WO2020090813A1

WO2020090813A1 - Method for producing flavonoid

Info

Publication number: WO2020090813A1
Application number: PCT/JP2019/042359
Authority: WO
Inventors: 有福　征宏; 佐野　理志; 上面　雅義; 北山　和弘
Original assignee: 日立化成株式会社
Priority date: 2018-10-30
Filing date: 2019-10-29
Publication date: 2020-05-07
Also published as: TW202017913A; JPWO2020090813A1

Abstract

A method for producing a flavonoid, which comprises: an extraction step wherein a material to be extracted containing a flavonoid is brought into contact with an extractant that contains liquefied dimethyl ether, thereby extracting an extract liquid that contains the flavonoid; and a vaporization step wherein at least some of the extractant is removed from the extract liquid by means of vaporization, thereby obtaining an extract that contains the flavonoid.

Description

Method for producing flavonoids

The present disclosure relates to a method for producing flavonoids.

Flavonoids are a group of naturally occurring organic compounds and are contained in flowers, leaves, roots, stems, fruits, seeds, etc. of various plants including citrus fruits and beans. Flavonoids have different characteristics and actions depending on the type, but most of them have a strong antioxidant action. For example, polymethoxyflavone, which is a flavonoid contained in citrus fruits, is known to have an antioxidant action, a carcinogenesis inhibiting action, an antibacterial action, an antiviral action, an antiallergic action, a melanin production inhibiting action, a blood glucose level inhibiting action, and the like. Therefore, it is expected to be applied to various uses such as pharmaceuticals, health foods, and cosmetics.

As a method for producing flavonoids from citrus fruits, for example, a method is known in which flavonoids are extracted from a citrus peel and the like with an aqueous ethanol solution, and the extracted flavonoids are recovered from the solution (see, for example, Patent Document 1).

JP, 2005-145824, A

However, the conventional flavonoid production method has a problem that the concentration of the obtained flavonoid is low. Therefore, it is required to develop a production method capable of improving the concentration of the obtained flavonoid.

The present disclosure has been made in view of the above problems of the conventional art, and an object of the present disclosure is to provide a method for producing flavonoids that can improve the concentration of the obtained flavonoids.

In order to achieve the above object, the present disclosure, an extraction step of contacting an extractant containing flavonoids with an extractant containing liquefied dimethyl ether, and extracting an extract containing flavonoids, and an extractant from the extract. Vaporizing and removing at least a part of the flavonoids to obtain an extract containing flavonoids, and a method for producing flavonoids.

According to the above production method, by extracting flavonoids from the extract using the extractant containing liquefied dimethyl ether, the extraction obtained is compared with the conventional method of extracting using an aqueous ethanol solution or the like. The concentration of flavonoids in the product can be greatly improved. Further, according to the above-mentioned production method, not only flavonoids but also glycosides thereof can be extracted. Here, the present inventors can extract flavonoids from dimethyl ether as a gas at normal temperature and normal pressure (25 ° C., 0.1 MPa) and a liquefied gas that can be easily liquefied at a pressure of 1 MPa or less. It was found that it is a solvent with extremely rare properties. Then, the present inventors have found that flavonoids can be extracted at a high concentration by liquefying this dimethyl ether and using it as an extractant. In addition, since the liquefied gas easily becomes a gas when the pressure is returned to normal pressure, the extractant contained in the extract can be easily reduced to zero and vaporized by setting the pressure to normal pressure. The dimethyl ether can be recovered as a gas, and the waste solvent can be significantly reduced as compared with the conventional technique.

In the above-mentioned production method, the flavonoids may contain at least one of sudatin and glycosidase. According to the above-mentioned production method, sudatin and glycosides can be extracted particularly efficiently.

In the above production method, the extraction material may include Sudachi pericarp. According to the above-mentioned production method, flavonoids can be extracted particularly efficiently from the extract containing the Sudachi peel.

The production method may further include an acid treatment step of acid-treating the extract before the extraction step. By performing the acid treatment step, at least a part of the flavonoid glycosides contained in the extract can be decomposed into flavonoids, and the extraction efficiency of flavonoids in the extraction step can be improved.

