WO2024210137A1 - Method for purifying 5,6-dihydroxyindole - Google Patents

Abstract

This method for purifying 5,6-dihydroxyindole comprises: a preparation step for preparing a raw material liquid comprising crude 5,6-dihydroxyindole and water; a first extraction step for adding an extractant to the raw material liquid to thereby obtain a first extract containing the 5,6-dihydroxyindole and a first raffinate; a simple purification step for simply purifying the 5,6-dihydroxyindole contained in the first extract; and a first solid-liquid separation step for recovering a first separated liquid containing the purified 5,6-dihydroxyindole, by solid-liquid separation. By the method, 5,6-dihydroxyindole can be purified to a high purity.

Description

Method for purifying 5,6-dihydroxyindole

The present invention relates to a method for purifying 5,6-dihydroxyindole.

5,6-Dihydroxyindole is used as a raw material for various applications, such as cosmetics, pharmaceuticals, and agricultural chemicals. For these applications, the raw material must be thoroughly purified in advance, as impurities in the raw material can affect the application.

For example, Patent Document 1 discloses that a melanin precursor-containing solution containing 5,6-dihydroxyindole, produced by adding a microbial suspension, is subjected to a solid-liquid separation process for removing microorganisms, and a deproteinization process using an ultrafiltration membrane. Through these processes, a melanin precursor solution from which microorganisms and proteins have been removed is obtained.

JP 2011-046658 A

The purification method described in Patent Document 1 can remove easily separable impurities such as microorganisms and proteins. However, other impurities are dissolved in the melanin precursor solution. The challenge is to remove the impurities dissolved in such a solution and increase the purity of 5,6-dihydroxyindole.

The object of the present invention is to provide a method for purifying 5,6-dihydroxyindole that can purify 5,6-dihydroxyindole to a high purity.

Such an object can be achieved by the present invention described in (1) to (19) below.
(1) a preparation step of preparing a raw material liquid containing unpurified 5,6-dihydroxyindole and water;
a first extraction step of adding an extractant to the raw material liquid to obtain a first extract containing the 5,6-dihydroxyindole and a first raffinate;
A simplified purification step for simply purifying the 5,6-dihydroxyindole contained in the first extract;
a first solid-liquid separation step of recovering a first separated liquid containing the purified 5,6-dihydroxyindole by solid-liquid separation treatment;
A method for purifying 5,6-dihydroxyindole, comprising the steps of:

(2) The method for purifying 5,6-dihydroxyindole described in (1) above, in which the simplified purification step includes a step of adding a poor solvent to the first extract, which supersaturates impurities without supersaturating the 5,6-dihydroxyindole, to cause precipitation.

(3) A method for purifying 5,6-dihydroxyindole as described in (2) above, which includes a first drying step of drying the first separated liquid recovered in the first solid-liquid separation step and recovering a first dried product containing the purified 5,6-dihydroxyindole.

(4) a second extraction step of adding water to the first dried product to obtain a second extract;
a second solid-liquid separation step of recovering a second separated liquid containing the purified 5,6-dihydroxyindole by solid-liquid separation treatment;
The method for purifying 5,6-dihydroxyindole according to the above (3).

(5) The method for purifying 5,6-dihydroxyindole described in (4) above, which includes a second drying step of drying the second separated liquid recovered in the second solid-liquid separation step and recovering a second dried product containing the purified 5,6-dihydroxyindole.

(6) The method for purifying 5,6-dihydroxyindole described in (4) above, which includes a concentration adjustment step of adjusting the concentration of the second separated liquid recovered in the second solid-liquid separation step and recovering an aqueous solution containing the purified 5,6-dihydroxyindole.

(7) A method for purifying 5,6-dihydroxyindole as described in (2) above, comprising a concentration step for concentrating the first extract, which is provided between the first extraction step and the poor solvent addition step.

(8) a water addition step of adding water to the first separated liquid recovered in the first solid-liquid separation step to obtain a mixed liquid containing the extractant, the poor solvent, and the water as a solvent;
and a solvent replacement step of removing the extractant and the poor solvent from the mixed solution to recover an aqueous solution containing the purified 5,6-dihydroxyindole.

(9) The method for purifying 5,6-dihydroxyindole described in (8) above, which includes a concentration adjustment step of adjusting the concentration of the aqueous solution recovered in the solvent replacement step and recovering the aqueous solution containing the purified 5,6-dihydroxyindole.

(10) A method for purifying 5,6-dihydroxyindole as described in (8) above, which includes a concentration step in which the first separated liquid recovered in the first solid-liquid separation step is concentrated in advance.

(11) The method for purifying 5,6-dihydroxyindole described in (1) above, wherein the simplified purification process includes a first drying process in which the first extract is dried to obtain a first dried product, and a second extraction process in which water is added to the first dried product to obtain a second extract.

(12) The method for purifying 5,6-dihydroxyindole described in (11) above, which includes a second drying step of drying the first separated liquid recovered in the first solid-liquid separation step and recovering a second dried product containing the purified 5,6-dihydroxyindole.

(13) The method for purifying 5,6-dihydroxyindole described in (11) above, which includes a concentration adjustment step of adjusting the concentration of the first separated liquid recovered in the first solid-liquid separation step and recovering an aqueous solution containing the purified 5,6-dihydroxyindole.

(14) The method for purifying 5,6-dihydroxyindole described in (1) above, wherein the simplified purification step includes a water addition step of adding water to the first extract obtained in the first extraction step to obtain a mixed solution containing the extractant and the water as a solvent, and a solvent replacement step of removing the extractant from the mixed solution to recover an aqueous solution containing the purified 5,6-dihydroxyindole.

(15) A method for purifying 5,6-dihydroxyindole according to (14) above, comprising a concentration adjustment step of adjusting the concentration of the aqueous solution recovered in the solvent replacement step and recovering the aqueous solution containing the purified 5,6-dihydroxyindole.

(16) A method for purifying 5,6-dihydroxyindole as described in (14) above, which includes a concentration step in which the first extract obtained in the first extraction step is concentrated in advance.

(17) A method for purifying 5,6-dihydroxyindole according to any one of (1) to (16) above, wherein the raw material liquid contains a fermentation liquid produced by fermenting sugar.

(18) A method for purifying 5,6-dihydroxyindole according to any one of (1) to (17) above, in which the raw material liquid contains the 5,6-dihydroxyindole before purification that has been converted from plant-derived L-tyrosine.

(19) A method for purifying 5,6-dihydroxyindole according to any one of (1) to (18) above, in which the 5,6-dihydroxyindole is purified in an oxygen-free environment.

According to the present invention, 5,6-dihydroxyindole can be purified to a high purity.

FIG. 1 is a process diagram for explaining the method for purifying 5,6-dihydroxyindole according to the first embodiment. FIG. 2 is a process diagram for explaining the method for purifying 5,6-dihydroxyindole according to the second embodiment. FIG. 3 is a process diagram for explaining a method for purifying 5,6-dihydroxyindole according to the third embodiment. FIG. 4 is a process diagram for explaining the method for purifying 5,6-dihydroxyindole according to the fourth embodiment.

The 5,6-dihydroxyindole purification method of the present invention will be described in detail below based on the preferred embodiment shown in the attached drawings.

FIG. 1 is a process diagram for explaining the method for purifying 5,6-dihydroxyindole according to the first embodiment.

In the method for purifying 5,6-dihydroxyindole according to this embodiment, unpurified 5,6-dihydroxyindole is purified and high-purity 5,6-dihydroxyindole is recovered. In other words, the method for purifying 5,6-dihydroxyindole according to this embodiment is a method for producing purified 5,6-dihydroxyindole. 5,6-dihydroxyindole is used as a raw material for various applications, such as cosmetics, pharmaceuticals, and agricultural chemicals. To realize these applications, it is essential to highly purify the raw material 5,6-dihydroxyindole.

The purification method of 5,6-dihydroxyindole shown in FIG. 1 includes a preparation step S102, a liquid-liquid extraction step S104 (first extraction step), a concentration step S106, a poor solvent addition step S108, a supernatant separation step S110 (first solid-liquid separation step), a drying step S112 (first drying step), a water extraction step S114 (second extraction step), a supernatant separation step S116 (second solid-liquid separation step), and a drying or concentration adjustment step S118 (second drying step or concentration adjustment step). In this embodiment, the poor solvent addition step S108 is a simplified purification step. Each step will be explained in turn below.

