Food and beverage composition, Anti-aging agent, mitochondrial dysfunction improving agent, nicotinamide phosphoribosyltransferase production promoter, nicotinamide adenine dinucleotide production promoter, sirtuin 1 production promoter, and melanin production inhibitor

Abstract

To provide a novel technology for suppressing aging.SOLUTION: The present invention provides a food and beverage composition comprising longan aril and pyrroloquinoline quinone.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to a food and drink composition, an anti-aging agent, an agent for improving mitochondrial dysfunction, a nicotinamide phosphoribosyltransferase production promoter, a nicotinamide adenine dinucleotide production promoter, a sirtuin 1 production promoter, and a melanin production inhibitor. [Background technology]

[0002] With the recent progress of aging society, the importance of technologies to suppress aging is increasing.

[0003] For example, Patent Document 1 focuses on nicotinamide phosphoribosyltransferase (NAMPT), an enzyme involved in the inhibition of aging, and proposes an NAMPT activator containing, as an active ingredient, one or more selected from the group consisting of grape seed, milk thistle, ginseng, houttuynia cordata, guava, pomegranate, HMB-Ca, cowberry, cinnamon bark, gardenia fruit, mulberry leaf, longan fruit, jujube, NMN, and chou-mei-sou. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Japanese Patent Publication No. 2024-022503 Summary of the Invention [Problem to be solved by the invention]

[0005] However, there is a need for further techniques to inhibit aging.

[0006] The present invention has been made in view of the above circumstances, and an object of the present invention is to provide a novel aging prevention technology. [Means for solving the problem]

[0007] As a result of investigations, the present inventors have found that the above-mentioned problems can be solved by combining Longan Fruit with Pyrroloquinoline Quinone, and have thus completed the present invention. More specifically, the present invention provides the following.

[0008] (1) A food or beverage composition comprising longan fruit and pyrroloquinoline quinone.

[0009] (2) The food and beverage composition according to (1), wherein the mass ratio of the component derived from longan fruit to the pyrroloquinoline quinone is 5:1 to 30:1.

[0010] (3) An anti-aging agent comprising longan fruit and pyrroloquinoline quinone.

[0011] (4) A mitochondrial dysfunction improving agent comprising longan berry and pyrroloquinoline quinone.

[0012] (5) A nicotinamide phosphoribosyltransferase production promoter comprising longan berry and pyrroloquinoline quinone.

[0013] (6) A nicotinamide adenine dinucleotide production promoter comprising longan berry and pyrroloquinoline quinone.

[0014] (7) A sirtuin 1 production promoter comprising longan berry and pyrroloquinoline quinone.

[0015] (8) A melanin production inhibitor comprising longan fruit and pyrroloquinoline quinone. [Effects of the Invention]

[0016] According to the present invention, a novel anti-aging technology is provided. [Brief explanation of the drawings]

[0017] [Figure 1] FIG. 1 is a diagram showing the results of the effect on melanin production in an example. [Figure 2] FIG. 1 shows the results of the influence on the amount of NAMPT produced in an example. [Figure 3] FIG. 1 shows the results of the influence on the amount of NAD+ production in an example. [Figure 4] FIG. 1 shows the results of the influence on the amount of IL-1β produced in an example. [Figure 5] FIG. 1 shows the results of the effect on the amount of GDF-15 produced in an example. [Figure 6] FIG. 1 shows the results of the effect on the amount of NLRP3 produced in an example. [Figure 7] FIG. 1 shows the results of the influence on Sirt1 production amount in an example. [Figure 8] FIG. 1 shows the results of the influence on Per2 production amount in an example. [Figure 9] FIG. 1 shows the results of the influence on the amount of NAD+ production in an example. [Figure 10] FIG. 1 shows the results of the influence on the amount of ATP produced in an example. [Figure 11] FIG. 1 is a graph showing the results of the influence on lactate levels in an example. [Figure 12] FIG. 1 shows the results of the influence on oxygen consumption rate in an example. [Figure 13] FIG. 1 shows the results of the effects on cell viability in an example. [Figure 14] FIG. 1 shows the results of the effect on SPiDER-βGal in an example. DETAILED DESCRIPTION OF THE INVENTION

[0018] Hereinafter, an embodiment of the present invention will be described, but the present invention is not limited to this.