The production method may further include, after the vaporization step, a decomposition step of decomposing at least a part of the flavonoid glycosides in the extract into flavonoids. By performing the decomposition step, the flavonoid concentration can be further improved.

According to the present disclosure, it is possible to provide a method for producing flavonoids that can improve the concentration of the obtained flavonoids.

FIG. 3 is a schematic diagram for explaining one embodiment of a method for producing flavonoids of the present disclosure.

Hereinafter, the present disclosure will be described in detail according to its preferred embodiments. However, the present disclosure is not limited to the following embodiments.

In the present specification, the numerical range indicated by using "to" indicates the range including the numerical values before and after "to" as the minimum value and the maximum value, respectively. In the numerical ranges described stepwise in the present specification, the upper limit value or the lower limit value of the numerical range of a certain stage can be arbitrarily combined with the upper limit value or the lower limit value of the numerical range of another stage. In the numerical range described in this specification, the upper limit value or the lower limit value of the numerical range may be replaced with the values shown in the examples. “A or B” may include either one of A and B, or may include both. Unless otherwise specified, the materials exemplified in the present specification can be used alone or in combination of two or more kinds.

The method for producing flavonoids of the present disclosure, an extractant containing flavonoids is brought into contact with an extractant containing liquefied dimethyl ether to extract an extract containing flavonoids, and at least the extractant is extracted from the extract. A part of which is vaporized and removed to obtain an extract containing flavonoids. Here, FIG. 1 is a schematic diagram for explaining one embodiment of the method for producing flavonoids of the present disclosure. Hereinafter, the method for producing flavonoids according to this embodiment will be described with reference to FIG.

[Extraction process]
In the extraction step, the extractant 21 containing flavonoids is brought into contact with the extractant 22 containing liquefied dimethyl ether in the extraction tank 12. Liquefied dimethyl ether is contained in the liquefied gas cylinder 11 and is supplied from the liquefied gas cylinder 11 to the extraction tank 12 by opening and closing a valve 31 provided between the liquefied gas cylinder 11 and the extraction tank 12. The extractant 22 is preferably supplied so that the entire extractant 21 is immersed in the extractant 22. A filtration filter 23 is installed in the extraction tank 12, and the extraction material 21 is arranged on the filtration filter 23.

In the extraction step, it is preferable to shake the extraction tank 12 and / or agitate the inside of the extraction tank 12 in order to promote extraction of flavonoids from the extraction material 21 to the extraction agent 22. By performing these operations, high-concentration flavonoids can be extracted in a shorter time.

The liquid temperature of the extractant 22 in the extraction tank 12 is not particularly limited as long as it can keep dimethyl ether in a liquid state, but may be, for example, 80 ° C. or lower and 10 to 40 ° C. Good.

The pressure in the extraction tank 12 is not particularly limited as long as dimethyl ether is liquid, and may be in a pressurized state or an atmospheric pressure state. When the extraction tank 12 is pressurized, the pressure is higher than atmospheric pressure and may be 100 MPa or less, 10 MPa or less, or 1 MPa or less. By making the inside of the extraction tank 12 under pressure, the liquid temperature of the extractant 22 can be raised, and the extraction of flavonoids from the extractant 21 to the extractant 22 can be promoted. When the inside of the extraction tank 12 is in a pressurized state, the liquid temperature of the extractant 22 in the extraction tank 12 can be made equal to or higher than the boiling point of dimethyl ether at normal pressure while keeping dimethyl ether in a liquid state.

The extraction time in the extraction step is not particularly limited, but may be 1 to 300 minutes, or 20 to 60 minutes. By setting the extraction time to 1 minute or more, the flavonoids contained in the extract 21 can be sufficiently extracted into the extract 22. The extraction time can be adjusted according to the conditions such as the above-described operations such as shaking and stirring, the liquid temperature of the extractant 22, the pressure in the extraction tank 12, and the like.