1. Preparation Step In the preparation step S102, first, a raw material liquid is prepared. The raw material liquid contains 5,6-dihydroxyindole before purification and water. Hereinafter, 5,6-dihydroxyindole before purification is referred to as "unpurified DHI". DHI refers to 5,6-dihydroxyindole.

Unrefined DHI may be a compound derived from fossil resources, but it may also be a compound derived from biomass. Biomass refers to organic resources derived from plants. Specifically, examples of biomass include resources that have been converted into sugars such as starch, glucose, and cellulose and stored, the bodies of animals that grow by eating plants, and products made by processing plants and animals. By using a compound derived from biomass as unrefined DHI, the DHI after purification is not derived from fossil resources. This can contribute to suppressing the increase of carbon dioxide in the atmosphere. Therefore, the preparation process S102 may be a process of obtaining a raw material liquid from any source, but may also be a process of crudely producing unrefined DHI from these biomasses. For crude production of unrefined DHI, for example, a fermentation production process using microorganisms, a process of conversion from hydrolysates of vegetable proteins, etc. are preferably used.

Among these, it is preferable to use a fermentation liquid as the raw material liquid. This makes it possible to increase the yield of crude DHI while suppressing the increase in atmospheric carbon dioxide during the crude production of crude DHI. In addition, it is preferable for the starting material to be fermented to be sugar. Sugar, particularly monosaccharides, is easily obtained since it is abundant in nature.

On the other hand, the raw material liquid may be a liquid containing crude DHI converted from plant-derived L-tyrosine. L-tyrosine is converted to DHI by the pathway described in JP 2006-158304 A. L-tyrosine is produced by fermentation of sugar as a raw material, hydrolysis of vegetable protein, etc. In other words, L-tyrosine can be produced from biomass. Therefore, the method of converting crude DHI from plant-derived L-tyrosine can also increase the yield of crude DHI while suppressing the increase in carbon dioxide in the atmosphere.

The concentration of unpurified DHI in the raw material solution is not particularly limited, but is preferably greater than 0% by mass and less than 10.0% by mass, more preferably greater than 0.05% by mass and less than 5.0% by mass, and even more preferably greater than 0.10% by mass and less than 3.0% by mass. This embodiment is capable of purifying compounds with excellent yields even when using such a relatively low concentration raw material solution.

Furthermore, the water content in the raw material liquid is not particularly limited, but is preferably 80% by mass or more, and more preferably 90% by mass or more. This embodiment is useful from the viewpoint of reducing the number of steps, since even when the raw material liquid contains water at such a ratio, the raw material liquid can be used without concentrating it. Furthermore, when the raw material liquid is a fermentation liquid, it generally contains water at the above-mentioned content ratio. This embodiment is easy to use from that viewpoint as well, since such a fermentation liquid can be applied to the raw material liquid.

In the liquid-liquid extraction step S104 (first extraction step), the raw material liquid is brought into contact with an extractant in an extraction tank containing the raw material liquid, and a liquid-liquid extraction process is performed to obtain a first extract. The liquid-liquid extraction process is a process for selectively extracting DHI contained in the raw material liquid by utilizing the difference in solubility in the extractant.

When an extractant is added to an extraction tank containing the raw material liquid, phase separation occurs into an upper layer of a first extract liquid and a lower layer of a first raffinate liquid. Through liquid-liquid extraction processing, DHI migrates to the first extract liquid and impurities migrate to the first raffinate liquid. Thereafter, for example, the lower layer of the first raffinate liquid can be discharged from the bottom of the extraction tank to recover the remaining first extract liquid. Note that the upper layer of the first extract liquid may be selectively recovered. Furthermore, the raw material liquid may be added to an extraction tank containing the extractant, or both the raw material liquid and the extractant may be added to the extraction tank at the same time.

The extractant used in the liquid-liquid extraction step S104 is not particularly limited as long as it is an organic solvent capable of extracting DHI. As the extractant, an organic solvent with a log P (octanol/water partition coefficient) of 0 or more is preferably used, more preferably an organic solvent with a log P of 0 to 1.5, and even more preferably an organic solvent with a log P of 0 to 1.0. This reduces the miscibility of the extractant with water. This makes it possible to further increase the separation between water and the extractant in the liquid-liquid extraction process. As a result, even when the concentration of DHI in the raw material liquid is low, the yield and purity of DHI can be increased, and the operability of the liquid-liquid extraction process can be improved.

Specific methods for measuring logP include, for example, the flask shaking method described in JIS Z 7260-107:2000.

In addition, instead of the measurement methods described above, logP can also be estimated by computational chemistry methods or empirical methods. Estimation by computational chemistry methods can be performed using computer software based on the structural formula of the compound. An example of this software is ChemDraw Ultra ver. 10.0 manufactured by PerkinElmer.

The extractant used is preferably an organic solvent whose Hansen solubility parameter distance dHSP with DHI is 17 or less, and more preferably an organic solvent whose Hansen solubility parameter distance dHSP with DHI is 14 or less.

The Hansen solubility parameter distance dHSP is one of the indices for judging the solubility of multiple compounds, i.e., the strength of the affinity between multiple compounds. For example, when the distance dHSP between the Hansen solubility parameters HSP of two types of compounds is small, the solubility between these compounds tends to be high. Therefore, by using an organic solvent whose Hansen solubility parameter distance dHSP with DHI is within the above range as an extractant, the extractability of DHI can be further improved. As a result, the yield and purity of DHI can be increased even when the concentration of DHI in the raw material liquid is low.

The definition and calculation method of the Hansen Solubility Parameter (HSP) are described in "Hansen Solubility Parameters: A User's Handbook" by Charles M. Hansen (CRC Press, 2007).

The Hansen solubility parameter HSP is divided into a dispersion force term (δD) corresponding to van der Waals interactions, a polar term (δP) resulting from the attractive force generated by the dipole moment, and a hydrogen bond term (δH) resulting from hydrogen bonds generated by active hydrogen and lone pairs. For example, the Hansen solubility parameter distance dHSP of two types of compounds a and b can be calculated using the following formula (1) based on the solubility parameters δDa, δPa, and δHa of compound a and the solubility parameters δDb, δPb, and δHb of compound b.

dHSP={4×(δDa-δDb) ² + (δPa-δPb) ² + (δHa-δHb) ² } ^1/2 ...(1)

This calculation can be performed using computer software based on the structural formula of the compound. An example of this software is HSPiP (Hansen Solubility Parameters in Practice) 4th Edition (4.1.07).

Examples of extractants include ethyl acetate, diethyl ether, and methyl isobutyl ketone, and one or a mixture of two or more of these is used.

The logP of ethyl acetate is 0.73, and the Hansen solubility parameter distance dHSP to DHI is 13.5.

The logP of diethyl ether is 0.89, and the Hansen solubility parameter distance dHSP to DHI is 15.2.

The logP of methyl isobutyl ketone is 1.4, and the Hansen solubility parameter distance dHSP to DHI is 16.4.

The boiling point of the extractant is preferably 100°C or less, and more preferably 90°C or less. This makes it possible to reduce the energy input for removing the extractant in the concentration step S106 (drying step S1061 in the second embodiment) described below.

The amount of extractant mixed into the raw liquid is preferably 20-400% by volume, more preferably 30-200% by volume, and even more preferably 50-100% by volume. This increases the extraction efficiency of DHI and prevents the generation of an excessive amount of extractant.

The liquid-liquid extraction step S104 may also include an extractant recovery operation in which the extractant is separated and recovered from the first raffinate. The separated extractant may be reused in the liquid-liquid extraction process. This can reduce the amount of extractant discarded, and can reduce running costs. For example, an oil-water separator using porous fibers is used for the extractant recovery operation.

3. Concentration Step In the concentration step S106, the first extract obtained in the liquid-liquid extraction step S104 is subjected to a concentration process. The concentration process is a process of removing the solvent contained in the first extract and concentrating it, for example, a process of evaporating the solvent. By performing the concentration process, the amount of the first extract can be reduced. This makes it easier for impurities to precipitate in the poor solvent addition step S108 described below. As a result, the purity of DHI can be ultimately increased. Specific operations of the concentration process include, for example, a heating operation, a decompression operation, a gas spraying operation, etc., and one or a combination of these operations is used.