[0019] The following abbreviations may be used hereinafter: Pyrroloquinoline quinone: PQQ Nicotinamide phosphoribosyltransferase: NAMPT Nicotinamide adenine dinucleotide: NAD + Nicotinamide: NAM β-nicotinamide mononucleotide: NMN Interleukin-1β: IL-1β Growth differentiation factor 15: GDF-15 Pyrin domain-containing protein 3: NLRP3 Adenosine triphosphate: ATP Oxygen Consumption Rate: OCR 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide: MTT Sirtuin 1: Sirt1 Clock gene Period2: Per2 Fluorescent probe for detecting intracellular β-galactosidase: SPiDER-βGal

[0020] (1) Food and drink composition The food and beverage composition of the present invention contains longan berry and pyrroloquinoline quinone.

[0021] As mentioned above, it has been reported that longan berry and the like have NAMPT activating activity on their own. On the other hand, the present inventors have made the unexpected discovery that combining longan fruit with pyrroloquinoline quinone can not only promote NAMPT production but also exert various effects related to the suppression of aging.

[0022] In the present invention, the "anti-aging effect" includes the promotion of the production of substances known to be involved in anti-aging, the inhibition of the production of substances known to be involved in accelerating aging, and various events resulting from these.

[0023] For example, in one embodiment of the present invention, the "aging inhibitory effect" includes the following effects and the anti-aging effects that can be achieved by the anti-aging agent of the present invention described below. -Inhibition of melanin production and the resulting skin whitening effect. - Promotes NAMPT production, and the resulting effects of skin whitening, antioxidant effects, life extension, and damage repair. NAD + It has the effect of promoting production, and the resulting effects of improving mitochondrial dysfunction, whitening the skin, antioxidant effects, extending lifespan, and repairing damage.

[0024] The constitution of the food and drink composition of the present invention will be described in detail below.

[0025] (1-1) Longan Niku Longan fruit (scientific name: Euphoria longana Lamarck (Sapindaceae) or Dimocarpus longan L.), also known as "longan," is a fruit tree of the genus Longan in the family Sapindaceae.

[0026] Longan fruit has traditionally been used in food and beverages for its arils (the skin that covers the seed coat, which is formed by the thickening of the pedicel and placenta) and extracts thereof. In the present invention, any of these longan fruits that have been conventionally used can be used.

[0027] The form of longan cherries is not particularly limited, and they may be in the form of powder, liquid, solid, paste, etc.

[0028] In a preferred embodiment of the present invention, the longan berry is a water extract of arils. The water extract may be subjected to appropriate procedures such as filtration, concentration, sterilization, powderization, etc. Furthermore, an excipient (dextrin, etc.) may be blended as necessary.

[0029] In a preferred embodiment of the present invention, the longan berry is a powder obtained by the following process: The arils of longan fruit are used as the starting material, which are extracted with water, filtered, concentrated (with pectinase treatment), and sterilized (at 125±10°C). Next, excipients are optionally added, followed by spray drying and granulation, and the resulting powder is collected as longan extract powder.

[0030] As longan rind, commercially available products (such as the longan rind extract powder used in the examples) may be used.

[0031] The content of longan cherries in the food and beverage composition is not particularly limited and can be adjusted appropriately depending on the effect to be obtained. The lower limit of the content of longan cherries (dry mass of components derived from longan cherries) is preferably 0.2% by mass or more, and more preferably 1.0% by mass or more, relative to the food and beverage composition. The upper limit of the content of longan berry (dry mass of components derived from longan berry) is preferably 10.0% by mass or less, and more preferably 5.0% by mass or less, of the food or drink composition.

[0032] In the present invention, the "dry mass of components derived from longan pulp" means the total amount of components excluding water from longan pulp blended in the food and beverage composition of the present invention.

[0033] (1-2) Pyrroloquinoline quinone PQQ(C 14 H6N2O8 (CAS registration number: 72909-34-3) functions as an oxidation-reduction coenzyme in the body and is known to have antioxidant and neuroprotective effects. However, the fact that a combination of PQQ and longan fruit has a good anti-aging effect was discovered for the first time by the present inventors.

[0034] In the present invention, PQQ includes not only the free form of PQQ but also salts of PQQ. These free forms and salts may be used alone or in combination of two or more.

[0035] Salts of PQQ include any salts that can be used as foods and drinks. Preferred salts include pyrroloquinoline quinone disodium salt.

[0036] The content of PQQ in the food and drink composition is not particularly limited and can be adjusted appropriately depending on the effect to be obtained. The lower limit of the PQQ content (equivalent to free PQQ) is preferably 0.00044% by mass or more, and more preferably 0.02200% by mass or more, relative to the food or drink composition. The upper limit of the PQQ content (equivalent to free PQQ) is preferably 0.44000% by mass or less, and more preferably 0.22000% by mass or less, relative to the food or drink composition.

