentiometer (for example, ORP Electrodes, manufactured by BROADLEY JAMES).

[0041] The reaction solution under reducing conditions can be prepared by any known method without any restrictions. For example, an aqueous solution for the reaction solution can be used instead of distilled water as the liquid medium for the reaction solution, and the aqueous solution for the reaction solution can be prepared by referring to, for example, a culture medium preparation method for obligate anaerobic microorganisms such as sulfate-reducing microorganisms (Pfennig, N. et al., (1981): The dissimilatory sulfate-reducing bacteria, In The Prokaryotes, A Handbook on Habitats Isolation and Identification of Bacteria, Ed. by Starr, MP et al., p926-940, Berlin, Springer Verlag.) or "Agricultural Chemistry Experiment Book, Vol. 3, Kyoto University Faculty of Agriculture, Agricultural Chemistry Class, 26th edition, 1990, Sangyo Tosho Publishing Co., Ltd.", and the like. The aqueous solution under the desired reducing conditions can be obtained. Specifically, an aqueous solution for a reaction solution under reducing conditions can be obtained by removing dissolved gases by heating or reducing the pressure of distilled water, etc. In this case, the aqueous solution for a reaction solution under reducing conditions can be prepared by treating the distilled water, etc. under reduced pressure of preferably 10 mmHg or less, more preferably 5 mmHg or less, and even more preferably 3 mmHg or less, preferably for 1 minute or more and 60 minutes or less, and more preferably for 5 minutes or more and 40 minutes or less, with the dissolved gases, particularly dissolved oxygen, being removed.

[0042] Furthermore, an appropriate reducing agent (eg, thioglycolic acid, ascorbic acid, cysteine ​​hydrochloride, mercaptoacetic acid, thiolacetic acid, glutathione, sodium sulfide, etc.) can be added to prepare an aqueous solution for the reaction under reducing conditions. An appropriate combination of these methods is also an effective method for adjusting an aqueous solution for a reaction liquid under reducing conditions.

[0043] When the reaction is carried out under reducing conditions, it is preferable to maintain the reaction solution under reducing conditions during the reaction. In order to maintain the reducing conditions during the reaction, it is desirable to prevent the inclusion of oxygen from outside the reaction system as much as possible. Specifically, the reaction system may be sealed with an inert gas such as nitrogen gas or carbon dioxide gas. As a method for more effectively preventing the inclusion of oxygen, it may be necessary to add a pH maintaining adjustment solution or various nutrient dissolving solutions to the reaction system in order to efficiently function the metabolic function within the cells of the aerobic bacteria of the present invention during the reaction. In such cases, it is effective to remove oxygen from the added solution in advance.

[0044] [(B) Step of removing impurities from the reaction solution] Next, (B) a step of removing impurities from the reaction solution is carried out. By culturing as described above, a 4-hydroxybenzoic acid composition is produced in the reaction solution. The 4-hydroxybenzoic acid composition can be recovered by recovering the reaction solution, but the reaction solution contains components that affect the color tone of the 4-hydroxybenzoic acid composition, such as the transformant, unreacted raw material compounds, by-products of the transformant, and components contained in the reaction solution itself. Therefore, by removing these components from the reaction solution, the color tone of the 4-hydroxybenzoic acid composition can be further improved. Components that affect the color tone of such 4-hydroxybenzoic acid compositions include various intermediate metabolic substances produced by transformants, substances contained in the aforementioned "other components" and "culture medium", and specific elements. Examples of the intermediate metabolic substances include, but are not limited to, shikimic acid, dehydroshikimic acid, quinic acid, protocatechuic acid, amino acids, etc. Examples of the specific elements include, but are not limited to, nitrogen, phosphorus, potassium, magnesium, sodium, iron, manganese, zinc, cobalt, etc.

[0045] In the present embodiment, the method for removing impurities from the reaction solution when preparing the 4-hydroxybenzoic acid composition includes both isolating the transformant and adsorbing the chemical components. By carrying out both of these steps, it is possible to remove more impurities from the 4-hydroxybenzoic acid composition, and as a result, it is possible to further improve the color tone of the 4-hydroxybenzoic acid composition.