Among these, the heating operation is preferably used. The temperature of the first extract during the heating operation is not particularly limited as long as it is a temperature at which the solvent (extractant) can be distilled off and is below the boiling point of DHI. The temperature of the first extract during the heating operation is preferably 40 to 90°C, more preferably 50 to 85°C, and even more preferably 60 to 80°C. This can increase the rate at which the solvent is distilled off, increasing the purification efficiency, and suppressing thermal decomposition and volatilization of DHI.

The pressure during the heating operation is not particularly limited and may be atmospheric pressure, but is preferably less than atmospheric pressure (101 kPa), more preferably 90 kPa or less, and even more preferably 80 kPa or less. This allows concentration at a relatively low temperature, making it easier to suppress oxidative polymerization of DHI. On the other hand, a lower limit for the pressure does not need to be set, but considering the cost of reducing pressure, etc., it is preferable for the pressure to be 20 kPa or more.

The heating time is not particularly limited, but is preferably 1 to 120 minutes, and more preferably 5 to 90 minutes.

The extractant distilled off in the concentration process may be reused in the liquid-liquid extraction process S104. This can reduce the cost of refining. The concentration process S106 may be provided as needed, and may be omitted.

4. Poor Solvent Addition Step In the poor solvent addition step S108, a poor solvent is added to the concentrated first extract. This causes precipitation in the first extract. The poor solvent is a solvent that supersaturates impurities without supersaturating DHI. Although the operation of adding a poor solvent is a simple operation, it is an operation that has good impurity separation properties by appropriately selecting a poor solvent. Therefore, by performing the poor solvent addition step S108, it is possible to efficiently produce DHI with high purity (for example, a purity of 80% or more, preferably a purity of 90% or more) at a high yield.

Examples of poor solvents include alkane solvents such as hexane, heptane, cyclohexane, and decalin; benzene solvents such as benzene, toluene, xylene, cyclobenzene, and dichlorobenzene; haloalkanes such as dichloromethane, chloroform, carbon tetrachloride, and 1,2-dichloroethane; ethers such as diethyl ether, methyl tertiary butyl ether (MTBE), diisopropyl ether, and diphenyl ether; and cyclic ethers such as 1,4-dioxane and tetrahydrofuran (THF). One or a mixture of two or more of these can be used.

Among these, alkane-based solvents are preferably used, and hexane is more preferably used. These have particularly low polarity, so they dissolve DHI and prevent it from becoming supersaturated, while acting to supersaturate and precipitate impurities, particularly those made of highly polar substances. This allows the impurities to be precipitated and removed with high separability, ultimately producing a refined DHI with a higher purity.

Furthermore, by taking into account the Snyder polarity parameter of the poor solvent, it becomes possible to supersaturate the impurities and precipitate them more accurately. The Snyder polarity parameter is a parameter defined by Snyder to quantitatively express the polarity of an organic solvent (L.R. Snyder, Journal of Chromatography A, Vol. 92, p. 223-230, 1974). For the Snyder polarity parameter for organic solvents, refer to the values described in, for example, Wako Analytical Circle No. 11, "Chromatography Q&A," which is a publicly known document, or http://www.sanderkok.com/techniques/elutropic_series_extended.html.

The Snyder polarity parameter of the poor solvent is preferably 4.3 or less, more preferably 4.0 or less, even more preferably 3.0 or less, and particularly preferably -0.3 or more and 2.5 or less. This allows impurities to be precipitated more accurately, and a more highly purified DHI can be produced. The Snyder polarity parameter of the poor solvent may be below the lower limit, but this may make it difficult to obtain the poor solvent. On the other hand, if the Snyder polarity parameter of the poor solvent exceeds the upper limit, the impurity removal rate may decrease.

The poor solvent is preferably a solvent that is more hydrophobic than the extractant. This allows impurities that dissolve in the extractant but are poorly soluble in the poor solvent to be effectively precipitated and separated. Hydrophobicity can be quantified by the aforementioned log P (octanol/water partition coefficient). Therefore, it is preferable that the log P of the poor solvent is higher than the log P of the extractant. This allows the poor solvent to be more hydrophobic than the extractant, and impurities mixed in the first extraction liquid can be more effectively separated.

The difference between the log P of the poor solvent and the log P of the extractant is not particularly limited, but is preferably 1 or more, more preferably 2 to 7, and even more preferably 3 to 5. This allows impurities contained in the first extract to be separated more effectively.

By adding the poor solvent, DHI in the first extract remains dissolved, while the impurities precipitate. Therefore, DHI and impurities can be accurately separated from each other based on the difference in solubility in the poor solvent. Furthermore, by going through the concentration step S106 described above, the amount of liquid phase in the first extract used in this step can be reduced. This also reduces the amount of poor solvent to be added in this step, and reduces the amount of impurities dissolved in the liquid phase. As a result, the purity of the purified DHI finally obtained can be increased.

Examples of impurities include organic acids and inorganic salts. These impurities have many highly polar functional groups and therefore have low solubility in poor solvents such as those described above. For this reason, these impurities are separated with particularly high separation properties in the poor solvent addition step S108.

The amount of poor solvent added to the first extract is preferably 20% by volume or more, and more preferably 50% by volume or more, relative to the first extract. The upper limit does not need to be set, but is preferably 1000% by volume or less, and more preferably 500% by volume or less. This makes it possible to sufficiently increase the extraction efficiency of DHI and to prevent the use of excessive poor solvent.

After the addition of the poor solvent, it is preferable to ensure a time for the poor solvent to act on the first extract before proceeding to the next step. This time is preferably 10 minutes or more, more preferably 20 minutes or more. The upper limit does not need to be set, but is preferably 120 minutes or less, more preferably 60 minutes or less. This makes it possible to sufficiently increase the precipitation efficiency of impurities and prevent the purification time from becoming excessively long.
Furthermore, after the poor solvent is added to obtain a mixture, the mixture may be stirred as necessary, which can promote the action of the poor solvent on the first extraction liquid.

5. Supernatant Separation Step In the supernatant separation step S110 (first solid-liquid separation step), a solid-liquid separation process is performed on the first extract in which a precipitate has occurred (first extract containing a precipitate). The solid-liquid separation process is a process for separating the supernatant and the precipitate from the first extract containing the precipitate. The separated supernatant is the first separated liquid. Examples of such solid-liquid separation processes include filtration separation, supernatant separation, reduced pressure dehydration, pressurized dehydration, and centrifugation, and the like. In this embodiment, supernatant separation is used. Supernatant separation is useful in that the operation is simple and the process can be performed efficiently. In addition, when separating the supernatant, other operations such as centrifugation may be used in combination.

The first separated liquid contains purified DHI at a high purity. Therefore, the first separated liquid may be recovered and used as a purified DHI product. In the following description, purified DHI is also referred to as "purified DHI." In this embodiment, the steps described below are carried out to further increase the purity of the purified DHI and optimize the product form.

6. Drying Step In the drying step S112 (first drying step), a drying process is performed on the first separated liquid. The drying process is a process of removing the solvent from the first separated liquid and drying the solute. The powder obtained by this process is called the first dried product. Examples of the drying process include processes involving heating, decompression, and gas blowing. By performing such a drying process, a powder of purified DHI can be obtained. This powder may be commercialized, but in this embodiment, the steps described below are performed to further increase the purity of the purified DHI and optimize the product form. In addition, by performing the drying process, the first separated liquid described above can be made into a form suitable for the water extraction step S114 described below, that is, a form in which DHI can be extracted with water at a high yield.

Among these, the heating operation is preferably used. The temperature of the first separated liquid during the heating operation is not particularly limited as long as it is a temperature at which the solvent can be distilled off and is below the boiling point of DHI. The temperature of the first separated liquid during the heating operation is preferably 40 to 90°C, more preferably 50 to 85°C, and even more preferably 60 to 80°C. This can increase the rate at which the solvent is distilled off, improving purification efficiency, and suppressing thermal decomposition and volatilization of DHI.

The pressure during the heating operation is not particularly limited and may be atmospheric pressure, but is preferably less than atmospheric pressure (101 kPa), more preferably 90 kPa or less, and even more preferably 80 kPa or less. This allows drying at a relatively low temperature, making it easier to suppress oxidative polymerization of DHI. On the other hand, a lower limit for the pressure does not need to be set, but considering the cost of reducing pressure, it is preferable for it to be 20 kPa or more.

The heating time is not particularly limited, but is preferably 1 to 120 minutes, and more preferably 5 to 90 minutes.