[0037] In the present invention, the term "amount converted into free PQQ" means the amount of PQQ incorporated into the food and drink composition of the present invention, converted into free PQQ. The amount of free PQQ in a food or drink composition is calculated based on the content of pyrroloquinoline quinone added and the molecular weight of pyrroloquinoline quinone or its salt.

[0038] (1-3) Total amount and ratio of longan and pyrroloquinoline quinone The total amount and ratio of longan berry and PQQ to be blended in the food and beverage composition are not particularly limited and can be set appropriately depending on the desired effect, etc.

[0039] From the viewpoint of more easily achieving the effects of the present invention, it is preferable to increase the ratio of longan nut to PQQ. In the food and beverage composition, the mass ratio of components derived from longan berry (dry mass of components derived from longan berry) to PQQ (equivalent to free PQQ) is preferably 5:1 to 30:1, more preferably 10:1 to 20:1, and even more preferably 12.5:1 to 15:1.

[0040] According to the present invention, an effective anti-aging effect can be achieved even when the total amount of longan berry and PQQ is lower than when longan berry or PQQ is blended alone. From this viewpoint, the lower limit of the total amount of components derived from longan rind (dry mass of components derived from longan rind) and PQQ (equivalent to free PQQ) is preferably 0.5% by mass or more, more preferably 1.0% by mass or more, relative to the food and beverage composition. In addition, the upper limit of the total amount of components derived from longan berry (dry mass of components derived from longan berry) and PQQ (equivalent to free PQQ) is preferably 10.0 mass% or less, more preferably 2.0 mass% or less, relative to the food and beverage composition.

[0041] (1-4) Other ingredients The food and drink composition may or may not contain ingredients other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0042] Ingredients other than longan berry and PQQ can be appropriately selected depending on the form of the food or beverage composition, and include any ingredients used as ingredients in food or beverages. Such ingredients include dyes, antioxidants, excipients, colorants, surfactants, preservatives, pH adjusters, and the like.

[0043] One embodiment of the present invention includes an embodiment in which the food or beverage composition consists solely of longan berry and PQQ.

[0044] (1-5) Form of food and beverage composition The form of the food or drink composition is not particularly limited and includes any form.

[0045] The form of the food or drink composition may be powder, liquid, solid, granule, particulate, paste, gel, or the like.

[0046] Specific examples of food and drink compositions include nutritional supplements, cooked foods, seasonings, soft drinks, jelly drinks, and confectioneries (for example, gummies, candies, Japanese sweets, and Western sweets).

[0047] The amount and frequency of intake of the food and drink composition are not particularly limited and can be set appropriately depending on the effects to be obtained, etc.

[0048] (1-6) Method for producing food and beverage compositions The method for producing the food and drink composition is not particularly limited, and any method can be adopted depending on the type of food and drink composition.

[0049] (2) Anti-aging agents The anti-aging agent of the present invention contains longan berry and pyrroloquinoline quinone.

[0050] In the present invention, the term "anti-aging (effect)" includes changes (increases or decreases) in aging markers, as well as inhibition of changes in aging markers. NAD is an aging marker whose increase indicates anti-aging. + , ATP, OCR, cell viability (MTT), Sirt1, etc. Aging markers whose decrease indicates anti-aging include lactic acid, SPiDER-βGal, GDF-15, IL-1β, and NLRP3. Among the aging markers, changes in Sirt1 can affect Per2 expression (Nakahata Y, Kaluzova M (2008) Cell 134:329-340, etc.). For example, an increase in Sirt1 can result in a decrease in Per2, which can lead to the formation of a circadian rhythm. Therefore, Per2 can also be used as an indirect indicator of anti-aging.

[0051] The presence or absence and degree of anti-aging effect can be evaluated by culturing using a melanocyte-containing three-dimensional skin model shown in the Examples or by culturing using human skin fibroblasts.

[0052] (2-1) Ingredients contained in anti-aging agents The components contained in the anti-aging agent can have the same constitution as that of the food and drink composition described above.

[0053] The anti-aging agent may or may not contain ingredients other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0054] One embodiment of the present invention includes an embodiment in which the anti-aging agent consists solely of longan fruit and PQQ.

[0055] (2-2) Form of anti-aging agent The form of the anti-aging agent is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0056] When the anti-aging agent is a pharmaceutical, its form is preferably an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0057] When the anti-aging agent is a food or drink composition, the same constitution as that of the food or drink composition described above can be adopted.

[0058] (2-3) Method for producing anti-aging agent The method for producing the anti-aging agent is not particularly limited, and any method can be adopted depending on the form of the agent.

