JP6333057B2 - Tyrosinase inhibitor and method for producing the same - Google Patents

Description

  The present invention relates to a tyrosinase inhibitor using plant rose as a raw material and a method for producing the same.

[Plant-derived Maillard reaction inhibitor]
As is well known, the Maillard reaction is a “non-enzymatic reaction” that produces a brown substance that occurs when reducing sugars and amino compounds (amino acids, peptides, proteins, etc.) are heated.
As this reaction progresses, it is said that the quality of foods will decrease. Therefore, in order to suppress the deterioration of food quality, it is desired to develop a substance that effectively suppresses the Maillard reaction.
In addition, the Maillard reaction occurs in the living body, and as a result, it is considered to be a cause of aging, blackening and browning of the skin, and a cause of diabetic complications. Therefore, the development of an effective and safe Maillard reaction inhibitor is desired.
Under such circumstances, various attempts have been made to obtain Maillard reaction inhibitory substances (or Maillard reaction inhibitors) from plants.

[Plant-derived tyrosinase inhibitor]
It is said that spots and buckwheat are caused by the production of melanin pigment. Melanin pigments are biosynthesized via dopa, dopaquinone, etc., with the action of “tyrosinase, an enzyme”, using tyrosine as a substrate. Therefore, if the action of tyrosinase can be inhibited, the production of melanin pigment can be suppressed, and a whitening effect can be expected.

[Maillard reaction inhibitor and tyrosinase inhibitor]
As described above, although the Maillard reaction inhibitor and the tyrosinase inhibitor have a common purpose for “whitening”, the former is “suppression of non-enzymatic reaction” and the latter is “the action of the enzyme tyrosinase. "Inhibition" and the mechanism of action of both is completely different.
Therefore, in order to reliably achieve the objective such as whitening, it is essential not only to suppress the Maillard reaction but also to inhibit or suppress the action of the enzyme tyrosinase.

  However, the present condition is that there has not been found a satisfactory “functional agent” that shows excellent action in both Maillard reaction inhibitor and tyrosinase inhibitor and has no concern about toxicity.

[Especially for tyrosinase inhibitors]
-1-
There is a great deal of literature on Maillard reaction inhibitors that use plants as raw materials, and there are quite a few references on tyrosinase inhibitors that use plants as raw materials, but they are effective for both Maillard reaction inhibitors and tyrosinase inhibitors. There are few literatures on functional agents that exhibit a good action, and there are few literatures on functional agents that show a reliable and excellent action for both of them.
In this case, even if it is acceptable as a Maillard reaction inhibitor, it seems that the action as a tyrosinase inhibitor is often insufficient. In connection with the present invention, the present inventors have conducted experiments using various plants, and although there are many plant extracts having Maillard reaction inhibitory activity, there is a substance having an effective inhibitory activity against tyrosinase. Some plants are only roses, and even if they are roses, it is difficult to say that a simple solvent extract is effective as a tyrosinase inhibitor.

-2-
If the action as a tyrosinase inhibitor is excellent, even if the action as a Maillard reaction inhibitor is small, it is possible to obtain both functions by administering or ingesting other Maillard reaction inhibitors in combination. It will be possible.
However, the present situation is that finding out what works as the latter tyrosinase inhibitor out of the action as the Maillard reaction inhibitor and the action as the tyrosinase inhibitor is a barrier.

-3-
Hereinafter, Patent Document 1 and Patent Document 2 as examples of documents related to plant-derived tyrosinase inhibitors will be described in detail.

(Patent Document 1)
-1-
Claims 1 to 3 of JP-T-2012-502022 (Patent Document 1) show a skin whitening composition containing “an extract of cypress tree, a solvent fraction thereof, or a compound separated and purified from it” as an active ingredient. Has been.
The solvent for extraction at this time is water, a C1-C4 lower alcohol or a mixed solvent thereof, and the solvent for fractionation is hexane, methylene chloride, ethyl acetate, or butanol (claim 12).

-2-
In claim 4, there is shown a skin whitening composition containing as an active ingredient one or more compounds selected from the group consisting of compounds represented by chemical formulas 1 to 10.
In claim 11, the compound represented by the chemical formula 10 is shown.

-3-
FIG. 28, FIG. 35, and FIG. 38 of this document show graphs showing tyrosinase inhibitory activity.