[0046] First, a method for isolating a transformant will be described. The transformant can be separated by a known separation method, for example, collection of the supernatant after standing, membrane filtration, centrifugation, etc. These methods can be used alone or in combination of two or more.

[0047] The membrane filtration can be carried out under general filtration conditions. From the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition, the membrane pore size is preferably 0.01 μm or more, more preferably 0.03 μm or more, more preferably 0.05 μm or more, more preferably 0.07 μm or more, more preferably 0.10 μm or more, more preferably 0.15 μm or more, and more preferably 0.20 μm or more. From the same viewpoint, it is preferably 3 μm or less, more preferably 2 μm or less, and more preferably 1 μm or less. In addition, the membrane pore size can be measured by general measurement methods such as mercury porosimetry, bubble point test, and bacterial filtration method, but it is preferable to use the value obtained by the bubble point test. Examples of the material of the membrane used in membrane filtration include polymer membrane, ceramic membrane, and stainless steel membrane.

[0048] As the centrifuge used for centrifugation, a common device such as a separation plate type, a cylindrical type, a decanter type, etc. can be used. From the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition, the lower limit of the temperature during centrifugation is preferably 5°C or more, more preferably 10°C or more, and even more preferably 20°C or more. From the same viewpoint, the upper limit of the temperature during centrifugation is preferably 70°C or less, more preferably 60°C or less, and even more preferably 40°C or less.

[0049] In addition, the relative centrifugal force and time during centrifugation can be appropriately set, but the lower limit of the relative centrifugal force is preferably 3000 g or more, more preferably 5000 g or more, even more preferably 7000 g or more, even more preferably 10000 g or more, and even more preferably 15000 g or more from the viewpoint of improving the color tone of the 4-hydroxybenzoic acid composition. In addition, the upper limit of the relative centrifugal force is preferably 50000 g or less, more preferably 25000 g or less, and even more preferably 20000 g or less from the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition. Similarly, the lower limit of the time is preferably 0.2 minutes or more, more preferably 0.5 minutes or more, and even more preferably 1 minute or more, from the viewpoint of improving the color tone of the 4-hydroxybenzoic acid composition, and the upper limit of the time is preferably 75 minutes or less, more preferably 60 minutes or less, and even more preferably 30 minutes or less, from the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition.

[0050] Next, a method for adsorbing chemical components will be described. An example of a method for adsorbing chemical components is adding an adsorbent that has no activity against 4-hydroxybenzoic acid to the reaction solution after removing the transformant by the above-mentioned method. The adsorbent is not particularly limited as long as it is inactive against 4-hydroxybenzoic acid and can adsorb the components contained in the unreacted raw material compounds and the reaction solution itself. Examples of such adsorbents include inorganic adsorbents such as natural adsorbents (natural zeolite, silver zeolite, acid clay, etc.) or synthetic adsorbents (synthetic zeolite, molecular sieve, bacteria adsorption polymer, hydroxyapatite, phosphate rock, titanium silicate, silica gel, silica alumina gel, porous glass, etc.); activated carbon such as powdered activated carbon, granular activated carbon, fibrous activated carbon, block-shaped activated carbon, extruded activated carbon, molded activated carbon, synthetic granular activated carbon, and synthetic fibrous activated carbon; and organic adsorbents such as molecular adsorption resins, ion exchange resins, ion exchange fibers, chelating resins, and chelating fibers. Among these, it is preferable to use activated carbon from the viewpoint of environmental compatibility.

[0051] The amount of adsorbent added is preferably 0.3 g or more, more preferably 0.5 g or more, even more preferably 0.8 g or more, even more preferably 1.0 g or more, even more preferably 1.5 g or more, even more preferably 1.8 g or more, and even more preferably 2.0 g or more per 100 g of reaction solution, from the viewpoint of further improving the color tone of the 4-hydroxybenzoic acid composition. In addition, the upper limit of the amount of adsorbent to be added is not particularly limited, but from the viewpoint of the work efficiency of the step (C) of recovering the 4-hydroxybenzoic acid composition in the reaction solution described below, the amount is preferably 5.0 g or less, more preferably 4.0 g or less, even more preferably 3.0 g or less, even more preferably 2.5 g or less, and even more preferably 2.1 g or less per 100 g of reaction solution.