[0059] (3) Mitochondrial dysfunction improver The agent for improving mitochondrial dysfunction of the present invention contains longan fruit and pyrroloquinoline quinone.

[0060] Mitochondria are intracellular organelles that play a vital role in living organisms, including ATP production. In recent years, mitochondrial dysfunction has been reported to be involved in cellular aging. According to the present invention, the effect of improving mitochondrial dysfunction in the body is exerted, and therefore, an anti-aging effect can be expected through the improvement of mitochondrial function.

[0061] In the present invention, "mitochondrial dysfunction amelioration (effect)" includes improvement of mitochondrial function in the living body compared to when the mitochondrial dysfunction ameliorating agent of the present invention is not ingested, and the like. Markers whose increase indicates improvement of mitochondrial dysfunction include ATP and OCR. Markers whose decrease indicates improvement of mitochondrial dysfunction include lactate and GDF-15.

[0062] The presence or absence and degree of the effect of improving mitochondrial dysfunction can be evaluated by culturing using the melanocyte-containing three-dimensional skin model shown in the Examples or by culturing using human skin fibroblasts.

[0063] (3-1) Ingredients contained in mitochondrial dysfunction improving agents The components contained in the mitochondrial dysfunction ameliorating agent can have the same configuration as the above-mentioned food and drink composition.

[0064] The agent for improving mitochondrial dysfunction may or may not contain ingredients other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0065] One embodiment of the present invention includes an embodiment in which the mitochondrial dysfunction ameliorating agent consists solely of longan fruit and PQQ.

[0066] (3-2) Form of mitochondrial dysfunction improving agent The form of the mitochondrial dysfunction ameliorating agent is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0067] When the agent for ameliorating mitochondrial dysfunction is a pharmaceutical product, its form is preferably an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0068] When the mitochondrial dysfunction ameliorating agent is a food or drink composition, the same constitution as the food or drink composition described above can be adopted.

[0069] (3-3) Method for producing mitochondrial dysfunction improving agent The method for producing the agent for ameliorating mitochondrial dysfunction is not particularly limited, and any method can be adopted depending on the form of the agent.

[0070] (4) Nicotinamide phosphoribosyltransferase production promoter The NAMPT production promoter of the present invention contains longan berry and pyrroloquinoline quinone.

[0071] NAMPT is an enzyme that converts NAM to NMN and regulates the production of NAD (see below). Activation of NAMPT has been suggested to be useful in treating and improving skin diseases and conditions through anti-aging, lifespan extension, and damage repair. According to the present invention, the effect of promoting NAMPT production in the body is exerted, and therefore, an anti-aging effect can be expected through the improvement of various biological functions by NAMPT. For example, NAD + The promotion of production can have the effect of suppressing melanin production.

[0072] In the present invention, the "NAMPT production promoting effect" includes the promotion of the amount of NAMPT produced in the body compared to when the NAMPT production promoter of the present invention is not ingested, through the promotion of NAMPT gene expression or the promotion of the NAMPT production mechanism, etc.

[0073] The presence or absence and degree of the NAMPT production promoting effect can be evaluated by culturing using the melanocyte-containing three-dimensional skin model shown in the Examples or by culturing using human skin fibroblasts.

[0074] (4-1) Ingredients contained in NAMPT production promoters The components contained in the NAMPT production promoter can have the same constitution as the above-mentioned food and drink composition.

[0075] The NAMPT production promoter may or may not contain components other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0076] One embodiment of the present invention includes an embodiment in which the NAMPT production promoter consists solely of longan berry and PQQ.

[0077] (4-2) Form of NAMPT production promoter The form of the NAMPT production promoter is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0078] When the NAMPT production promoter is a pharmaceutical, it is preferably in the form of an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0079] When the NAMPT production promoter is a food or drink composition, it can have the same structure as the food or drink composition described above.

[0080] (4-3) Method for producing NAMPT production promoter The method for producing the NAMPT production promoter is not particularly limited, and any method can be adopted depending on the form of the agent.

[0081] (5) Nicotinamide adenine dinucleotide production promoter The NAD of the present invention + The production promoter includes longan berry and pyrroloquinoline quinone.

[0082] NAD + NAD is an important coenzyme that exists in various parts of the body and plays various roles in energy metabolism, cell metabolism, gene expression, DNA repair, etc. + It has been reported that the molecular weight of α-glucan is large, making it difficult to absorb when administered externally, and that it is decomposed in the intestines. According to the present invention, NAD in vivo + NAD + It is expected that this will have an anti-aging effect through the improvement of various biological functions. For example, NAD + The promotion of production can have the effect of suppressing melanin production.