In the drying step S112, it is not necessary to remove all of the solvent contained in the first separation liquid. Specifically, the amount of solvent remaining in the first dried product is preferably 30% by mass or less of the entire first dried product, and more preferably 10% by mass or less. If the drying has progressed to this extent, it can be considered that the solvent, such as the extractant, has been almost completely removed. Therefore, when performing the water extraction step S114 described below, the extraction of DHI with water can be performed more smoothly, and ultimately, a purified DHI with higher purity can be produced. In addition, the amount of solvent remaining in the first dried product can be measured, for example, by the loss on drying method.

7. Water Extraction Step In the water extraction step S114 (second extraction step), a water extraction process is performed in which water is added to the first dried product to obtain a second extract. The water extraction process is a process for selectively extracting DHI contained in the first dried product by utilizing the difference in solubility in water. Although the water extraction process is a simple process, it is a process that has good separation properties for water-insoluble impurities. Therefore, by performing the water extraction step S114, it is possible to efficiently produce purified DHI with a higher purity.

When water is added to the extraction tank containing the first dried material, the DHI transfers to the water. In other words, the second extract obtained by the water extraction process is an aqueous solution in which DHI is dissolved in water. On the other hand, impurities that are insoluble in water do not dissolve even after water is added. Therefore, DHI and impurities can be separated from each other based on the difference in solubility in water. Examples of impurities include substances that are insoluble in water. In addition to pure water, the water may be distilled water, ion-exchanged water, tap water, industrial water, etc. The first dried material may be added to the extraction tank containing water, or both the first dried material and water may be added to the extraction tank at the same time.

The amount of water added to the first dried product is preferably 400% by mass or more, and more preferably 800% by mass or more, relative to the first dried product. The upper limit does not have to be set, but is preferably 2400% by mass or less, and more preferably 1600% by mass or less. This makes it possible to sufficiently increase the extraction efficiency of DHI and to prevent excessive water use.

After adding water, it is preferable to ensure that there is time for the water to act on the first dried product before moving to the next step. This time is preferably 10 minutes or more, and more preferably 20 minutes or more. Also, no particular upper limit need be set, but it is preferably 120 minutes or less, and more preferably 60 minutes or less. This makes it possible to sufficiently increase the extraction efficiency of DHI and prevent the purification time from becoming excessively long.

In addition, after adding water to obtain a mixture (aqueous solution), the mixture may be stirred as necessary. This can promote the action of water on the first dried material.

8. Supernatant Separation Step In the supernatant separation step S116 (second solid-liquid separation step), a solid-liquid separation process is performed on the second extract containing impurities insoluble in water. The solid-liquid separation process is a process for separating the supernatant and impurities from the second extract containing impurities insoluble in water. The separated supernatant is used as the second separated liquid. Examples of such solid-liquid separation processes include filtration separation, supernatant separation, reduced pressure dehydration, pressurized dehydration, and centrifugation, and the like. In this embodiment, supernatant separation is used. Since the operation is simple, the supernatant separation is useful in that the process can be performed efficiently. In addition, when separating the supernatant, other operations such as centrifugation may be used in combination.

The second separated liquid contains purified DHI at a high purity. Therefore, the second separated liquid may be recovered and used as a purified DHI product.

9. Step of Drying or Concentration Adjustment In the step S118 of drying or concentration adjustment (second drying step or concentration adjustment step), the second separated liquid is subjected to a drying treatment or a concentration adjustment treatment.

The drying process is a process in which the solvent is removed from the second separated liquid and the solute is dried. The powder obtained by this process is called the second dried product. Examples of the drying process include processes involving heating, reducing pressure, and spraying gas. By carrying out such a drying process, a powder of purified DHI can be obtained.

Among these, the heating operation is preferably used. The temperature of the second separated liquid during the heating operation is not particularly limited as long as it is a temperature at which the solvent (water) can be distilled off and is below the boiling point of DHI. The temperature of the second separated liquid during the heating operation is preferably 60°C or higher, and more preferably 90°C or higher. This increases the rate at which the solvent is distilled off, improving purification efficiency and suppressing thermal decomposition and volatilization of DHI. From the viewpoint of suppressing oxidative polymerization of DHI, the upper limit of the temperature of the second separated liquid during the heating operation is preferably 200°C or lower, and more preferably 150°C or lower.

The pressure during the heating operation is not particularly limited and may be atmospheric pressure, but is preferably less than atmospheric pressure (101 kPa), more preferably 90 kPa or less, and even more preferably 80 kPa or less. This allows drying at a relatively low temperature, making it easier to suppress oxidative polymerization of DHI. On the other hand, the lower limit of the pressure does not need to be set, but is preferably 20 kPa or more when considering the cost of reducing pressure, etc.
The heating time is not particularly limited, but is preferably from 1 to 120 minutes, and more preferably from 5 to 90 minutes.

The concentration adjustment process is a process for adjusting the concentration by removing the solvent contained in the second separated liquid or by diluting the second separated liquid. Specific operations of the concentration adjustment process include, for example, an operation for removing the solvent and an operation for adding a solvent, and one or a combination of these operations is used. The operation for removing the solvent is, for example, appropriately selected from the operations described above. On the other hand, the operation for adding the solvent is an operation for adding water to the second separated liquid to dilute it. By carrying out such a concentration adjustment process, an aqueous solution of purified DHI can be obtained.

10. Environment for each step Each of the above steps is preferably performed in an oxygen-free environment. This makes the environment for each step inert, so that the denaturation of purified DHI due to oxidation can be suppressed. An example of the denaturation of purified DHI is melanization. By suppressing the denaturation of purified DHI, it is possible to produce purified DHI with higher purity and at a higher yield.

An oxygen-free environment refers to an environment in which, for example, an inert gas is continuously supplied to a processing tank in which each process is carried out, thereby reducing the oxygen concentration. The oxygen concentration of the gas phase in the processing tank is preferably 10 ppm or less by volume, and more preferably 5 ppm or less. The oxygen concentration of the gas phase in the processing tank is measured by an oxygen concentration meter.

Examples of the inert gas include nitrogen gas and argon gas.
In addition to the water used in the water extraction step S114, the raw material liquid and the extract may have their dissolved oxygen levels reduced in advance, thereby preventing oxidation of DHI by dissolved oxygen.

11. Effects of the First Embodiment As described above, the method for purifying 5,6-dihydroxyindole according to the first embodiment includes a preparation step S102, a liquid-liquid extraction step S104 (first extraction step), a poor solvent addition step S108, and a supernatant separation step S110 (first solid-liquid separation step). In the preparation step S102, a raw material liquid containing 5,6-dihydroxyindole and water before purification is prepared. In the liquid-liquid extraction step S104, an extractant is added to the raw material liquid to obtain a first extract and a first raffinate. In the poor solvent addition step S108, i.e., a simplified purification step, a poor solvent that supersaturates impurities without supersaturating 5,6-dihydroxyindole is added to the first extract to cause precipitation. In other words, the simplified purification step is a step for simply purifying 5,6-dihydroxyindole contained in the first extract. In the supernatant fractionation step S110, a first separation solution containing purified 5,6-dihydroxyindole is recovered by supernatant fractionation (fixed separation treatment).

In this configuration, the simple operation of adding a poor solvent (simple purification process) can efficiently remove relatively polar impurities, making it possible to produce high-purity 5,6-dihydroxyindole.

The method for purifying 5,6-dihydroxyindole according to the first embodiment also includes a drying step S112 (first drying step). In the drying step S112, the first separated liquid recovered in the supernatant fractionation step S110 (first solid-liquid separation step) is dried to recover a first dried product of 5,6-dihydroxyindole after purification.

With this configuration, the first dried material can be directly manufactured into a product, or the first dried material can be subjected to the water extraction process S114 to produce 5,6-dihydroxyindole of higher purity.

The method for purifying 5,6-dihydroxyindole according to the first embodiment includes a water extraction step S114 (second extraction step) and a supernatant separation step S116 (second solid-liquid separation step). In the water extraction step S114, water is added to the first dried product to obtain a second extract. In the supernatant separation step S116, a second separation liquid containing purified 5,6-dihydroxyindole is recovered by supernatant separation (solid-liquid separation process).