[0052] [(C) Step of recovering 4-hydroxybenzoic acid composition from the reaction solution] Then, (C) a step of recovering the 4-hydroxybenzoic acid composition in the reaction solution is carried out. By removing the impurities as described above, a reaction solution containing a 4-hydroxybenzoic acid composition with a relatively high purity can be obtained, and the 4-hydroxybenzoic acid composition can also be separated from the reaction solution by a known method. Such known methods include crystallization, membrane separation, organic solvent extraction, and various adsorption methods (using ion exchange resins, synthetic adsorbents, etc.).

[0053] Furthermore, after the 4-hydroxybenzoic acid composition is recovered, a step of washing or purifying the 4-hydroxybenzoic acid composition may be provided, if necessary.

[0054] The step of washing the 4-hydroxybenzoic acid composition can be carried out by a known washing method. Examples of known washing methods include ultrasonic washing, stirring washing, flow washing, immersion washing, etc. Among these, stirring washing is preferred. The washing solvent is preferably one that does not dissolve the 4-hydroxybenzoic acid composition but dissolves only impurities. For example, pure water is preferable as such a washing solvent. Here, pure water refers to water with a conductivity of 1 MΩ·cm or more. Water that satisfies the above conductivity has high purity and contains very few impurities such as chlorine and salts.

[0055] The amount of washing solvent in the step of washing the 4-hydroxybenzoic acid composition is, from the viewpoint of further improving the color tone of the 4-hydroxybenzoic acid composition, preferably 7 times or more, more preferably 8 times or more, even more preferably 9 times or more, and even more preferably 10 times or more, the mass of the obtained 4-hydroxybenzoic acid composition. In addition, from the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition, the amount of the washing solvent is preferably 50 times or less, more preferably 25 times or less, even more preferably 15 times or less, and even more preferably 12 times or less.

[0056] The number of washing steps in the step of washing the 4-hydroxybenzoic acid composition is preferably one or more times, more preferably two or more times, and even more preferably three or more times, from the viewpoint of further improving the color tone of the 4-hydroxybenzoic acid composition. From the viewpoint of improving the production efficiency of the 4-hydroxybenzoic acid composition, the number of washings is preferably 10 times or less, more preferably 7 times or less, even more preferably 5 times or less, and even more preferably 4 times or less.

[0057] [Application] The 4-hydroxybenzoic acid composition of the present embodiment can be used in the same applications as those of 4-hydroxybenzoic acid compositions synthesized by conventional petrochemical processes using fossil resources as raw materials. That is, the applications of the 4-hydroxybenzoic acid composition of the present embodiment include polymer raw materials such as polyester, polyester resin, polyester fiber, liquid crystal polyester, liquid crystal polyester resin, and liquid crystal polyester fiber, raw materials for synthesizing paraben, which is a preservative, dyes, pigments, adhesive resins, and plasticizers. Examples of products using the polymer using the 4-hydroxybenzoic acid composition of the present embodiment include various power cords, electrical devices, various parts, building materials, furniture, fishing tackle such as fishing lines, fishing nets, ropes, liquid crystal polymers, etc.

[0058] Although the embodiments of the present invention have been described above, these are merely examples of the present invention, and various configurations other than those described above can be adopted. Furthermore, the present invention is not limited to the above-described embodiments, and modifications and improvements within the scope of the present invention are included in the present invention.

[0059] Specifically, in this embodiment, 4-hydroxybenzoic acid is given as an example of a chemical substance made from biomass resources as a raw material, but the technical concept of the present disclosure is applicable to various substances produced using biomass resources as a raw material, including phenol. EXAMPLES

[0060] The embodiments of the present invention will be described in detail based on Examples and Comparative Examples. It should be noted that the present invention is not limited to the Examples.

