Persulfide-containing composition

Abstract

[Problem] The present invention addresses the problem of providing: a persulfide-containing composition containing a compound that is derived from a natural product or is widely used as an additive and that can stabilize a glutathione analogue persulfide or a cysteine analogue persulfide; and a method for stabilizing a persulfide. [Solution] By having a polyphenol coexist with a glutathione analogue persulfide or a cysteine analogue persulfide, it is possible to suppress degradation of the glutathione analogue persulfide or the cysteine analogue persulfide and stabilize the persulfide.

Description

Persulfide-containing composition

[0001] The present invention relates to persulfide-containing compositions and methods for stabilizing persulfides.

[0002] In recent years, persulfides (also referred to as active sulfur compounds, supersulfur compounds, or supersulfur molecules) have attracted attention as glutathione-related substances and N-acetylcysteine-related substances, and their high reactivity and physiological activity are expected. However, persulfides have a problem of low stability due to their high reactivity. For this reason, various studies have been conducted, and for example, a method for stabilizing persulfides using a novel tyrosine derivative is known (Patent Document 1). In addition, a method for stabilizing persulfides using a hydroxy compound has been reported (Patent Document 2).

[0003] However, when using persulfides in cosmetics or topical agents, there is often a reluctance to incorporate compounds that do not exist in nature or that are not used in existing products for stabilization, and there is a tendency to prefer products that use ingredients derived from natural products whenever possible. Furthermore, as shown in Figure 2A of Patent Document 2, it has been suggested that nucleophiles such as hydroxy anions contribute to the decomposition of persulfides, indicating that not all hydroxy compounds contribute to stabilization. Furthermore, it is well known that tyrosine is converted into melanin, the cause of age spots, via dopa or dopaquinone in the body.

[0004] Therefore, there has been a demand for a method for stabilizing persulfides using existing compounds derived from natural products and that are not tyrosine derivatives.

[0005] WO2021 / 172523 Pamphlet Japanese Patent Application Laid-Open No. 2021-66722

[0006] Br J Pharmacol., 176(4):671-683 (2019)

[0007] An object of the present invention is to provide a persulfide-containing composition containing a compound derived from a natural product or widely used as an additive, which is capable of stabilizing persulfides, and to provide a method for stabilizing persulfides.

[0008] As a result of extensive research into the above-mentioned problems, the present inventors have found that polyphenols contribute to stabilization by inhibiting the decomposition of persulfides, and have thus completed the present invention.

[0009] That is, the present invention relates to the following (1) to (8). (1) A persulfide-containing composition containing a persulfide of a glutathione analog or a persulfide of a cysteine ​​analog and a polyphenol. (2) The persulfide-containing composition according to (1), in which the polyphenol is present in an amount of 0.01 to 15 parts by weight per part by weight of the persulfide of a glutathione analog or the persulfide of a cysteine ​​analog. (3) A method for producing the persulfide-containing composition according to either (1) or (2), comprising a step of allowing the persulfide of a glutathione analog or the persulfide of a cysteine ​​analog to coexist with a polyphenol. (4) The persulfide-containing composition according to either (1) or (2), in which the polyphenol is a polyphenol having a galloyl group or a resorcinol group. (5) The persulfide-containing composition according to (3), in which the polyphenol is a polyphenol having a galloyl group or a resorcinol group. (6) A composition for external use comprising the composition according to (1) or (2). (7) A composition for oral use comprising the composition according to (1) or (2). (8) A method for stabilizing a persulfide of a glutathione analog or a persulfide of a cysteine ​​analog, characterized by allowing the persulfide of a glutathione analog or the persulfide of a cysteine ​​analog to coexist with a polyphenol.

[0010] According to the present invention, a composition containing a glutathione analog persulfide or a cysteine ​​analog persulfide and a polyphenol inhibits decomposition of the glutathione analog persulfide or the cysteine ​​analog persulfide.Furthermore, a method for stabilizing the glutathione analog persulfide or the cysteine ​​analog persulfide by using a polyphenol can be provided.