In this configuration, water-insoluble impurities can be efficiently removed by the simple operation of adding water, and 5,6-dihydroxyindole of higher purity can be produced. For this reason, in this embodiment, the poor solvent addition step S108 can be considered a primary simplified purification step, and the water extraction step S114 can be considered a secondary simplified purification step. In other words, in this embodiment, by performing multiple simplified purification steps including the poor solvent addition step S108 and the water extraction step S114, 5,6-dihydroxyindole of higher purity (preferably a purity of 95% or more) can be produced.

The method for purifying 5,6-dihydroxyindole according to the first embodiment also includes a step S118 (second drying step or concentration adjustment step) of drying or adjusting the concentration. The second drying step dries the second separated liquid recovered in the supernatant collection step S116 (second solid-liquid separation step) and recovers a second dried product of purified 5,6-dihydroxyindole. The concentration adjustment step adjusts the concentration of the second separated liquid recovered in the supernatant collection step S116 (second solid-liquid separation step) and recovers an aqueous solution containing purified 5,6-dihydroxyindole.

With this configuration, it is possible to recover purified 5,6-dihydroxyindole powder or an aqueous solution of purified 5,6-dihydroxyindole.

In addition, in the method for purifying 5,6-dihydroxyindole according to the first embodiment, the raw material liquid may contain a fermentation liquid produced by fermenting sugar.

With this configuration, the raw material liquid can be obtained from sugars that are abundant in nature, making it easier to obtain. In addition, since the refined 5,6-dihydroxyindole is not derived from fossil resources, it contributes to suppressing the increase of carbon dioxide in the atmosphere.

In addition, in the method for purifying 5,6-dihydroxyindole according to the first embodiment, the raw material liquid may contain unpurified 5,6-dihydroxyindole that has been converted from plant-derived L-tyrosine.

This configuration allows the use of plant-derived starting materials, making it possible to obtain purified 5,6-dihydroxyindole while suppressing the increase in atmospheric carbon dioxide.

The method for purifying 5,6-dihydroxyindole according to the first embodiment also includes a concentration step S106. The concentration step S106 is provided between the liquid-liquid extraction step S104 (first extraction step) and the poor solvent addition step S108, and concentrates the first extraction liquid.

With this configuration, the amount of liquid phase in the first extraction liquid provided to the poor solvent addition step S108 can be reduced. This allows the amount of poor solvent to be added in the poor solvent addition step S108 to be reduced, and the amount of impurities dissolved in the liquid phase to be reduced. As a result, the purity of the purified DHI finally obtained can be further increased.

In addition, in the method for purifying 5,6-dihydroxyindole according to the first embodiment, 5,6-dihydroxyindole is purified in an oxygen-free environment.

This configuration makes it possible to suppress the denaturation of purified DHI that occurs due to oxidation. This makes it possible to produce purified DHI with even higher purity and at a higher yield.

The method for purifying 5,6-dihydroxyindole of the present invention has been described above based on the first embodiment, but the present invention is not limited to the first embodiment. For example, the method for purifying 5,6-dihydroxyindole of the present invention may be a purification method in which any step is added to the first embodiment.

Next, a second embodiment of the 5,6-dihydroxyindole purification method of the present invention will be described in detail with reference to the attached drawings.

FIG. 2 is a process diagram for explaining the purification method of 5,6-dihydroxyindole according to the second embodiment.

In the method for purifying 5,6-dihydroxyindole according to this embodiment, unpurified 5,6-dihydroxyindole is purified and high-purity 5,6-dihydroxyindole is recovered. In other words, the method for purifying 5,6-dihydroxyindole according to this embodiment is a method for producing purified 5,6-dihydroxyindole. 5,6-dihydroxyindole is used as a raw material for various applications, such as cosmetics, pharmaceuticals, and agricultural chemicals. To realize these applications, it is essential to highly purify the raw material, 5,6-dihydroxyindole.

The purification method of 5,6-dihydroxyindole shown in FIG. 2 includes a preparation step S1021, a liquid-liquid extraction step S1041 (first extraction step), a drying step S1061 (first drying step), a water extraction step S1081 (second extraction step), a supernatant separation step S1101 (first solid-liquid separation step), and a drying or concentration adjustment step S1121 (second drying step or concentration adjustment step). In this embodiment, the drying step S1061 (first drying step) and the water extraction step S1081 (second extraction step) are simplified purification steps. In this embodiment, too, the simplified purification step is a step for simply purifying 5,6-dihydroxyindole contained in the first extract. The preparation step S1021 and the liquid-liquid extraction step S1041 are the same as the preparation step S102 and the liquid-liquid extraction step S104 of the first embodiment described above. That is, the first extract is obtained in the liquid-liquid extraction step S1041. Below, the explanation of the preparation step S1021 and the liquid-liquid extraction step S1041 will be omitted, and the explanation will start from the drying step S1061.

12. Drying Step In the drying step S1061 (first drying step), a drying process is performed on the first extract. The drying process is a process for removing the solvent from the first extract and drying the solute. In this embodiment, the powder obtained by this process is the first dried product. Examples of the drying process include processes involving heating, decompression, and gas blowing. According to such a drying process, the solute can be dried while minimizing the loss of DHI associated with the drying process. In this specification, the purified DHI is also referred to as "purified DHI".

For the drying process, a heating operation is preferably used. The temperature of the first extract during the heating operation is not particularly limited as long as it is a temperature at which the solvent can be distilled off and is below the boiling point of DHI. The temperature of the first extract during the heating operation is preferably 40 to 90°C, more preferably 50 to 85°C, and even more preferably 60 to 80°C. This increases the rate at which the solvent is distilled off, improving purification efficiency and suppressing thermal decomposition and volatilization of DHI.

The pressure during the heating operation is not particularly limited and may be atmospheric pressure, but is preferably less than atmospheric pressure (101 kPa), more preferably 90 kPa or less, and even more preferably 80 kPa or less. This allows drying at a relatively low temperature, making it easier to suppress oxidative polymerization of DHI. On the other hand, the lower limit of the pressure does not need to be set, but is preferably 20 kPa or more when considering the cost of reducing pressure, etc.
The heating time is not particularly limited, but is preferably from 1 to 120 minutes, and more preferably from 5 to 90 minutes.

In the drying step S1061, it is not necessary to remove all of the solvent contained in the first extract. Specifically, the amount of solvent remaining in the first dried product is preferably 30% by mass or less of the entire first dried product, and more preferably 10% by mass or less. If the drying has progressed to this extent, it can be considered that the solvent, such as the extractant, has been almost completely removed. Therefore, when performing the water extraction step S1081 described below, the extraction of DHI with water can be performed more smoothly, and ultimately, a purified DHI with higher purity can be produced. In addition, the amount of solvent remaining in the first dried product can be measured, for example, by the loss on drying method.

13. Water Extraction Step In the water extraction step S1081 (second extraction step), a water extraction process is performed in which water is added to the first dried product to obtain a second extract. The water extraction process is a process for selectively extracting DHI contained in the first dried product by utilizing the difference in solubility in water. Although the water extraction process is a simple process, it is a process that has good separation properties for water-insoluble impurities. Therefore, by performing the water extraction step S1081, it is possible to efficiently produce purified DHI with a higher purity.

When water is added to the extraction tank containing the first dried material, the DHI transfers to the water. In other words, the second extract obtained by the water extraction process is an aqueous solution in which DHI is dissolved in water. On the other hand, impurities that are insoluble in water do not dissolve even after water is added. Therefore, DHI and impurities can be separated from each other based on the difference in solubility in water. Examples of impurities include substances that are insoluble in water. In addition to pure water, the water may be distilled water, ion-exchanged water, tap water, industrial water, etc. The first dried material may be added to the extraction tank containing water, or both the first dried material and water may be added to the extraction tank at the same time.

The amount of water added to the first dried product is preferably 400% by mass or more, and more preferably 800% by mass or more, relative to the first dried product. The upper limit does not have to be set, but is preferably 2400% by mass or less, and more preferably 1600% by mass or less. This makes it possible to sufficiently increase the extraction efficiency of DHI and to prevent excessive water use.

After adding water, it is preferable to ensure that there is time for the water to act on the first dried product before moving to the next step. This time is preferably 10 minutes or more, and more preferably 20 minutes or more. Also, no particular upper limit need be set, but it is preferably 120 minutes or less, and more preferably 60 minutes or less. This makes it possible to sufficiently increase the extraction efficiency of DHI and prevent the purification time from becoming excessively long.

In addition, after adding water to obtain a mixture (aqueous solution), the mixture may be stirred as necessary. This can promote the action of water on the first dried material.