[0061] <Examples and Comparative Examples> Example 1 A test tube was prepared by dissolving 10 ml of A liquid medium (2 g of (NH2)2CO, 7 g of (NH4)2SO4, 0.5 g of KH2PO4, 0.5 g of K2HPO4, 0.5 g of MgSO4·7H2O, 1 ml of 0.06% (w / v) FeSO4·7H2O + 0.042% (w / v) MnSO4·2H2O, 1 ml of 0.02% (w / v) biotin solution, 2 ml of 0.01% (w / v) thiamin solution, 2 g of yeast extract, 7 g of vitamin assay casamino acid, and 40 g of glucose in 1 L of distilled water) containing 50 μg / ml of kanamycin, and further containing 2% by mass of calcium carbonate. A loopful of 4-hydroxybenzoic acid (BP-01849) was inoculated into the culture and cultured aerobically at 33°C for 24 hours with shaking at 200 rpm to obtain a test solution containing 4-hydroxybenzoic acid synthesized from glucose (4-hydroxybenzoic acid concentration: 7.3% by mass). The test liquid was subjected to vacuum filtration using a membrane filter (Merck Millipore, Steritop 0.22 μm×33 mm) with a pore size of 0.22 μm, to separate the HBA-47 from the test liquid. Next, powdered activated carbon was added to the test liquid, and the activated carbon treatment was performed. Note that 2.0 g of powdered activated carbon was used per 100 g of the test liquid. Next, 25% sulfuric acid was added to the test liquid that had been subjected to the activated carbon treatment, and the pH was lowered from 8 to 3, thereby performing a crystallization operation to precipitate the solute. Next, the precipitated solute was separated by filtration, and the solid precipitate was collected. Next, the obtained precipitate was washed three times with pure water in an amount 10 times the mass of the precipitate, stirred at 200 rpm using a stirrer, and then dried to obtain a solid 4-hydroxybenzoic acid composition.

[0062] Example 2 A solid 4-hydroxybenzoic acid composition was obtained in the same manner as in Example 1, except that the amount of powdered activated carbon added to the test liquid was changed to 1.0 g.

[0063] Example 3 A solid 4-hydroxybenzoic acid composition was obtained in the same manner as in Example 1, except that the amount of powdered activated carbon added to the test liquid was changed to 0.4 g.

[0064] Comparative Example 1 A solid 4-hydroxybenzoic acid composition was obtained in the same manner as in Example 1, except that the amount of powdered activated carbon added to the test liquid was changed to 0.2 g.

[0065] Comparative Example 2 A solid 4-hydroxybenzoic acid composition was obtained in the same manner as in Example 1, except that the amount of powdered activated carbon added to the test liquid was changed to 0.4 g, and the number of times that the obtained precipitate was stirred and washed with pure water in an amount 10 times the mass of the precipitate was changed to two times.

[0066] Comparative Example 3 Commercially available 4-hydroxybenzoic acid (Tokyo Chemical Industry Co., Ltd., product code: H0207) was prepared.

[0067] (Physical property evaluation) The physical properties of the 4-hydroxybenzoic acid compositions obtained in each of the Examples and Comparative Examples were evaluated by the following methods.

[0068] (concentration analysis) The amount of 4-hydroxybenzoic acid in the 4-hydroxybenzoic acid compositions of each Example and Comparative Example was determined by high performance liquid chromatography using a high performance liquid chromatograph equipped with a UV detector (manufactured by Hitachi High-Tech Science Corporation, Chromaster (registered trademark)) under the following <analysis conditions>. The results are shown in Table 1. <Analysis conditions> Column: COSMOSIL 5C18-AR-II (φ4.6mm×250mm) manufactured by Nacalai Tesque Mobile phase: Water / methanol / perchloric acid = 4 / 1 / 0.0075 (vol / vol / vol) isocratic elution Flow rate: 1mL / mmin Column temperature: 40℃ Detection method: Photodiode array (PDA) detector (210 nm)

[0069] (Actual bio-based rate) The 4-hydroxybenzoic acid (A) in the 4-hydroxybenzoic acid composition of each Example and Comparative Example was subjected to the radioactive carbon concentration test based on the biobased concentration test standard ASTM D6866-20 established by the American Society for Testing and Materials (ASTM). 14 The C content was measured. The measurement results and the radioactive carbon in the circulating carbon in 1950 were then compared. 14 From the concentration of C, the radioactive carbon in the circulating carbon of 4-hydroxybenzoic acid (A) in the 4-hydroxybenzoic acid compositions of each Example and Comparative Example was calculated. 14 Radioactive carbon in the total carbon atoms when the C concentration is taken as 100% 14 The C concentration was calculated as the actual bio-based rate and evaluated as an index of environmental compatibility. The calculation results are shown in Table 1.