[0011] Screening results Storage test results at 60°C Storage test results at 40°C

[0012] The compositions of the present invention contain glutathione analog persulfide or cysteine ​​analog persulfide (hereinafter, sometimes collectively referred to as "persulfides"). Here, glutathione analogs in the present invention refer to reduced or oxidized glutathione, reduced or oxidized γ-glutamylcysteine, reduced or oxidized cysteinylglycine, and compounds in which one or more amino or carboxyl groups in these structures are modified with a linear or branched alkyl group having 1 to 24 carbon atoms, as well as compounds esterified with an organic or inorganic acid. Furthermore, cysteine ​​analogs in the present invention refer to cysteine, cystine, reduced or oxidized N-acetylcysteine ​​(hereinafter, sometimes referred to as "NAC"), compounds in which one or more amino or carboxyl groups in these structures are modified with a linear or branched alkyl group having 1 to 24 carbon atoms, and compounds esterified with an organic or inorganic acid. The term "persulfide" as used herein refers to a molecule having a structure in which one or more sulfur atoms are directly bonded to a sulfur atom constituting a thiol group or disulfide bond present in a typical glutathione analog or cysteine ​​analog molecule (e.g., RS-SH or RSSS-R'; R and R' are any functional groups). Any glutathione analog or cysteine ​​analog having one or more sulfur atoms directly bonded to a sulfur atom in the molecule can be used in the present invention. A mixture of multiple compounds with different numbers of sulfur atoms in the molecule may also be used. The persulfide contained in the composition of the present invention is preferably an oxidized form, more preferably oxidized glutathione persulfide or oxidized NAC persulfide. Oxidized glutathione persulfide (oxidized glutathione trisulfide) or oxidized NAC persulfide (oxidized NAC trisulfide), which contain three sulfur atoms per molecule, is even more preferred.

[0013] Persulfides can be obtained using commonly available glutathione analogs or cysteine ​​analogs. For example, oxidized glutathione and peracetic acid are reacted on ice for 30 minutes, followed by the addition of a solvent consisting of equal parts ethanol and tetrahydrofuran (THF). The precipitate obtained by centrifugation is then further reacted with sodium hydrogen sulfide at room temperature to obtain oxidized glutathione persulfide. Purification of this oxidized glutathione persulfide produces oxidized glutathione trisulfide, which can be obtained in crystalline, amorphous, solution, or suspension forms. Oxidized NAC persulfide can be obtained using a similar process using oxidized N-acetylcysteine ​​as a raw material.

[0014] The composition of the present invention contains the persulfide and a polyphenol. The polyphenol in the present invention refers to a compound having two or more phenolic hydroxyl groups on an aromatic ring contained in one molecule. Among polyphenols, those having a galloyl group or a resorcinol group and a molecular weight of 120 to 2000 are particularly preferred. Examples of such polyphenols include pyrogallol (CAS No.: 87-66-1), hydroxyquinol (CAS No.: 533-73-3), galacetophenone (CAS No.: 528-21-2), gallic acid (CAS No.: 149-91-7), methyl gallate (CAS No.: 99-24-1), ethyl gallate (CAS No.: 831-61-8), propyl gallate (CAS No.: 121-79-9), octyl gallate (CAS No.: 1034-01-1), and dodecyl gallate. Examples of suitable polyphenols include decyl catechin (CAS No. 1166-52-5), tannic acid (CAS No. 1401-55-4), quercetin (CAS No. 117-39-5), epigallocatechin gallate (CAS No. 989-51-5), hesperidin (CAS No. 520-26-3), ellagic acid (CAS No. 476-66-4), and resveratrol (CAS No. 501-36-0). Particularly preferred are pyrogallol, propyl gallate, tannic acid, and quercetin. Glycosides in which sugars are bound to the polyphenols may also be used. Each polyphenol can be quantified by standard analytical techniques. For example, the amount of polyphenols contained in an analytical sample can be determined from a calibration curve prepared by HPLC using a standard sample of known concentration.

[0015] The composition of the present invention may be used in combination with other materials, as long as the effects of the present invention are not impaired. In addition to the persulfides and polyphenols, these may include thickeners, pH adjusters, fragrances, bases for topical preparations, excipients, etc.