-4-
The description of the above-mentioned compound in the column of the detailed description of the invention of Patent Document 1 is inconsistent with the description of Claims 4 and 9 as described later in the section -6. Many things make it difficult to grasp the invention of Patent Document 1.
If the chemical formula is displayed correctly, there are many cases where the compound name is wrong, and if the compound name is correct, there are many cases where the chemical formula is wrong. Since it cannot be determined which of the chemical formula and the compound name is correct and which is wrong (or both are wrong), the invention of Patent Document 1 cannot be grasped.

-5-
By way of example, compound 3 of formula 3 in claim 4 and paragraph 0015, according to paragraph 0066, “compound 3 is kaempferol 3-O-rhamnopyranoside”, paragraphs 0097 and 0100 according to experimental example 5 Is confused because “Compound 9 has been identified as Canocgiol of formula 9” in paragraph 0078.
Moreover, compound 3 of formula 3 is “quercetin-3-O-arabinofuranoside” if it is assumed that formula 3 is correct.
Therefore, in patent document 1, as a compound of Chemical Formula 3,
Quenferol 3-O-rhamnopyranoside Canacgiol Quercetin-3-O-arabinofuranoside is a completely different substance.

-6
The description or explanation in Patent Document 1 is also confusing in the following points.
A: Compound of Chemical Formula 1 in Claim 4 and Paragraph 0016 Description of Paragraphs 0062-0063: ○ Quercetin Description of Paragraph 0095: ▲ Lucidone (Compound 1)
Description of paragraph 0098: ▲ lucidone (compound 1)
(Note: If chemical formula 1 is correct, the ○ mark is correct and the ▲ mark is incorrect.)
A: Compound of Chemical Formula 2 in Claim 4 and Paragraph 0017 Description of Paragraphs 0064-0065: ○ Quercitrin Description of Paragraph 0096: Methyl Linderone (Compound 2)
Description of paragraph 0099: ▲ Methyl Linderone (Compound 2)
(Note: If chemical formula 2 is correct, the ○ mark is correct and the ▲ mark is incorrect.)
C: Compound of Chemical Formula 3 in Claim 4 and Paragraph 0018 Description of Paragraphs 0066 to 0067: Kaempferol 3-O-rhamnopyranoside Paragraph 0097 Description: ▲ Kanakugiol (Compound 3)
Description of paragraph 0100: ▲ canacguiol (compound 3)
(Note: If chemical formula 3 is correct, the ▲ mark will be wrong.)
D: Compound of Chemical Formula 4 in Claim 4 and Paragraph 0019 Explanation of Paragraphs 0068-0069: Quercetin 3-O-arabinofuranoside (Note: If Chemical Formula 4 is correct, the ▲ mark is incorrect.)

−7−
In any case, in Patent Document 1, an “extract or a solvent fraction thereof” is obtained from the plant Anagony, so that the component difficult to extract with a solvent is the component acquired in Patent Document 1. There is little or not.

(Patent Document 2)
Japanese Patent Application Laid-Open No. 2010-18530 (Patent Document 2) relates to “a method for extracting a tyrosinase inhibitory active agent using an apple secondary residue or tishimazasa as a raw material and its tyrosinase inhibitory active composition”.
In addition, according to the description in paragraph 0010, “apple secondary residue” in Patent Document 2 is “squeezed residue generated when squeezing apple juice”.
About the content of invention of Claims 1-8 of patent document 2, since it is hard to understand just by quoting description of a claim, it will show a summary in the form of Table 1-Table 8 as follows.

Special table 2012-502022 JP2010-018530

(Regarding Patent Document 1)
-1-
Patent Document 1 (Japanese Translation of PCT International Publication No. 2012-502022) describes a composition for whitening the skin containing an extract of an Anthracnaceae (or a solvent fraction thereof) as an active ingredient. These extracts and fractions There is also a description that has a tyrosinase inhibitory activity.
There is also a description that the active ingredient is one or more compounds selected from the group consisting of the compounds represented by Chemical Formulas 1 to 10.
FIG. 28 shows a graph showing the cell tyrosinase inhibitory activity of various concentrations of the ethanol extract of Anemone and its solvent fraction. FIG. 35 shows a graph showing the tyrosinase inhibitory activity represented by lucidone.