[0083] In the present invention, "NAD + The "production promoting effect" is NAD + Increased gene expression and NAD + Through the enhancement of the production mechanism, NAD in the body + The amount of NAD produced by the present invention + This includes promotion compared to when the production promoter is not taken.

[0084] NAD + The presence or absence and degree of the production-promoting effect can be evaluated by culturing using the melanocyte-containing three-dimensional skin model shown in the Examples or by culturing using human skin fibroblasts.

[0085] (5-1) NAD + Ingredients contained in the production promoter NAD + The components contained in the production promoter can have the same configuration as the above-mentioned food and drink composition.

[0086] NAD + The production promoter may or may not contain components other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0087] One aspect of the present invention is a NAD + This includes an embodiment in which the production enhancer consists solely of longan berry and PQQ.

[0088] (5-2) NAD + Form of production promoter NAD + The form of the production enhancer is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0089] NAD + When the production-enhancing agent is a pharmaceutical, it is preferably in the form of an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0090] NAD + When the production promoter is a food or drink composition, the same composition as the food or drink composition described above can be adopted.

[0091] (5-3) NAD + Method for producing a production promoter NAD + There are no particular limitations on the method for producing the production promoter, and any method can be used depending on the form of the agent.

[0092] (6) Sirtuin 1 production promoter The sirtuin 1 production promoter of the present invention contains longan berry and pyrroloquinoline quinone.

[0093] Sirtuin 1 inhibits NAD + It is one of the enzymes responsible for the dependent deacetylases and is also known as the longevity gene.

[0094] In the present invention, the "effect of promoting sirtuin 1 production" includes an increase in sirtuin 1 gene expression and an increase in the amount of sirtuin 1 produced in the body compared to when the sirtuin 1 production promoter of the present invention is not ingested.

[0095] The presence or absence and degree of the effect of promoting sirtuin 1 production can be evaluated by culturing using the melanocyte-containing three-dimensional skin model shown in the Examples or by culturing using human skin fibroblasts.

[0096] (6-1) Ingredients contained in Sirtuin 1 production promoters The components contained in the sirtuin 1 production enhancer may have the same configuration as the above-mentioned food and drink composition.

[0097] The sirtuin 1 production promoter may or may not contain components other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0098] One embodiment of the present invention includes an embodiment in which the sirtuin 1 production promoter consists solely of longan berry and PQQ.

[0099] (6-2) Form of Sirtuin 1 Production Promoter The form of the sirtuin 1 production enhancer is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0100] When the sirtuin 1 production enhancer is a pharmaceutical, it is preferably in the form of an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0101] When the sirtuin 1 production enhancer is a food or drink composition, it may have the same composition as the food or drink composition described above.

[0102] (6-3) Method for producing sirtuin 1 production promoter The method for producing the sirtuin 1 production promoter is not particularly limited, and any method can be used depending on the form of the agent.

[0103] (7) Melanin production inhibitors The melanin production inhibitor of the present invention contains longan fruit and pyrroloquinoline quinone.

[0104] Melanin (C 18 H 10 N2O4 (CAS number: 8049-97-6) is a colored substance produced by melanocytes (pigment cells) present in the skin, etc., and can cause skin aging.

[0105] In the present invention, the "melanin production inhibitory effect" includes a reduction in the amount of melanin produced in the body compared to when the melanin production inhibitor of the present invention is not ingested, through a reduction in anti-inflammatory action, a reduction in melanin gene expression, an inhibition of the melanin production mechanism in melanocytes, etc.

[0106] In the present invention, the "melanin production inhibitory effect" includes changes (increases or decreases) in melanin production markers, inhibition of changes in melanin production markers, and the like. Examples of melanin production markers whose reduction indicates an inhibitory effect on melanin production include IL-1β and NLRP3.

[0107] The presence or absence and degree of the melanin production inhibitory effect can be evaluated by culturing using the melanocyte-containing three-dimensional skin model shown in the Examples.

[0108] (7-1) Ingredients contained in melanin production inhibitors The components contained in the melanin production inhibitor can have the same configuration as the above-mentioned food and drink composition.

[0109] The melanin production inhibitor may or may not contain ingredients other than longan berry and PQQ, as long as the effects of the present invention are not impaired.

[0110] One embodiment of the present invention includes an embodiment in which the melanin production inhibitor consists solely of longan fruit and PQQ.

[0111] (7-2) Form of melanin production inhibitor The form of the melanin production inhibitor is not particularly limited, and includes any form (pharmaceutical, food and drink composition, etc.).