The ratio of the mass of the extractant 22 to the mass of the extractant 21 (mass of the extractant 22 / mass of the extractant 21) is not particularly limited, but may be, for example, 1 to 1000 or 2 to 20. Good. When the mass ratio is 1 or more, the extractant 21 and the extractant 22 are easily brought into contact with each other, and the flavonoids contained in the extractant 21 are easily extracted into the extractant 22. Further, even if this mass ratio exceeds 1000, the extraction efficiency of flavonoids is difficult to further improve, so from the viewpoint of reducing the amount of extractant 22 used and suppressing the enlargement of the extraction tank 12, the mass ratio. May be 1000 or less.

After extracting flavonoids from the extraction material 21 into the extraction agent 22 by the extraction process, the extraction material 24 is separated from the extraction material 21 and the extraction agent 22 by the filtration filter 23, and the extraction liquid 24 containing the extracted flavonoids and the extraction agent 22 is obtained. Transfer from the extraction tank 12 to the vaporization tank 13. The extraction liquid 24 is transferred by opening and

closing valves

32 and 33 provided between the extraction tank 12 and the vaporization tank 13. Only one of the

valves

32 and 33 may be provided.

The extraction process is usually performed with the valve 32 closed. Thereby, the flavonoids contained in the extraction material 21 can be easily extracted into the extraction material 22 sufficiently. It is also possible to continuously supply the extractant 22, extract flavonoids, and discharge the extract 24 with the

valves

31, 32, and 33 opened.

(Drawing fee)
The lottery material 21 is not particularly limited as long as it contains flavonoids. In the present specification, flavonoids include flavonoids and their glycosides (hereinafter, also referred to as “flavonoid glycosides”), sugar adducts in which sugars are further bound to flavonoid glycosides, and those treated with an enzyme. Including things.

Flavonoids are aromatic compounds having a basic structure of phenylchroman skeleton, and include flavones, flavonols, flavanones, flavanonols, isoflavones, anthocyanins, flavanols, chalcones, aurones and the like. Among these, the flavonoid may be polymethoxyflavone which is a flavone.

Examples of polymethoxyflavone include sudatitin, demethoxysudacitin, nobiletin, tangeretin, pentamethoxyflavone, tetramethoxyflavone, and heptamethoxyflavone. Among these, the polymethoxyflavone may be sudatin or demethoxysudatin.

Alternatively, the flavonoid may be a flavanone, hesperetin. Moreover, the flavonoid may contain quercetin or anthocyanidin.

Flavonoid glycosides are hydrophilic compounds having a structure in which flavonoids and sugars are linked by glycoside bonds as described above. The sugar that constitutes the flavonoid glycoside is not particularly limited, and examples thereof include known sugars that can form a glycoside by binding to the flavonoid described above through a glycoside bond.

The extract 21 may include one or more of the above-mentioned flavonoids.

The extraction fee 21 may include components other than flavonoids. Examples of the other component include sugar, cellulose, acid and the like. The content of flavonoids in the extract 21 is preferably 5 mass ppm or more, and more preferably 10 to 1000 mass ppm, based on the total solid content of the extract 21.

Specifically, as the extraction material 21, plants, seaweed flowers, leaves, roots, stems, fruits, seeds and the like can be used. In particular, since the pericarp contains a large amount of polymethoxyflavone and glycosides thereof, the squeezed residue of citrus fruits can be preferably used. Further, the extraction material 21 may be a dry powder obtained from citrus fruits, or may be a dry powder obtained from citrus peels. Examples of citrus fruits include Sudachi, Wenshu mandarin orange, ponkan, and shikwasa. The citrus fruits may be polymethoxyflavone such as sudatin and demethoxysudatin, and sudachi containing a large amount of glycosides thereof. The dry powder can be obtained by crushing plants, seaweed flowers, leaves, roots, stems, fruits, seeds and the like with a mixer or the like. The particle size of the dry powder may be 0.1 to 5 mm in terms of major axis diameter from the viewpoint of extraction efficiency of flavonoids.