-2-
However, Patent Document 1 does not describe the following points that are characteristic constituent features of the tyrosinase inhibitor of the present invention.
I: Use roses as plant material.
B: “Low-temperature vacuum distillation residue (R) itself” or its “Extract with solvent for extraction from low-temperature vacuum distillation residue (R) (E)” as an active ingredient.

(Regarding Patent Document 2)
Patent Document 2 (Japanese Patent Application Laid-Open No. 2010-018530) relates to an apple secondary residue and ticasamasa, and does not describe the following points that are special characteristic constituents in the tyrosinase inhibitor of the present invention.
I: Use roses as plant material.
B: “Low-temperature vacuum distillation residue (R) itself” or its “Extract with solvent for extraction from low-temperature vacuum distillation residue (R) (E)” as an active ingredient.

(Object of the present invention)
Under such a background, the present invention provides:
(1) To provide a method for obtaining a tyrosinase inhibitor having an excellent tyrosinase inhibitory effect by using rose as a plant raw material and taking special production means; and
(2) providing a tyrosinase inhibitor that can be obtained in such a manner;
It is intended.

The tyrosinase inhibitor of the present invention is
A component distilled when a water-containing plant raw material is subjected to dry distillation under low temperature and reduced pressure conditions using a dry distillation apparatus equipped with a pressure reduction mechanism is referred to as “low temperature reduced pressure dry distillation fraction (d)” and the low temperature reduced pressure dry distillation thereof. When a dry solid or solid residue (that is, “can residue”) remaining in the carbonization apparatus after operation is referred to as “low temperature vacuum distillation residue (R)”,
The plant material is a rose, and
The above-mentioned “low-temperature vacuum distillation residue (R) itself” or its “extract with a solvent for extraction from low-temperature vacuum distillation residue (R) (E)” as an active ingredient,
It is characterized by.

  Here, the low-temperature vacuum distillation operation is typically a reduced pressure of a plant raw material in a water-containing state having a water content of 90% by weight or less under a low-temperature condition of 60 to 20 ° C. and a gauge pressure of −88 kPa or less. This is a carbonization operation performed under conditions.

  A typical example of the extraction solvent is ethanol water having an ethanol concentration of 25 to 75% by volume (that is, “mixed solution of 25 to 75 volumes of ethanol and 75 to 25 volumes of water”).

The method for producing the tyrosinase inhibitor of the present invention comprises:
Dry distillation of the water-containing plant material under low temperature and reduced pressure conditions using a dry distillation apparatus equipped with a vacuum mechanism;
The plant material is a rose,
Acquire “low temperature vacuum distillation residue (R)” which is a solid or solid residue (ie “can residue”) in the dry state remaining in the carbonization apparatus after the low temperature vacuum distillation operation, or further Performing extraction with a solvent for extraction from the "vacuum dry distillation residue (R)" to obtain an extract (E);
It is characterized by.

-1-
Regarding the high-temperature vacuum-distilled product that is distilled when the plant raw material is dry-distilled under “high-temperature vacuum conditions (more specifically, a vacuum condition of 100 mmHg or less under a high temperature of about 120 to 350 ° C.)” Products (deodorants and antioxidants), such as those containing tea leaves and other plant vacuum-distilled products as active ingredients or added as active ingredients, are widely available on the market, including drug stores. It is well known because there are many documents.

-2-
However, it is well known in the art about “dry-distilling plant raw materials under low temperature and reduced pressure conditions (typically under a reduced pressure condition of −88 kPa or less at a gauge pressure of 60 to 20 ° C.)”. It seems that it is not.

-3-
When the vacuum distillation of the plant material is performed, the active ingredient contained in the plant material is transferred into a vacuum distillation solution (distillate) which is a distillate, whether it is a high temperature vacuum distillation or a low temperature vacuum distillation. The active ingredient produced by the decomposition or modification of the components contained in the plant raw material during the vacuum distillation operation is also transferred to the “vacuum dry distillation liquid (distilled portion)”. Thus, in order to achieve the transfer of the active ingredient, vacuum distillation is performed.

-4-
However, the present inventors have tested the “distilled fraction” that is distilled during low-temperature vacuum dry distillation using rose as a plant raw material. As a result, the tyrosinase inhibition rate is extremely small or close to zero. Oops.