14. Supernatant Separation Step In the supernatant separation step S1101 (first solid-liquid separation step), a solid-liquid separation process is performed on the second extract containing impurities insoluble in water. The solid-liquid separation process is a process for separating the supernatant and impurities from the second extract containing impurities insoluble in water. The separated supernatant is used as a separation liquid (first separation liquid). Examples of such solid-liquid separation processes include filtration separation, supernatant separation, reduced pressure dehydration, pressurized dehydration, and centrifugation, and the like. In this embodiment, supernatant separation is used. Supernatant separation is useful in that the operation is simple and the process can be performed efficiently. In addition, when separating the supernatant, other operations such as centrifugation may be used in combination.

The separated liquid contains purified DHI at a high purity. Therefore, the separated liquid can be recovered and used as a purified DHI product.

15. Step of Drying or Concentration Adjustment In the step S1121 of drying or concentration adjustment (second drying step or concentration adjustment step), the separated liquid is subjected to a drying treatment or a concentration adjustment treatment.

The drying process is a process in which the solvent is removed from the separated liquid and the solute is dried. The powder obtained by this process is called the second dried product. Examples of the drying process include processes involving heating, reducing pressure, and spraying gas. By carrying out such a drying process, a powder of purified DHI can be obtained.

Among these, the heating operation is preferably used. The temperature of the separated liquid during the heating operation is not particularly limited as long as it is a temperature at which the solvent (water) can be distilled off and is below the boiling point of DHI. The temperature of the separated liquid during the heating operation is preferably 60°C or higher, more preferably 90°C or higher. This can increase the distillation rate of the solvent, improve the purification efficiency, and suppress the thermal decomposition and volatilization of DHI. From the viewpoint of suppressing the oxidative polymerization of DHI, the upper limit of the temperature of the separated liquid during the heating operation is preferably 200°C or lower, more preferably 150°C or lower.
The various conditions during the heating operation (pressure, heating time, etc.) can be the same as those during the heating operation in the first embodiment described above. The concentration adjustment process can be the same as that in the first embodiment described above. The environment in which each process is performed can be the same as that in the first embodiment described above.

16. Effects of the Second Embodiment As described above, the purification method of 5,6-dihydroxyindole according to the second embodiment includes a preparation step S1021, a liquid-liquid extraction step S1041 (first extraction step), a drying step S1061 (first drying step), a water extraction step S1081 (second extraction step), and a supernatant separation step S1101 (first solid-liquid separation step). In the preparation step S1021, a raw material liquid containing 5,6-dihydroxyindole and water before purification is prepared. In the liquid-liquid extraction step S1041, an extractant is added to the raw material liquid to obtain a first extract and a first raffinate. In the drying step S1061, the first extract is dried to obtain a first dried product. In the water extraction step S1081 (second extraction step), water is added to the first dried product to obtain a second extract. That is, the 5,6-dihydroxyindole contained in the first extract is simply purified to obtain a second extract by a simplified purification process including a drying process S1061 and a water extraction process S1081. In the supernatant separation process S1101, a separation liquid containing the purified 5,6-dihydroxyindole is recovered by supernatant separation (fixed separation process).

In this configuration, by performing a simple operation (simple purification process) of performing an extraction process using water after obtaining the first dried product, water-insoluble impurities can be efficiently removed, making it possible to produce high-purity 5,6-dihydroxyindole.

The purification method for 5,6-dihydroxyindole according to the second embodiment also includes a step S1121 (second drying step or concentration adjustment step) of drying or adjusting the concentration. The second drying step dries the separated liquid recovered in the supernatant separation step S1101 (first solid-liquid separation step) and recovers a second dried product of purified 5,6-dihydroxyindole. The concentration adjustment step adjusts the concentration of the separated liquid recovered in the supernatant separation step S1101 (first solid-liquid separation step) and recovers an aqueous solution containing purified 5,6-dihydroxyindole.

With this configuration, it is possible to recover purified 5,6-dihydroxyindole powder or an aqueous solution of purified 5,6-dihydroxyindole.

In addition, in the method for purifying 5,6-dihydroxyindole according to the second embodiment, the raw material liquid contains a fermentation liquid produced by fermenting sugar.

With this configuration, the raw material liquid can be obtained from sugars that are abundant in nature, making it easier to obtain. In addition, since the refined 5,6-dihydroxyindole is not derived from fossil resources, it contributes to suppressing the increase of carbon dioxide in the atmosphere.

In addition, in the method for purifying 5,6-dihydroxyindole according to the second embodiment, the raw material liquid may contain unpurified 5,6-dihydroxyindole that has been converted from plant-derived L-tyrosine.

This configuration allows the use of plant-derived starting materials, making it possible to obtain purified 5,6-dihydroxyindole while suppressing the increase in atmospheric carbon dioxide.

In addition, in the method for purifying 5,6-dihydroxyindole according to the second embodiment, 5,6-dihydroxyindole is purified in an oxygen-free environment.

This configuration makes it possible to suppress the denaturation of purified DHI that occurs due to oxidation. This makes it possible to produce purified DHI with even higher purity and at a higher yield.

The method for purifying 5,6-dihydroxyindole of the present invention has been described above based on the second embodiment, but the present invention is not limited to the second embodiment. For example, the method for purifying 5,6-dihydroxyindole of the present invention may be a purification method in which any step is added to the second embodiment.

Next, a third embodiment of the 5,6-dihydroxyindole purification method of the present invention will be described in detail with reference to the attached drawings.

FIG. 3 is a process diagram for explaining a method for purifying 5,6-dihydroxyindole according to a third embodiment. The third embodiment will be explained below, focusing on the differences from the first embodiment described above, and omitting explanations of similar points.

The method for purifying 5,6-dihydroxyindole shown in Figure 3 includes a preparation step S1, a liquid-liquid extraction step S2, a concentration step (first concentration step) S3, a poor solvent addition step S4, a supernatant separation step S5, a concentration step (second concentration step) S6, a water addition step S7, a solvent replacement step S8, a supernatant separation step S9, and a concentration adjustment step S10. This embodiment differs from the first embodiment described above mainly in that it includes a concentration step (second concentration step) S6, a water addition step S7, and a solvent replacement step S8 after the supernatant separation step S5.

In other words, the preparation step S1, the liquid-liquid extraction step S2, the concentration step (first concentration step) S3, the poor solvent addition step S4, and the supernatant separation step S5 are the same as or similar to the preparation step S102, the liquid-liquid extraction step S104 (first extraction step), the concentration step S106, the poor solvent addition step S108, and the supernatant separation step S110 (first solid-liquid separation step) of the first embodiment described above. Also, in this embodiment, the poor solvent addition step S108 is a simplified purification step. Also, the first separated liquid is obtained in the supernatant separation step S5. The second concentration step S6 will be described below.

17. Second Concentration Step In the second concentration step S6, the first separation liquid obtained in the supernatant separation step S5 is subjected to a concentration treatment. The concentration treatment method can be the same as the method described above in the first embodiment (for example, the method in the concentration step S106). The second concentration step S6 may be provided as necessary, and may be omitted.

18. Water Addition Step In the water addition step S7, water is added to the concentrated first separation liquid to obtain a mixed liquid. The mixed liquid contains the extractant, poor solvent, and purified DHI contained in the first separation liquid, and water. In this manner, DHI is dissolved in a plurality of solvents (extractant, poor solvent, and water). In this embodiment, the type of water, the addition method, etc. described above in the first embodiment (for example, the type of water, the addition method, etc. described in the water extraction step S114) can be appropriately used.

19. Solvent Replacement Step In the solvent replacement step S8, a process for removing a specific solvent (in this embodiment, an extractant and a poor solvent) is performed on the mixed liquid obtained in the water addition step S7. That is, the solvent replacement step can be called a specific solvent removal process for removing a specific solvent from the mixed liquid. As a process for removing a specific solvent, a heat treatment (evaporation treatment or distillation treatment) of the mixed liquid is preferable. However, the process for removing a specific solvent is not limited to a heat treatment. For example, the specific solvent may be removed by an adsorption process in which an adsorbent that selectively adsorbs a specific solvent is added to the mixed liquid, or a membrane separation process in which the mixed liquid is passed through a membrane that selectively allows the specific solvent to pass therethrough. The case where a heat treatment (evaporation treatment) is used will be described below.