The extraction fee 21 may be previously subjected to acid treatment. In this case, the manufacturing method of the present embodiment may further include an acid treatment step of acidizing the extractant before the extraction step. The acid treatment can be performed by reacting the extract with an acid such as hydrochloric acid or sulfuric acid. The reaction is not particularly limited, but can be carried out, for example, at a temperature of 60 to 120 ° C. for 0.5 to 3 hours. By performing the acid treatment step, at least a part of the flavonoid glycosides contained in the extract can be decomposed into flavonoids, and the extraction efficiency of flavonoids in the extraction step can be improved. In addition, when the acid treatment step is performed, even if the decomposition step described below is not performed after the vaporization step, the flavonoid glycoside has been decomposed into flavonoids in advance, so that an extract with a high concentration of flavonoids can be obtained. . When the acid treatment is performed, it is preferable to neutralize the acid with a base such as an aqueous sodium hydroxide solution before the extraction step.

(Extractant)
The extractant 22 may be made of only liquefied dimethyl ether or may contain a solvent other than liquefied dimethyl ether. Examples of the other solvent include water, ethanol, supercritical carbon dioxide, isobutane, isobutene, normal butane, propane and the like. From the viewpoint of extraction efficiency of flavonoids in the extraction step, the content of liquefied dimethyl ether in the extractant 22 is preferably 50% by mass or more, and 90% by mass or more, based on the total amount of the extractant 22. Is more preferable.

[Vaporization process]
In the vaporizing step, at least a part of the extractant 22 is vaporized and removed from the extract liquid 24 obtained in the extracting step to obtain an extract 26 containing flavonoids. In the vaporization step, the extraction liquid 24 transferred to the vaporization tank 13 is brought to normal pressure or heated to a temperature of 20 ° C. or higher to vaporize at least a part of the extractant 22 and exhaust gas provided in the vaporization tank 13. By opening and closing the valve 34 from the mouth, the gas 25 is discharged to the outside of the vaporization tank 13. As a result, the extract 26 containing flavonoids from which at least a part of the extraction agent 22 is removed can be obtained. The obtained extract 26 can be discharged from the discharge port provided in the vaporization tank 13 by opening and closing the valve 35 and collected in the container 14.

The extract 26 may be a liquid or a solid, but it is preferable to collect it in a liquid state from the viewpoint of work efficiency.

In the vaporization process, it is not necessary to remove the entire amount of the extractant 22, and a part may remain. By leaving a part of the extractant 22, it becomes easy to collect the extract 26 in a liquid state. When the extract 26 is recovered in a liquid state, it is preferable that the extractant 22 contains a solvent that is liquid at room temperature and normal pressure in addition to liquefied dimethyl ether. Solvents other than liquefied dimethyl ether include alcohols (ethanol and the like), alkanes (hexane and the like), esters (ethyl acetate and the like), water and the like.

When the extract 26 obtained through the vaporization step is recovered in a liquid state, it can be dried to be a solid after the recovery. Thereby, the solid extract 26 can be obtained.

[Disassembly process]
The production method of the present embodiment may further include a decomposition step of decomposing at least a part of the flavonoid glycosides in the obtained extract 26 into flavonoids after the vaporization step. As a result, a decomposition product containing a high concentration of flavonoid can be obtained. Degradation of flavonoid glycosides can be carried out in the same manner as the acid treatment of the extract described above. The extract 26 to be subjected to the acid treatment is preferably in a solid state.

[Purification process]
When flavonoid is the target substance, the production method of the present embodiment may further include a purification step of extracting and purifying flavonoid from the extract 26 or the degradation product after the vaporization step or after the decomposition step. .. The extract 26 or the decomposed product may contain, in addition to flavonoids, sugar, flavonoid glycosides, citric acid, limonene, and the like. Here, flavonoids are hydrophobic, while sugars, flavonoid glycosides, and citric acid are hydrophilic. Therefore, the flavonoid can be further extracted and purified by introducing the extract 26 or the decomposed product into an organic solvent such as ethanol, ethyl acetate, hexane and a mixture thereof, and removing the insoluble matter by filtration or the like.

The flavonoids can be efficiently produced at a high concentration by the production method of the present embodiment described above. The flavonoids produced by the production method of the present embodiment may be polymethoxyflavone, and may be sudatin and / or demethoxysudatin. The production method of the present embodiment is suitable for producing polymethoxyflavone, particularly sudatinine and demethoxysudacitin, and the concentration thereof can be greatly improved.