-5-
As described above, since the “low-temperature vacuum dry distillation liquid (distillate)” was completely disappointing, the present inventors were discouraged and colored “lightly brown” together with the “low-temperature vacuum dry distillation liquid”. The solid or solid “residue after low-temperature vacuum distillation (residue can)” was also discarded together.
However, probably because rose (damask rose in this test) was used as a raw material, the “residue” had a good scent unlike the appearance. Therefore, the degree of expectation was zero, but when the “residues” were extracted using “ethanol”, “50% by volume ethanol water”, and “water” as solvents for disposal before disposal, When “50% by volume ethanol water” was used, a tyrosinase inhibition rate that was good enough to think that it was a mistake was obtained.

-6
In other words, “active ingredients” having a tyrosinase inhibitory action (function) in roses and “active ingredients” generated by decomposition or denaturation of components contained in roses during reduced-pressure distillation operations are distilled during low-temperature vacuum distillation. Almost no shift during the distillate, (b) increased to the residue side during low temperature vacuum distillation, or (b) increased due to changes in specific components or reaction of various components on the residue side .

−7−
In addition, with regard to “frozen storage roses” that have not undergone the low-temperature reduced-pressure dry distillation step, “ethanol”, “50% by volume ethanol water”, and “water” are used as solvents for the frozen flower parts as they are. When the tyrosinase inhibition rate was measured, the sample extracted with “50 vol% ethanol water” was 1/3 as compared with the sample derived from the above “residue after low-temperature vacuum distillation”. An inhibition rate of about 1/4 was obtained.

-8-
This means that “rose” used as a raw material has a “substance” compared to other plants in terms of tyrosinase inhibitory action (function). It can be said that the stage was greatly extended and blossomed.

-9-
In general, the residue after performing extraction, distillation, steam distillation, dry distillation, etc. on plant raw materials is so-called waste, and it is actually difficult to dispose of it in terms of its disposal and disposal costs. It is.
However, the specific “low-temperature vacuum distillation residue” in the present invention is an unexpected development that it is a “residue” and is a treasure house where a tyrosinase inhibitor having an excellent tyrosinase inhibitory action (function) can be obtained. Was planned.

It is a HPLC chromatogram (detection wavelength is 280 nm) of "extract (E)" when "residue (R)" in an Example is extracted with 50 volume% ethanol water. It is a HPLC chromatogram (detection wavelength is 280 nm) of "extract (E)" when "residue (R)" in an Example is extracted with ethanol. It is a HPLC chromatogram (detection wavelength is 280 nm) of "extract (E)" when "residue (R)" in an Example is extracted with water.

(Plant raw material)
-1-
In the present invention, roses (of the family Rosaceae) are used as plant materials. In the object of the present invention to obtain a tyrosinase inhibitor, as described above in the section “Effects of the Invention”, when rose is used as a raw material, a unique action is achieved and excellent results are obtained. is there.
Among roses, damask rose cultivated (or imported) mainly for the purpose of obtaining rose water or rose oil is particularly suitable. By the way, Damascus is one of the biggest producers in Bulgaria, but it is also cultivated in Japan.

-2-
Since roses are cultivated for the purpose of flower parts, there is a temporal restriction that they can be obtained from producers during the flowering season.
However, for the purpose of the present invention, even if it is a frozen preserved product, there is almost no difference in action and effect from that collected at the time of flowering. Even if it exists, since it has been confirmed by a test in the laboratory that the decrease in the effect is small, it is usual to use a frozen storage product that can secure the material stably.

-3-
As for the rose part, any of whole plant, flower part, leaf part and rhizome may be used.

(Low temperature vacuum distillation)
In the present invention, a water-containing rose is subjected to dry distillation under low temperature and reduced pressure using a dry distillation apparatus equipped with a pressure reduction mechanism.
The desired product can be industrially efficiently obtained by low-temperature vacuum distillation of roses under the following conditions.

(Condition of moisture content during low-temperature vacuum distillation)
It is preferable that the moisture content of the rose subjected to low temperature vacuum distillation is limited to 90% by weight or less. The lower limit of the moisture content is suitably up to about 50% by weight. A preferable range of the moisture content of the rose subjected to low temperature vacuum distillation is 60 to 85% by weight, and a more preferable range is 70 to 83% by weight.
This is because the target active ingredient is maximized on the residue (R) side after the low-temperature vacuum dry distillation when the above moisture content rose is subjected to low-temperature vacuum dry distillation.