The heating process is a process in which the mixed liquid is heated to remove the extractant and poor solvent while leaving water behind. This allows an aqueous solution containing purified DHI to be recovered. In other words, the heating process in the solvent replacement step S8 is a process that utilizes the boiling point difference between multiple solvents (extractant, poor solvent, and water) to evaporate the extractant and poor solvent while leaving water behind.

For example, when ethyl acetate (boiling point 77°C) is used as the extractant and hexane (boiling point 69°C) is used as the poor solvent, the extraction agent and poor solvent can be removed while leaving water by performing a heating process at a heating temperature of about 90°C. From this perspective, the difference in boiling points between the multiple solvents (extractant, poor solvent, and water) is preferably 5°C or more, and more preferably 10°C or more and 30°C or less.

The heating treatment can be carried out by appropriately applying the heating operation described above in the first embodiment (for example, the heating operation described in the drying step S112). As a result of removing the extractant and poor solvent in the solvent replacement step S8, impurities that are dissolved in the extractant or poor solvent may precipitate, but are insoluble in water, so it is preferable to remove such impurities.

20. Supernatant collection step In the supernatant collection step S9, a solid-liquid separation process is performed on the aqueous solution containing water-insoluble impurities. The solid-liquid separation process can be performed by appropriately applying the solid-liquid separation process described above in the first embodiment (for example, the solid-liquid separation process described in the supernatant collection step S116). This results in a separated supernatant (second separated liquid). The supernatant collection step S9 may be provided as necessary, and may be omitted.

21. Concentration Adjustment Step In the concentration adjustment step S10, the second separated liquid is subjected to a concentration adjustment process. The concentration adjustment process can be carried out by appropriately applying the concentration adjustment process described above in the first embodiment (for example, the concentration adjustment process described in the drying or concentration adjustment step S118). This results in an aqueous solution of purified DHI adjusted to a predetermined concentration.

22. Advantages of the Third Embodiment In the first embodiment, water is added to the first dried product obtained by subjecting the first separated liquid to a drying process. On the other hand, in the present embodiment, water is added to the first separated liquid. Thus, in the present embodiment, the timing of adding water is different from that in the first embodiment. This allows DHI to be handled in a liquid state at all times. As a result, the stirring load can be reduced, and purified DHI can be efficiently produced.
In this embodiment, the poor solvent addition step S4 can be regarded as a first simplified purification step, and the solvent substitution step S8 can be regarded as a secondary simplified purification step. In other words, in this embodiment, by carrying out a plurality of simplified purification steps including the poor solvent addition step S4 and the solvent substitution step S8, it is possible to produce 5,6-dihydroxyindole with a higher purity (preferably a purity of 95% or more).

The method for purifying 5,6-dihydroxyindole of the present invention has been described above based on the third embodiment, but the present invention is not limited to the third embodiment. For example, the method for purifying 5,6-dihydroxyindole of the present invention may be a purification method in which any step is added to the third embodiment.

Next, a fourth embodiment of the 5,6-dihydroxyindole purification method of the present invention will be described in detail with reference to the attached drawings.

FIG. 4 is a process diagram for explaining a method for purifying 5,6-dihydroxyindole according to the fourth embodiment. The fourth embodiment will be explained below, focusing on the differences from the second embodiment described above, and omitting explanations of similar points.

The purification method of 5,6-dihydroxyindole shown in FIG. 4 includes a preparation step S11, a liquid-liquid extraction step S12, a concentration step S13, a water addition step S14, a solvent replacement step S15, a supernatant separation step S16, and a concentration adjustment step S17. This embodiment differs from the second embodiment described above mainly in that it includes a water addition step S14 and a solvent replacement step S15 instead of the drying step S1061 and water extraction step S1081 of the second embodiment. In other words, in this embodiment, the water addition step S14 and the solvent replacement step S15 are simplified purification steps, and the steps other than the simplified purification steps are the same as or similar to the steps of the second embodiment described above. Then, a first extract is obtained in the liquid-liquid extraction step S12. The following will begin with the concentration step S13.

23. Concentration Step In the concentration step S13, the first extract obtained in the liquid-liquid extraction step S12 is subjected to a concentration treatment. The concentration treatment method can be the same as the method described above in the first embodiment (for example, the method in the concentration step S106). In addition, the concentration step S13 may be provided as necessary, and may be omitted.

24. Water Addition Step In the water addition step S14, water is added to the concentrated first extract to obtain a mixed solution. The mixed solution contains the extractant and purified DHI contained in the first extract, and water. In this manner, DHI is dissolved in a plurality of solvents (extractant and water). In this embodiment, the type of water, the addition method, etc. described in the first embodiment (for example, the type of water, the addition method, etc. described in the water extraction step S114) can be appropriately used.

25. Solvent Replacement Step In the solvent replacement step S15, a process for removing a specific solvent (in this embodiment, an extractant) is performed on the mixed liquid obtained in the water addition step S14. The solvent replacement step can be called a specific solvent removal step in which a specific solvent is removed by the process (e.g., heat treatment) described above in the third embodiment. The following describes the case where a heat treatment (evaporation treatment) is used.

The heating process in the solvent replacement step S15 is a process in which the mixed liquid is heated to remove the extractant while leaving water behind. This makes it possible to recover an aqueous solution containing purified DHI. In other words, the heating process in the solvent replacement step S15 is a process in which the boiling point difference between multiple solvents (extractant and water) is used to evaporate the extractant while leaving water behind.

For example, when ethyl acetate (boiling point 77°C) is used as the extractant, the heating process can be performed at a heating temperature of about 90°C, thereby removing the extractant while leaving water behind. From this perspective, it is preferable that the boiling point difference between the multiple solvents (extractant and water) is 5°C or more, and more preferably 10°C or more and 30°C or less.

The heating treatment can be carried out by appropriately applying the heating operation described above in the first embodiment (for example, the heating operation described in the drying step S112). As a result of removing the extractant in the solvent replacement step S15, impurities that are dissolved in the extractant may precipitate, but are insoluble in water, so it is preferable to remove such impurities.

26. Supernatant collection step In the supernatant collection step S16, a solid-liquid separation process is performed on the aqueous solution containing water-insoluble impurities. The solid-liquid separation process can be performed by appropriately applying the solid-liquid separation process described above in the first embodiment (for example, the solid-liquid separation process described in the supernatant collection step S116). This allows the separated supernatant (separated liquid) to be obtained. The supernatant collection step S16 may be provided as necessary, and may be omitted.

27. Concentration Adjustment Step In the concentration adjustment step S17, the separated liquid is subjected to a concentration adjustment process. The concentration adjustment process can be carried out by appropriately applying the concentration adjustment process described above in the first embodiment (for example, the concentration adjustment process described in the drying or concentration adjustment step S118). As a result, an aqueous solution of purified DHI adjusted to a predetermined concentration is obtained.

28. Effects of the Fourth Embodiment In the second embodiment, water is added to the first dried product obtained by drying the first extract. On the other hand, in the present embodiment, water is added to the first extract. Thus, in the present embodiment, the timing of adding water is different from that in the second embodiment. This allows DHI to be handled in a liquid state at all times. As a result, the stirring load can be reduced, and purified DHI can be efficiently produced.

The method for purifying 5,6-dihydroxyindole of the present invention has been described above based on the fourth embodiment, but the present invention is not limited to the fourth embodiment. For example, the method for purifying 5,6-dihydroxyindole of the present invention may be a purification method in which any step is added to the fourth embodiment.

Next, a specific example A of the present invention will be described.
12. Preparation of Purified DHI 12.1. Example 1A
First, a fermentation liquid produced by fermentation of glucose was prepared as a raw material liquid. The water content in the raw material liquid was 90% by mass. Next, a liquid-liquid extraction process was carried out by contacting the raw material liquid with an extractant. As a result, a first extract was obtained. As the extractant, ethyl acetate was used in an amount of 50% by volume relative to the raw material liquid.

Then, the obtained first extract was concentrated by heating. Next, a poor solvent was added to the concentrated first extract. This operation caused a precipitate to form in the first extract. Hexane was used as the poor solvent. The amount of poor solvent added to the concentrated first extract was 200% by volume.

Next, the supernatant of the first extract containing the precipitate was separated to obtain a first separated liquid. The separated first separated liquid was then subjected to a drying treatment by heating. This resulted in a first dried product.

Next, a water extraction process was performed in which water was added to the first dried product. This resulted in a second extract containing water-insoluble impurities. The amount of water added to the first dried product was 900% by mass.