The preferred embodiments of the present disclosure have been described above in detail, but the present disclosure is not limited to the above embodiments. For example, in the manufacturing method of the present disclosure, it is necessary to perform each step continuously or intermittently by using the liquefied gas extraction device to which the liquefied gas cylinder 11, the extraction tank 12, and the vaporization tank 13 shown in FIG. 1 are connected. Alternatively, each step may be performed independently. Further, the filtration filter 23 may not be provided in the extraction tank 12, and the contact between the extraction material 21 and the extraction material 22 and the separation of the extraction material 21 and the extraction material 22 are not performed in the same tank but separately. You can go.

Hereinafter, the present disclosure will be described more specifically based on Examples and Comparative Examples, but the present disclosure is not limited to the following Examples.

(Example 1)
Flavonoids were extracted from the extract by the method shown in FIG. First, the frozen Sudachi juice residue containing the Sudachi peel after squeezing Sudachi juice was crushed with a mixer to obtain a crushed frozen Sudachi juice residue having a particle size (major axis diameter) of about 0.1 to 5 mm. 30 g of the obtained crushed frozen Sudachi juice residue is put into the extraction tank of the liquefied gas extraction device as an extractant, and 60 ml of liquefied DME (dimethyl ether) is injected as an extractant from the liquefied gas cylinder into the extraction tank to extract 5 extraction tanks. Shaking for minutes, the flavonoids (extractant) in the extract were extracted into the extractant (extraction step). The extraction step was carried out under a room temperature (25 ° C.) environment in a state where the liquefied DME vaporized in the closed extraction tank and reached the saturated vapor pressure (about 0.6 MPa). Then, the extract-containing liquefied DME (extract) was transferred from the extraction tank to the vaporization tank through a filter having a pore size of 1 μm. By keeping the inside of the vaporization tank at normal pressure, DME was vaporized and removed in the vaporization tank (vaporization step), and the liquid extract was separated and collected. The liquid extract contains water contained in the pericarp of Sudachi. The operation from the injection of liquefied DME to the extraction tank containing the crushed frozen Sudachi juice residue to the separation and recovery of the liquid extract was repeated 3 times, and the collected liquid extracts for 3 times were combined to extract Sudachi peel. A liquid was obtained.

The obtained Sudachi peel skin extract was dried in a vacuum dryer at 60 ° C for 300 minutes to obtain a solid Sudachi peel skin extract. Then, 0.1 g of the extract of Sudachi pericarp was immersed in 2 g of 1N hydrochloric acid, and the flavonoid glycosides in the extract were decomposed into flavonoids by reacting at 120 ° C. for 1 hour. Neutralized. The neutralized solution was dried in a vacuum dryer at 60 ° C. for 300 minutes to obtain a solid decomposition product of glycosides of Sudachi pericarp.

(Example 2)
A crushed frozen Sudachi juice residue was obtained in the same manner as in Example 1. 100 g of the obtained crushed frozen Sudachi juice residue was immersed in 300 g of 1N hydrochloric acid and reacted at 120 ° C. for 1 hour to decompose the flavonoid glycosides in the crushed frozen Sudachi juice residue into flavonoids, and then 1N hydroxylated. The solution was neutralized with an aqueous sodium solution (acid treatment step). The neutralized solution was dried in a vacuum dryer at 60 ° C. for 300 minutes to obtain an acid-treated ground Sudachi juice residue. A solid Sudachi peel extract and a solid Sudachi peel extract glycoside decomposition product were obtained in the same manner as in Example 1 except that 30 g of the acid-treated ground Sudachi juice residue was used as the extractant.

(Comparative Example 1)
A crushed frozen Sudachi juice residue was obtained in the same manner as in Example 1. 100 g of the obtained crushed frozen Sudachi juice residue was weighed into a 1 L flask as an extractant, 120 g of ethanol and 280 g of pure water were added thereto as an extractant, and the liquid temperature was 60 ° C. for 5 hours in an oil bath. Extraction was performed, and flavonoids (extractant) in the extract were extracted as an extractant. Then, the solid matter was separated using a PTFE filter having a pore size of 1 μm to obtain a water / ethanol extract. The obtained water / ethanol extract was dried at 60 ° C. for 300 minutes in a vacuum dryer to obtain a solid Sudachi peel extract (water / ethanol extract). The obtained Sudachi pericarp extract was subjected to acid treatment (glycoside decomposition) in the same manner as in Example 1 to obtain a solid Sudachi pericarp extract glycoside decomposition product.