(Temperature conditions for low temperature vacuum distillation)
As a temperature condition when performing low-temperature vacuum distillation, a low temperature in the range of about 60 to 20 ° C is appropriate, a more preferable range is 55 to 25 ° C, and a further preferable range is 50 to 30 ° C.
When vacuum distillation is performed under conditions where the temperature exceeds 60 ° C., the active ingredient in the extract (E) is extracted even if the solid or solid residue (R) remaining in the distillation apparatus is extracted with a solvent. In addition to the reduced amount, the tyrosinase inhibitory action (function) tends to become insufficient when the obtained extract (E) is used.

(Pressure conditions for low temperature vacuum distillation)
The pressure condition (depressurization condition) when performing low temperature vacuum distillation is preferably -88 kPa or less (-660 mmHg or less), usually -96 to -100 kPa (-720 to -750 mmHg) in gauge pressure notation.
In absolute pressure notation, it is preferably 13.3 kPa or less (100 mmHg or less), usually 1.3 to 5.3 kPa (10 to 40 mmHg).
If the degree of decompression is less than the above range (when the degree of decompression is insufficient), it will take a long time for dry distillation. On the other hand, too large a degree of decompression has restrictions on the vacuum equipment, so both are industrial. Lack of sex.

(One of the objects of the present invention / low temperature vacuum distillation residue (R))
-1-
“Low-temperature vacuum distillation residue (R)”, which is a solid or solid residue remaining in the carbonization apparatus when the rose is subjected to low-temperature vacuum distillation as described above, is one of the objects of the present invention. .
-2-
However, since this “low-temperature vacuum dry distillation residue (R) itself” contains many contaminants such as plant tissues, depending on how it is used (for example, when used for cosmetics), it may be negative in terms of usability. There is. In addition, since the ratio of the active ingredient is also limited, the tyrosinase inhibitory action (function) is also limited.
-3-
Therefore, this “low-temperature vacuum distillation residue (R)” is often used as a raw material or an intermediate material of “extract (E”) described below.
The purchaser of this “low-temperature vacuum distillation residue (R)” uses the residue (R) to perform “extraction with a solvent” to be described later to form an extract (E), and uses the extract (E). Thus, secondary products and tertiary products are manufactured and sold, and the extract (E) is further sold to a third party.

(Another product of the present invention / Extract (E) with a solvent for extraction from low-temperature vacuum distillation residue (R))
-1-
And the above-mentioned "extract (E) by the solvent for extraction from low-temperature vacuum distillation residue (R)" is also one of the main objectives of this invention.
-2-
Here, as the “solvent for extraction”, various solvents including water, ethanol, and a water-ethanol mixed solvent can be used.
-3-
However, the extract (E) extracted with ethanol water having an ethanol concentration of 25 to 75% by volume (preferably 30 to 70% by volume) exhibits particularly excellent tyrosinase inhibitory action (function). It is optimal for the purposes of the present invention to use ethanol water of composition as the extraction solvent. (As a reminder, “ethanol water having an ethanol concentration of 25 to 75% by volume” is a mixed solution of 25 to 75 volumes of ethanol and 75 to 25 volumes of water.)
This is because, when ethanol water having an ethanol concentration of 25 to 75% by volume is used as an extraction solvent, the active ingredient contained in the low-temperature vacuum distillation residue (R) is concentrated in the extract (E). Because. (In addition, ethanol water having such a solvent composition is advantageous in that it is a non-toxic and non-flammable solvent and is inflammable.)
-4-
Here, the active ingredient is a component that exerts a tyrosinase inhibitory action (function), and some of the compound names are known, but individual compounds are isolated and, if necessary, those individual compounds. Rather than using a mixture of, for example, “the component extracted with ethanol water selected from within the range of 25 to 75% by volume of ethanol” (in other words, the celestial composition contained in the rose as it is It is “practical” and “purposed” (that fits a certain purpose).