[0070] (absorbance) The 4-hydroxybenzoic acid composition of each Example and Comparative Example was dissolved in ethanol to a concentration of 20 g / L to prepare a measurement sample. The absorbance of this measurement sample at a wavelength of 425 nm was measured using a UV-visible spectrophotometer (Shimadzu Corporation, UV-2400PC) with pure water as a blank. The results are shown in Table 1.

[0071] (color tone) A 1L reactor equipped with a stirrer and a nitrogen inlet tube was charged with 241.7 g (4-hydroxybenzoic acid content: 1.75 mol) of the 4-hydroxybenzoic acid composition of each Example and Comparative Example. After 1L of nitrogen was introduced into this reactor, the temperature was raised to 240°C and heated under a nitrogen atmosphere for 6 hours. After heating, the 4-hydroxybenzoic acid composition in the reactor was dissolved in ethanol to a concentration of 20% by mass, and the color of the solution was quantified based on the iodine color number measured in accordance with DIN6162 and evaluated according to the following criteria. The evaluation results are shown in Table 1. A: Iodine color number is 1 to 10 B: Iodine color number is more than 10 and less than 20 C: Iodine color number exceeds 20

[0072] [Table 1]

[0073] As shown in Table 1, by setting the absorbance of the 4-hydroxybenzoic acid composition to 0.015 or less, the balance between environmental compatibility and color tone was improved.

[0074] Corynebacterium glutamicum HBA-2 was deposited at the Patent Microorganisms Depositary of the National Institute of Technology and Evaluation, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan (postal code 292-0818) (domestic deposit date: March 27, 2014, international deposit date under the Budapest Treaty: February 23, 2015, deposit number: NITE BP-01838).

[0075] Corynebacterium glutamicum HBA-47 was deposited at the National Institute of Technology and Evaluation, Patent Microorganisms Depositary, 2-5-8 Kazusa Kamatari, Kisarazu, Chiba, Japan (postal code 292-0818) (date of international deposit under the Budapest Treaty: April 25, 2014, deposit number: NITE BP-01849). These strains have been deposited internationally under the terms of the Budapest Treaty and are publicly available under the conditions set forth in 37 CFR 1.808.

Claims

1. Radiocarbon derived from biomass resources, determined based on the concentration of radiocarbon in circulating carbon in the 1950s, measured according to ASTM D6866-20 １４ The present invention comprises at least 4-hydroxybenzoic acid (A) having a measured bio-based content rate of 80% or more and 100% or less, which indicates the proportion of C; A 4-hydroxybenzoic acid composition having an absorbance of 0.015 or less at a wavelength of 425 nm when dissolved in ethanol at a concentration of 20 g / L.

2. The 4-hydroxybenzoic acid composition according to claim 1, The 4-hydroxybenzoic acid composition has a content of the 4-hydroxybenzoic acid (A) of 95% by mass or more and 100% by mass or less, when the entire 4-hydroxybenzoic acid composition is taken as 100% by mass.

3. The 4-hydroxybenzoic acid composition according to claim 1 or 2, The 4-hydroxybenzoic acid composition, wherein the measured bio-content rate of the 4-hydroxybenzoic acid (A) is 95% or more and 100% or less.

4. The 4-hydroxybenzoic acid composition according to claim 1 or 2, The 4-hydroxybenzoic acid composition has an absorbance of 0.010 or less.

5. The 4-hydroxybenzoic acid composition according to claim 1 or 2, The 4-hydroxybenzoic acid composition has an absorbance of 0.001 or more.

6. The 4-hydroxybenzoic acid composition according to claim 1 or 2, The 4-hydroxybenzoic acid (A) is a 4-hydroxybenzoic acid composition comprising at least a product of one or two transformants selected from the transformant HBA-2 (accession number: NITE BP-01838) and the transformant HBA-47 (accession number: NITE BP-01849).

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