Next, the supernatant of the second extract containing water-insoluble impurities was separated to obtain a second separated liquid. The separated second separated liquid was then subjected to a drying treatment by heating. This resulted in a second dried product, which was a powder of purified DHI. Each of the above steps was carried out in an oxygen-free environment while supplying nitrogen gas.

12.2. Examples 2A to 6A and Comparative Examples 1A to 2A
Purified DHI was produced in the same manner as in Example 1A, except that the production process of purified DHI was changed as shown in Table 1.

13. Evaluation of purified DHI The purity of the purified DHI obtained in each Example A and each Comparative Example A was measured by high performance liquid chromatography (HPLC). The purity was defined as the ratio of the mass of DHI to the total mass of the recovered material. The measurement results are shown in Table 1.
The conditions for HPLC analysis of DHI are as follows:

Column: COSMOSIL 5C18-AR-II (φ4.6 mm × 250 mm) manufactured by Nacalai Tesque Mobile phase: water/methanol/perchloric acid = 4/1/0.0075 (vol/vol/vol) isocratic elution Flow rate: 1 mL/min
Column temperature: 40°C
Detection method: Photodiode array (PDA) detector

As is clear from Table 1, in each Example A, by simply carrying out the simple operation of adding a poor solvent (simplified purification step), it was possible to produce purified DHI of higher purity than in each Comparative Example A. Furthermore, when the poor solvent addition step was used in combination with the water extraction step or the solvent replacement step (when multiple simplified purification steps were carried out), this tendency was more pronounced. This is thought to be due to the different types of impurities that can be removed in each step. Furthermore, by adding a concentration step, it was possible to achieve high purity even if the amount of poor solvent used in the poor solvent addition step was reduced.

From the above, it has become clear that the present invention makes it possible to efficiently produce highly purified DHI.

Next, a specific embodiment B of the present invention will be described.
9. Preparation of Purified DHI 9.1. Example 1B
First, a fermentation liquid produced by fermentation of glucose was prepared as a raw material liquid. The water content in the raw material liquid was 90% by mass. Next, a liquid-liquid extraction process was carried out by contacting the raw material liquid with an extractant. As a result, a first extract was obtained. As the extractant, ethyl acetate was used in an amount of 50% by volume relative to the raw material liquid.

Then, the first extract obtained was subjected to a drying process by heating. This resulted in the first dried product.

Next, a water extraction process was performed in which water was added to the first dried product. This resulted in a second extract containing water-insoluble impurities. The amount of water added to the first dried product was 1000% by mass.

Next, the supernatant of the second extract containing water-insoluble impurities was separated to obtain a separated liquid. The separated liquid was then dried by heating. This resulted in a second dried product, which was a powder of purified DHI. Each of the above steps was carried out in an oxygen-free environment while supplying nitrogen gas.

9.2. Examples 2B to 4B and Comparative Examples 1B to 3B
Purified DHI was produced in the same manner as in Example 1B, except that the production process of purified DHI was changed as shown in Table 2.

10. Evaluation of purified DHI The purity of the purified DHI obtained in each Example B and each Comparative Example B was measured by high performance liquid chromatography (HPLC). The purity was defined as the ratio of the mass of DHI to the total mass of the recovered material. The measurement results are shown in Table 2.
The conditions for HPLC analysis of DHI are as follows.

Column: COSMOSIL 5C18-AR-II (φ4.6 mm × 250 mm) manufactured by Nacalai Tesque Mobile phase: water/methanol/perchloric acid = 4/1/0.0075 (vol/vol/vol) isocratic elution Flow rate: 1 mL/min
Column temperature: 40°C
Detection method: Photodiode array (PDA) detector

As is clear from Table 2, in each Example B, it was possible to produce purified DHI of higher purity than in each Comparative Example B by simply performing the simple operations of drying and water extraction (simple purification process) or the simple operations of water addition and solvent replacement (simple purification process).

From the above, it has become clear that the present invention makes it possible to efficiently produce highly purified DHI.

According to the present invention, 5,6-dihydroxyindole can be purified to a high purity. Therefore, the present invention has industrial applicability.

S102 Preparation step S104 Liquid-liquid extraction step S106 Concentration step S108 Poor solvent addition step S110 Supernatant separation step S112 Drying step S114 Water extraction step S116 Supernatant separation step S118 Drying or concentration adjustment step S1021 Preparation step S1041 Liquid-liquid extraction Step S1061 Drying step S1081 Water extraction step S1101 Supernatant separation step S1121 Drying or concentration adjustment step S6 Concentration step S7 Water addition step S8 Solvent replacement step S13 Concentration step S14 Water addition step S15 Solvent replacement step

Claims (19)

A preparation step of preparing a raw material solution containing unpurified 5,6-dihydroxyindole and water;
a first extraction step of adding an extractant to the raw material liquid to obtain a first extract containing the 5,6-dihydroxyindole and a first raffinate;
A simplified purification step for simply purifying the 5,6-dihydroxyindole contained in the first extract;
a first solid-liquid separation step of recovering a first separated liquid containing the purified 5,6-dihydroxyindole by solid-liquid separation treatment;
A method for purifying 5,6-dihydroxyindole, comprising the steps of: The method for purifying 5,6-dihydroxyindole according to claim 1, wherein the simplified purification step includes a step of adding a poor solvent to the first extract, which causes a precipitation by adding a poor solvent that supersaturates impurities without supersaturating the 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 2, further comprising a first drying step of drying the first separated liquid recovered in the first solid-liquid separation step and recovering a first dried product containing the purified 5,6-dihydroxyindole. a second extraction step of adding water to the first dried product to obtain a second extract;
a second solid-liquid separation step of recovering a second separated liquid containing the purified 5,6-dihydroxyindole by solid-liquid separation treatment;
The method for purifying 5,6-dihydroxyindole according to claim 3, comprising the steps of: The method for purifying 5,6-dihydroxyindole according to claim 4, further comprising a second drying step of drying the second separated liquid recovered in the second solid-liquid separation step and recovering a second dried product containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 4, further comprising a concentration adjustment step of adjusting the concentration of the second separated liquid recovered in the second solid-liquid separation step and recovering an aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 2, further comprising a concentration step for concentrating the first extract, which is provided between the first extraction step and the poor solvent addition step. a water addition step of adding water to the first separated liquid recovered in the first solid-liquid separation step to obtain a mixed liquid containing the extractant, the poor solvent, and the water as a solvent;
The method for purifying 5,6-dihydroxyindole according to claim 2, further comprising a solvent replacement step of removing the extractant and the poor solvent from the mixed solution to recover an aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 8, further comprising a concentration adjustment step of adjusting the concentration of the aqueous solution recovered in the solvent replacement step and recovering the aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 8, further comprising a concentration step in which the first separated liquid recovered in the first solid-liquid separation step is concentrated in advance. The method for purifying 5,6-dihydroxyindole according to claim 1, wherein the simplified purification process includes a first drying process for drying the first extract to obtain a first dried product, and a second extraction process for adding water to the first dried product to obtain a second extract. The method for purifying 5,6-dihydroxyindole according to claim 11, further comprising a second drying step of drying the first separated liquid recovered in the first solid-liquid separation step and recovering a second dried product containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 11, further comprising a concentration adjustment step of adjusting the concentration of the first separated liquid recovered in the first solid-liquid separation step and recovering an aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 1, wherein the simplified purification step includes a water addition step of adding water to the first extract obtained in the first extraction step to obtain a mixed solution containing the extractant and the water as a solvent, and a solvent replacement step of removing the extractant from the mixed solution to recover an aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 14, further comprising a concentration adjustment step of adjusting the concentration of the aqueous solution recovered in the solvent replacement step and recovering the aqueous solution containing the purified 5,6-dihydroxyindole. The method for purifying 5,6-dihydroxyindole according to claim 14, further comprising a concentration step in which the first extract obtained in the first extraction step is concentrated in advance. The method for purifying 5,6-dihydroxyindole according to claim 1, wherein the raw material liquid contains a fermentation liquid produced by fermenting sugar. The method for purifying 5,6-dihydroxyindole according to claim 1, wherein the raw material liquid contains the 5,6-dihydroxyindole before purification that has been converted from plant-derived L-tyrosine. The method for purifying 5,6-dihydroxyindole according to claim 1, in which the 5,6-dihydroxyindole is purified in an oxygen-free environment.