(About low-temperature vacuum distillation distillation (d))
-1-
On the other hand, it has been confirmed that the “low temperature vacuum distillation distillation fraction (d)” in the above low temperature vacuum distillation has substantially no tyrosinase inhibitory action (function) or is small even if the action (function) is exhibited. Therefore, it is not the “tyrosinase inhibitor” itself which is the object of the present invention.
However, since it has been confirmed that this “low-temperature vacuum distillation distillation (d)” has a corresponding function as another purpose, for example, a Maillard reaction suppression function or an antioxidant function, such a function is required. Can be used.
-2-
Therefore, “residue (R)” and “distillation (d)” obtained by low-temperature vacuum distillation of the above-mentioned rose, and “extract (E)” obtained from the former “residue (R)” 3 Anyone will be used effectively.

(Abbreviation of the relationship between the three)
In the above description, (low temperature vacuum distillation) residue (R) (that is, “can residue”),
-Extract (E) (with solvent for extraction from the low temperature vacuum distillation residue (R)),
・ (Low temperature vacuum distillation) Distillate (d)
The terms are easily confused, and the relationship is shown in the following “Table 0”.
The “residue (R)” marked with “◎” and the “extract (E)” marked with “◎” obtained from the residue (R) are the direct objects of the present invention.

  Next, the present invention will be further described with reference to examples.

[Plant preparation]
-1-
The following 1 to 14 were prepared as plant materials.
1: Rose from 2011 (2011) 1 / Damascus (Rose with gouge, moisture is around 80% by weight)
2: Roses produced in 2012 (Heisei 24) 2 / Damascus (Rose flowers with gouge, moisture is about 80% by weight; production area is the same as rose 1)
3: Shikuwasa (squeezed rice cake when manufacturing the juice of the fruit, moisture is about 80% by weight)
4: Bellows (The squeezer used to produce the juice of the fruit, the water content is about 80% by weight)
5: Ginger (chip at the time of commercialization, moisture is about 60% by weight)
6: mandarin oranges (squeezed straw at the time of manufacturing the juice of the fruit, moisture is about 80% by weight)
7: Turmeric (cutting chips at the time of commercialization, moisture is 60% by weight or less)
8: Ghetto (boiled rice cake when making moon peach water, moisture is about 80% by weight)
9: Lavender (flower after the ornamental flowering period, moisture is about 80% by weight)
10: Sudachi (squeezed rice cake when producing the juice of the fruit, moisture is about 80% by weight)
11: Yuzu (It is a squeezed koji at the time of juice production of the fruit, the water is about 80% by weight)
12: Green onion (the part that did not become a product, moisture is about 80% by weight)
13: Casablanca (flowers at flowering, moisture is about 80% by weight)
14: Frozen rose (damask rose)

[Low temperature vacuum distillation operation]
-1-
About 30 kg of each of the 1 to 13 plant raw materials prepared above was put into a tank of a tank-shaped dry distillation apparatus equipped with a pressure reducing mechanism, and under stirring, low temperature (35 to 40 ° C.) and reduced pressure ( Dry distillation was performed under the conditions of -98 kPa (-735 mmHg) in gauge pressure and 3.3 kPa (25 mmHg) in absolute pressure.
The time required for dry distillation is approximately 8 hours, excluding the time required for charging the raw materials and taking out the residues in the tank, and the time required for decompression and the time for returning to normal pressure.
-2-
The liquid distilled from the carbonization apparatus is “low-temperature vacuum distillation distillation (d)” (abbreviated as “distillation (d)”).
-3-
On the other hand, the solid residue remaining in the carbonization apparatus (dry distillation kettle) after the above low-temperature vacuum distillation operation is the “low-temperature vacuum distillation residue (R)” (abbreviated as “residue (R)”) in the present invention.
This residue (R) is a flake-like lump or a powdery form in which the lump is broken.
In addition, since the water | moisture content contained in the plant raw material at the time of charging enters into the reduced pressure dry distillation liquid side (low temperature reduced pressure dry distillation distillate (d) side), this residue (R) has a water content of about 2 to 5% by weight or less Virtually dry body. (By the way, even if the residue (R) is a flaky lump, it can be easily crushed into particles or powder.)