[0112] When the melanin production inhibitor is a pharmaceutical, it is preferably in the form of an oral agent, although the present invention does not exclude parenteral agents (such as transdermal agents).

[0113] When the melanin production inhibitor is a food or drink composition, the same constitution as the food or drink composition described above can be adopted.

[0114] (7-3) Method for producing melanin production inhibitor The method for producing the melanin production inhibitor is not particularly limited, and any method can be used depending on the form of the agent. [Example]

[0115] The present invention will be explained in more detail below with reference to examples, but the present invention is not limited to these examples.

[0116] <Test using a melanin-containing 3D skin model-1> Using the following method, various evaluations were carried out under various addition conditions using a melanin-containing three-dimensional skin model.

[0117] (1) Sample preparation The following three types of samples were prepared and subjected to the culture test described below.

[0118] (1-1) Longan fruit sample Longan extract powder (manufactured by Matsuura Pharmaceutical Co., Ltd.) was prepared. This sample was prepared from the arils of longan fruit (scientific name: Euphoria longana (Sapindaceae)), which were extracted with water, filtered, concentrated (with pectinase treatment), and sterilized (125±10°C), followed by the addition of dextrin, spray drying, and sieving (sieve: 0.5 mm opening, with heavy metal removal by magnetic treatment). This resulted in a powder sample. The longan extract powder used in this example contains an excipient (dextrin) as well as components derived from longan. Therefore, the longan concentration hereinafter indicates the amount (dry mass) converted to components derived from longan.

[0119] (1-2) Pyrroloquinoline quinone sample Pyrroloquinoline quinone disodium salt (purity 99.0% or higher, manufactured by Ryusendo Co., Ltd.) was prepared. Pyrroloquinoline quinone disodium salt will hereinafter also be referred to as "PQQ." This sample is a powdered sample obtained by fermentation production using microbial fermentation and purification processes (hydroethanol extraction, concentration, drying, filtration, etc.). Hereinafter, the concentration of the pyrroloquinoline quinone sample (sodium salt) is expressed as the amount converted into the free pyrroloquinoline quinone.

[0120] (1-3) Positive control sample As a positive control substance, L-(-) ascorbic acid (powder sample, manufactured by Kanto Chemical Co., Ltd.) was prepared. Hereinafter, L-(-) ascorbic acid will also be referred to as "VC."

[0121] (2) Culture test In this example, the following culture methods (system using a three-dimensional skin model) were commonly adopted.

[0122] The culture was carried out in an incubator under specified ingredient addition conditions for 3 weeks at 37°C and 5% CO2 using a melanocyte-containing three-dimensional skin model ("MEL-300", manufactured by Kurabo Industries, Ltd.).

[0123] The culture medium used was EPI-100NMM113, which contains melanin-stimulating factors bFGF, α-MSH, and KGF. These components function as melanin-forming factors, pro-inflammatory factors, and external stimulatory factors in this culture system. The medium was changed every other day, and each time the test substance was dissolved in the medium, the biological activity of the test substance was evaluated from the bottom side of the skin model, which is intended for food use.

[0124] The conditions for adding ingredients to the medium are as follows. Control: Medium only VC(100): Positive control sample 100μg / ml Longan (30): Longan sample 30 μg / ml Longan (100): Longan sample 100 μg / ml PQQ(5): Pyrroloquinoline quinone sample 5μg / ml PQQ(10): Pyrroloquinoline quinone sample 10 μg / ml Longan (50) + PQQ (5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 5 μg / ml Longan (15) + PQQ (2.5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 2.5 μg / ml

[0125] Four wells were set up for each sample (n=4).

[0126] Statistical analysis of the obtained results was performed using Bonferroni multiple comparisons or Dunnett test as a multiple comparison test. In both methods, whether or not a significant improvement was shown compared to the control group was determined, and this was expressed as *p<0.05, **p<0.01. Note that a significant trend was determined to exist when 0.05≦p<0.1, and the p-value was expressed accordingly.

[0127] (3) Test-1: Effect on melanin production A melanocyte-containing three-dimensional skin model was cultured under various addition conditions, and the amount of melanin produced was measured.