[0016] The method for making the persulfide and polyphenol coexist is not particularly limited, and they may be mixed together with other raw materials when preparing the composition, or they may be mixed at a different time from the other raw materials. The form of the composition is not limited as long as it can be taken orally or applied to the skin, and can be prepared in the form of a solution, dispersion, powder, granules, capsules, tablets, paste, gel, or sheet according to a conventional method using solvents or bases used in general foods, supplements, cosmetic compositions, pharmaceutical compositions, and topical skin preparations.

[0017] The content of each component in the composition of the present invention can be appropriately adjusted by adding excipients and the like that are generally used in the above forms.

[0018] The content of persulfides in the composition of the present invention can be determined by a common method using HPLC. The content of persulfides in the composition of the present invention can be adjusted as appropriate, and typically the composition contains 0.001 to 0.7 wt %, preferably 0.0012 to 0.5 wt % of persulfides. The amount of polyphenols contained in the composition of the present invention is preferably 0.01 to 15 wt %, more preferably 0.02 to 10 wt %, and even more preferably 0.04 to 8 wt %, per 1 wt % of persulfides.

[0019] The method for stabilizing persulfides in the present invention can be achieved by allowing persulfides and polyphenols to coexist. The mechanism of stabilization is thought to be inhibition of persulfide decomposition. The degree of stabilization of persulfides can be determined, for example, from the residual rate of persulfides when the initial amount of persulfides is taken as 100. The stabilization of persulfides in the present invention can be determined as a relative value based on the amount of persulfides obtained when a 0.01 wt% aqueous solution (pH 5.5) is stored under conditions equivalent to one year of storage at room temperature, for example, at 85°C for 16 hours. The effect of adding a stabilizer is such that the residual rate of persulfides in the test group to which the stabilizer is added is preferably 1.2 times or more, more preferably 1.5 times or more, and even more preferably 1.8 times, compared to the residual rate of persulfides when stored under the above conditions (pH 5.5, 85°C, 16 hours) without the stabilizer.

[0020] The persulfide-containing composition can be applied to the skin as an external preparation or taken orally. It can also be prepared into a desired dosage form as described above. The desired dosage form can be prepared according to a general method.

[0021] The present invention will be specifically explained below using examples, but the present invention is not limited to these examples.

[0022] The persulfide was produced using the following method based on a previously published paper (Non-Patent Document 1). Commercially available oxidized glutathione as a reagent was dissolved in ultrapure water to prepare a 170 mM oxidized glutathione aqueous solution. 4 mL of 32 wt% peracetic acid solution was added to 2.5 mL of this oxidized glutathione aqueous solution and allowed to react on ice for 30 minutes. 30 mL of a solvent consisting of an equal mixture of ethanol and tetrahydrofuran was added to the resulting reaction solution, which was then centrifuged at 10,000 rpm at 4°C for 10 minutes. The recovered precipitate was dried under reduced pressure for 1 hour. 2.5 mL of ultrapure water was added to the resulting dried product and dissolved it. An equal volume of sodium hydrogen sulfide solution adjusted to 60 mM with 0.3 M sodium hydroxide aqueous solution was then added, and the mixture was allowed to react at room temperature for 1 hour to obtain oxidized glutathione persulfide. The resulting oxidized glutathione persulfide was purified using a synthetic adsorption resin, concentrated, and dried to obtain oxidized glutathione trisulfide powder.

[0023] (Test to confirm the stabilizing effect of oxidized glutathione trisulfide) Oxidized glutathione trisulfide (GSSSG) powder was dissolved in water to prepare a 0.01 wt% aqueous solution. The pH at this time was 3.9. Each test substance was added to this solution to the final concentration listed in Table 1, setting up test groups 2 to 19. The pH was adjusted to 5.5 with NaOH and then allowed to stand at 85°C for 16 hours. A test group to which no test substance was added served as a control (test group 1). The amount of residual GSSSG in each sample was then quantified by HPLC, and the relative value was calculated as a percentage, with the value before storage at 85°C set to 100.