[Test solution]
-1-
Since the “distilled fraction (d)” obtained by the above low-temperature vacuum drying operation was a liquid, it was used as it was for the test of the tyrosinase inhibition rate.
-2-
About the powdery “residue (R)” obtained by the above low-temperature vacuum distillation operation, each 5.0 g was weighed, and 50 mL of “ethanol” and 50 mL of “50% by volume ethanol water” as an extraction solvent. , 50 mL of “water” was added, and the mixture was extracted by stirring at room temperature for 4 hours. After extraction, the solid content was separated and removed by filtering through filter paper.
In addition, since the water extraction filtrate was turbid, filtration was further performed using a 0.45 μm filter. Since some dry powder extracts were cloudy and could not be tested when used in a test without dilution, the sample solution was “diluted 10 times” with the solvent used for extraction.

[test]
(1) 1.8 mL of 30 mM phosphate buffer (pH 6.8), 1.0 mL of a 1.66 mM L-tyrosine solution, and 0.1 mL of each sample solution are mixed and preliminarily added for 5 minutes in a 37 ° C. constant temperature bath. Warm was done.
(2) After adding 0.1 mL of tyrosinase solution, the mixture was stirred and again heated in a thermostatic bath at 37 ° C. for 10 minutes.
(3) 0.1 mL of 1M sodium azide was added, the absorbance at 475 nm was measured, and the tyrosinase inhibition rate (%) was calculated. A 3 mM arbutin solution was used as a “positive control”.

[Test results]
Table 9 below shows the test results of the “distilled fraction (d)” obtained by the above low-temperature vacuum drying operation.
Moreover, the test result about the extract (E) by "ethanol", "50 volume% ethanol water", and "water" of "residue (R)" which remains in a carbonization apparatus (dry distillation kettle) after the above-mentioned low-temperature vacuum distillation operation Is shown in Table 10 below.

[Reproducibility about roses]
In order to see whether the results in Table 10 above are reproducible, the “residue (R) extract (E)” and “ The tyrosinase inhibition rate for the “frozen rose extract” was measured.
In addition, about the frozen rose, for the comparison with the solvent extract (E) of a residue (R), using the extract obtained by extracting the rose flower preserve | saved frozen with an extraction solvent, respectively. A test was conducted.
The results are shown in Table 11.

  From Table 11 above, a high tyrosinase inhibition rate was confirmed in the “extract with 50 volume% ethanol water (E)” from the residue (R) (dry powder that is a can residue) when roses were distilled at low temperature under reduced pressure. Is done. (This tyrosinase inhibition rate, as noted in the above [Test Solution] section, is a “sample solution” that is “10-fold diluted” with the solvent used for extraction, so the tyrosinase inhibition rate is very high. It can be said.)

Since the tyrosinase inhibitor of the present invention comprising a low-temperature vacuum distillation residue (R) of a plant raw material or an extract (E) using a solvent for its extraction as an active ingredient exhibits a preferable tyrosinase inhibitory action, it is particularly intended for whitening. It is useful as an additive for various uses including cosmetics, quasi drugs and health foods.

Claims (3)

A component distilled out when a plant raw material in a water-containing state is subjected to dry distillation under a temperature condition of 20 to 60 ° C. and a pressure condition of −100 to −96 kPa using a dry distillation apparatus equipped with a pressure reduction mechanism is referred to as “low temperature reduced pressure reduction”. When the solid or solid residue remaining in the carbonization apparatus after the low-temperature vacuum distillation operation is referred to as “low-temperature vacuum distillation residue (R)”
The plant material is a rose, and
The above-mentioned “low-temperature vacuum distillation residue (R) itself” or its “extract (E) with ethanol water having an ethanol concentration of 25 to 75% by volume from the low-temperature vacuum distillation residue (R)” as an active ingredient,
A tyrosinase inhibitor characterized by the above. The tyrosinase inhibitor according to claim 1, wherein the rose has a moisture content of 90% by weight or less. Carbonizing a plant raw material in a water-containing state using a carbonization apparatus equipped with a decompression mechanism under a temperature condition of 20 to 60 ° C. and a pressure condition of −100 to −96 kPa ,
The plant material is a rose,
Obtain “low temperature vacuum distillation residue (R)” which is a solid or solid residue remaining in the carbonization apparatus after the low temperature vacuum distillation operation, or further ethanol from the “low temperature vacuum distillation residue (R)” Obtaining an extract (E) by performing extraction with ethanol water having a concentration of 25 to 75% by volume ;
A process for producing a tyrosinase inhibitor characterized by