[0128] The amount of melanin produced was measured according to the following method. After culturing, the skin model was washed with PBS(-), immersed in 0.1N NaOH overnight at 60°C for extraction, and then centrifuged to remove the film and insoluble fraction contained in the skin model as a precipitate, thereby obtaining a solution fraction. The absorbance (405 nm) of the obtained solution fraction was measured and determined as the amount of melanin produced. Next, the melanin suppression rate for each group was calculated using the following formula, assuming that the amount of melanin produced in the "control" was 100%. Melanin suppression rate (%) = 100 - 100 * (absorbance of each group / absorbance of control)

[0129] The results are shown in Figure 1. A higher melanin suppression rate indicates a higher melanin suppression effect. As shown in FIG. 1, the melanin-suppressing effect was observed when the positive control (VC), longan berry, and pyrroloquinoline quinone were administered alone. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone. In particular, it was a very surprising finding that the combination of longan and pyrroloquinoline quinone, even when the total amount was low, exhibited a better melanin suppression effect than the addition of higher amounts of longan or pyrroloquinoline quinone alone.

[0130] Although data is not shown, when the positive control sample was combined with longan berry or pyrroloquinoline quinone, no synergistic effect was observed at the small doses mentioned above, and the effect was the same as when the same amount of each was administered alone.

[0131] (4) Test-2: Effect on nicotinamide phosphoribosyltransferase (NAMPT) production A melanocyte-containing three-dimensional skin model was cultured under various addition conditions, and the amount of NAMPT produced was measured.

[0132] The amount of NAMPT produced was measured by the following method. After culturing, the culture medium of the skin model was treated using a spin column (10kD Spin Columns, manufactured by Abcam) to remove proteins, and the eluate containing the low molecular weight fraction was collected. The amount of NAMPT produced from the obtained low molecular weight fraction was quantified using the "NAD / NADH Assay Kit" (Abcam).

[0133] The results are shown in Figure 2. The higher the amount of NAMPT produced, the greater the anti-aging effect on the skin can be expected. As shown in FIG. 2, the administration of longan berry and pyrroloquinoline quinone alone was found to have the effect of improving NAMPT production. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone. In particular, it was a very surprising finding that the combination of longan and pyrroloquinoline quinone, even when the total amount was low, had a good effect on improving NAMPT production compared to the addition of higher amounts of longan or pyrroloquinoline quinone alone.

[0134] (5) Test-3: Nicotinamide adenine dinucleotide (NAD + )Effect on production volume We cultured a three-dimensional skin model containing melanocytes under various conditions and measured NAD + The amount of production was measured.

[0135] NAD + The amount of production was measured by the following method. After culturing, the culture medium of the skin model was treated using a spin column (10kD Spin Columns, manufactured by Abcam) to remove proteins, and the eluate containing the low molecular weight fraction was collected. The obtained low molecular weight fraction was analyzed for NAD using the "NAD / NADH Assay Kit" (Colorimetric). +The production amount was quantified.

[0136] The results are shown in Figure 3. NAD + The higher the production amount, the greater the anti-aging effect on the skin can be expected. As shown in Figure 3, the administration of longan and pyrroloquinoline quinone alone significantly increased NAD + The effect of improving production volume was observed. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone. In particular, the combination of longan berry and pyrroloquinoline quinone, even when the total amount is low, exhibits a better NAD than the addition of higher amounts of longan berry or pyrroloquinoline quinone alone. + The fact that the production amount was improved was a very unexpected finding.

[0137] <Test using a melanin-containing 3D skin model-2> Using the following method, various evaluations were carried out under various addition conditions using a melanin-containing three-dimensional skin model.

[0138] (1) Sample preparation The following three types of samples were prepared and subjected to the culture test described below.

[0139] (1-1) Longan fruit sample Longan extract powder (manufactured by Matsuura Pharmaceutical Co., Ltd.) was prepared. This sample was prepared from the arils of longan fruit (scientific name: Euphoria longana (Sapindaceae)), which were extracted with water, filtered, concentrated (with pectinase treatment), and sterilized (125±10°C), followed by the addition of dextrin, spray drying, and sieving (sieve: 0.5 mm opening, with heavy metal removal by magnetic treatment). This resulted in a powder sample. The longan extract powder used in this example contains an excipient (dextrin) as well as components derived from longan. Therefore, the longan concentration hereinafter indicates the amount (dry mass) converted to components derived from longan.

[0140] (1-2) Pyrroloquinoline quinone sample Pyrroloquinoline quinone disodium salt (purity 99.0% or higher, manufactured by Ryusendo Co., Ltd.) was prepared. Pyrroloquinoline quinone disodium salt will hereinafter also be referred to as "PQQ." This sample is a powdered sample obtained by fermentation production using microbial fermentation and purification processes (hydroethanol extraction, concentration, drying, filtration, etc.). Hereinafter, the concentration of the pyrroloquinoline quinone sample (sodium salt) is expressed as the amount converted into the free pyrroloquinoline quinone.