[0024]

[0025] (Results of the Stabilizing Effect Confirmation Test) The results are shown in Figure 1. (The vertical axis represents the GSSSG residual rate at 0 hr = 100%). The residual rate of GSSSG in the control (Test Group 1) was 27%. In contrast, the test groups containing polyphenolic compounds such as propyl gallate and tannic acid (Test Groups 5-19) exhibited residual rates 1.5 times (40.5%) higher than the control. Among these, propyl gallate (Test Groups 5 and 6), tannic acid (Test Groups 7 and 8), and quercetin (Test Groups 16 and 17) exhibited residual rates more than twice that of the control. On the other hand, tyrosine (Test Group 2) and inositol (Test Groups 3 and 4), both of which contain hydroxyl groups, did not exhibit any stabilizing effect on GSSSG. These results demonstrate that polyphenols have the effect of stabilizing GSSSG even under heated conditions, and that the compositions of the present invention are stable under heated conditions.

[0026] (Stabilization test of oxidized glutathione trisulfide) Oxidized glutathione trisulfide (GSSSG) powder was dissolved in citric acid-sodium citrate buffer to prepare a 0.1 wt% GSSSG solution at pH 5.0. To this solution, propyl gallate (final concentration: 0.05 wt%, 0.1 wt%, or 0.2 wt%), tannic acid (final concentration: 0.05 wt%, 0.1 wt%, or 0.2 wt%), or quercetin (final concentration: 0.005 wt%, 0.01 wt%, or 0.02 wt%) was added. After storage at 60°C for 1 week or 40°C for 10 weeks, which corresponds to one year of storage at room temperature for persulfide stabilization, the amount of remaining oxidized glutathione trisulfide was quantified by HPLC and calculated relative to the amount of persulfide before storage (100).

[0027] The results are shown in Figure 2 (1 week at 60°C) and Figure 3 (10 weeks at 40°C). Under both conditions, the control showed a residual rate of approximately 30%, whereas the addition of stabilizers such as polyphenols, such as propyl gallate, tannic acid, and quercetin, inhibited the decomposition of persulfides and improved the residual rate. Furthermore, although not shown, the pyrogallol-added sample showed a GSSSG residual rate of 59.9% after storage at 60°C for 1 week, demonstrating its contribution to the stabilization of persulfides. These results demonstrate that polyphenols are effective in stabilizing GSSSG even under prolonged heating conditions, and that the compositions of the present invention are stable even under prolonged heating conditions. The vertical axis of Figures 2 and 3 represents the GSSSG residual rate (0 hr = 100%), and the horizontal axis represents Control, 1: Propyl gallate, 2: Tannic acid, and 3: Quercetin.

[0028] From the above, it has been revealed that the coexistence of glutathione persulfide and polyphenols can suppress the decomposition of persulfide and stabilize it even under long-term heating conditions.Since the polyphenols used in the present invention can be used as cosmetics and foods, the composition of the present invention can be used in general foods, beverages, cosmetics, pharmaceuticals, etc.

Claims

1. A persulfide-containing composition containing a persulfide of a glutathione analog or a persulfide of a cysteine ​​analog and a polyphenol.

2. The persulfide-containing composition according to claim 1, wherein the polyphenol is present in an amount of 0.01 to 15 parts by weight per part by weight of the persulfide of a glutathione analog or the persulfide of a cysteine ​​analog.

3. A method for producing the persulfide-containing composition according to claim 1 or 2, comprising a step of allowing a persulfide of a glutathione analog or a persulfide of a cysteine ​​analog to coexist with a polyphenol.

4. The persulfide-containing composition according to claim 1 or 2, wherein the polyphenol is a polyphenol having a galloyl group or a resorcinol group.

5. The persulfide-containing composition according to claim 3, wherein the polyphenol is a polyphenol having a galloyl group or a resorcinol group.

6. A topical composition comprising the composition of claim 1 or 2.

7. An oral composition comprising the composition of claim 1 or 2.

8. A method for stabilizing a persulfide of a glutathione analog or a persulfide of a cysteine ​​analog, characterized by coexisting the persulfide of a glutathione analog or the persulfide of a cysteine ​​analog with a polyphenol.

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