(Comparative example 2)
A solid extract of Sudachi pericarp (water extract) was obtained in the same manner as in Comparative Example 1 except that the extractant was changed to 400 g of pure water. In Comparative Example 2, the acid treatment (degradation of glycoside) of the Sudachi peel extract was not performed.

(Comparative example 3)
A solid Sudachi peel extract (isobutane extract) was obtained in the same manner as in Example 1 except that liquefied isobutane was used as the extractant. The extraction step was carried out under a room temperature (25 ° C.) environment in a state where the liquefied isobutane was vaporized in the closed extraction tank to reach the saturated vapor pressure. In Comparative Example 3, the extract was a syrup-like semi-solid state, and 0.1 g or more of the extract could not be obtained. Therefore, acid treatment (glycoside decomposition) of the Sudachi peel extract was not performed. Further, since the extract was in the above state, there was almost no waste liquid described later.

<Measurement of flavonoids concentration>
The flavonoids in the Sudachi peel extract obtained in each Example and Comparative Example were measured by the following method. First, 1 g of a solid Sudachi pericarp extract was dissolved / dispersed in 100 g of ethanol and filtered through a PTFE filter having a pore size of 0.1 μm to obtain an ethanol solution. The components of this ethanol solution were analyzed by high performance liquid chromatography (HPLC). A calibration curve was prepared using a commercially available standard purified sample of the target substance as a standard substance, and the concentration of the target substance in the extract of perch of Sudachi was roughly calculated using the calibration curve. As the HPLC apparatus, "Chrome Master" manufactured by Hitachi High-Tech was used. The target substances for measuring the concentrations were sudatin, hesperidin, and demethoxysudatin. Further, in Examples 1 and 2 and Comparative Example 1, the concentration of sudatinine glycoside was also measured. The concentration of sudachitin glycoside was determined from the difference between the sudachitin concentration of the acid-treated sudachi peel extract glycoside decomposition product and the sudachitin concentration of the sudachi peel extract. The results are summarized in Table 1.

<Measurement of waste liquid (waste solvent)>
The amount of the solvent (water and ethanol) that was liquid at room temperature and pressure during the vaporization step and the subsequent drying was measured as the amount of waste liquid. The measurement results are shown in Table 1.

As shown in Table 1, in Examples 1 and 2, flavonoids having different structures, sudacitin, hesperidin, and demethoxysudacitin were extracted at a higher concentration than those of Comparative Examples 1 to 3, and sudacithin glycosides were also extracted. Was done. Further, in Examples 1 and 2, the amount of waste liquid was significantly reduced as compared with Comparative Examples 1 and 2. In Comparative Example 3, extraction was performed with liquefied isobutane, which is a liquefied gas, but unlike the case where liquefied DME was used, flavonoids could not be extracted.

11 ... Liquefied gas cylinder, 12 ... Extraction tank, 13 ... Vaporization tank, 14 ... Container, 21 ... Extraction material, 22 ... Extraction agent, 23 ... Filtration filter, 24 ... Extraction liquid, 25 ... Gas, 26 ... Extract, 31, 32, 33, 34, 35 ... Valves.

Claims

An extraction step of extracting an extract containing flavonoids by contacting an extract containing flavonoids with an extract containing liquefied dimethyl ether,
A vaporization step of vaporizing and removing at least a part of the extractant from the extract to obtain an extract containing flavonoids,
A method for producing flavonoids having:
The production method according to claim 1, wherein the flavonoids include at least one of sudatin and glycoside.
The manufacturing method according to claim 1 or 2, wherein the lottery material includes Sudachi pericarp.
The manufacturing method according to any one of claims 1 to 3, further comprising an acid treatment step of treating the extract with an acid before the extraction step.
The production method according to any one of claims 1 to 4, further comprising a decomposition step of decomposing at least a part of the flavonoid glycosides in the extract into flavonoids after the vaporization step.