[0141] (1-3) Positive control sample As a positive control substance, L-(-) ascorbic acid (powder sample, manufactured by Kanto Chemical Co., Ltd.) was prepared. Hereinafter, L-(-) ascorbic acid will also be referred to as "VC."

[0142] (2) Culture test In this example, the following culture methods (system using a three-dimensional skin model) were commonly adopted.

[0143] The culture was carried out in an incubator under specified ingredient addition conditions for 3 weeks at 37°C and 5% CO2 using a melanocyte-containing three-dimensional skin model ("MEL-300", manufactured by Kurabo Industries, Ltd.).

[0144] The culture medium used was EPI-100NMM113, which contains melanin-stimulating factors bFGF, α-MSH, and KGF. These components function as melanin-forming factors, pro-inflammatory factors, and external stimulatory factors in this culture system. The medium was changed every other day, and each time the test substance was dissolved in the medium, the biological activity of the test substance was evaluated from the bottom side of the skin model, which is intended for food use.

[0145] The conditions for adding ingredients to the medium are as follows. Control: Medium only VC(100): Positive control sample 100μg / ml Longan (15): Longan sample 15 μg / ml Longan (50): Longan sample 50μg / ml Longan (100): Longan sample 100 μg / ml PQQ(2.5): Pyrroloquinoline quinone sample 2.5μg / ml PQQ(3): Pyrroloquinoline quinone sample 3 μg / ml PQQ(5): Pyrroloquinoline quinone sample 5μg / ml PQQ(10): Pyrroloquinoline quinone sample 10 μg / ml Longan (50) + PQQ (5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 5 μg / ml Longan (15) + PQQ (2.5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 2.5 μg / ml

[0146] Four wells were set up for each sample (n=4).

[0147] Statistical analysis of the results was performed using the Dunnett test as a multiple comparison test to determine whether or not there was a significant improvement compared to the control group, and the results were expressed as *p<0.05, **p<0.01. Note that a significant trend was determined to exist when 0.05≦p<0.1, and the p-value was expressed accordingly.

[0148] (3) Test-1: Effect on IL-1β production A melanocyte-containing three-dimensional skin model was cultured under various addition conditions, and the amount of IL-1β produced was measured.

[0149] The amount of IL-1β produced was measured by the following method. 100 μl of the culture medium was collected, and the amount of IL-1β produced was measured using "IL-1β, ELISA kit, Human" (ENZ).

[0150] The results are shown in Figure 4. A lower value for the amount of IL-1β produced means a higher effect of suppressing melanin production. As shown in FIG. 4, the IL-1β suppression effect was observed when longan berry and pyrroloquinoline quinone were administered alone. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone.

[0151] (4) Test-2: Effect on the production of GDF-15 and NLRP3 A melanocyte-containing three-dimensional skin model was cultured under various addition conditions, and the production levels of GDF-15 and NLRP3 were measured.

[0152] The production levels of GDF-15 and NLRP3 were measured according to the following method. After the culture, the skin model was washed with PBS(-), and 250 μL of "T-PER (trademark) Tissue Protein Extraction Reagent buffer" (Thermo Scientific) was added, followed by crushing and extraction at 4°C to prepare a lysate. The crushing and extraction was carried out as follows: the skin model and two glass beads were placed in a polypropylene tube, followed by the addition of T-PER Buffer. Extraction was then carried out using a crushing and extracting device (a micro-smash MS-100R bead-type cell crusher manufactured by Tomy Seiko) at 4°C, 2000 rpm, and for 5 minutes.

[0153] Using a portion of the lysate (20 μL), the total protein concentration was quantified by relative comparison with BSA (bovine serum albumin) using the standard BCA protein assay method (measured at OD562 nm).

[0154] The lysate (100 μL) was subjected to the Quantikine Human GDF-15 ELISA Kit and the Human NLRP3 ELISA Kit (ab274401, Abcam), and the amount of GDF-15 protein and NLRP3 protein per unit protein was quantified.

[0155] The results for GDF-15 are shown in Figure 5. The lower the amount of GDF-15 produced, the greater the anti-aging effect can be expected. As shown in FIG. 5, the GDF-15 suppression effect was observed when longan fruit and pyrroloquinoline quinone were administered alone. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone. In particular, it was a very surprising finding that the combination of longan and pyrroloquinoline quinone, even when the total amount was low, exhibited a better GDF-15 inhibitory effect than the addition of higher amounts of longan or pyrroloquinoline quinone alone.

[0156] The results for NLRP3 are shown in Figure 6. The lower the amount of NLRP3 produced, the greater the effect of suppressing melanin production can be expected. As shown in Figure 6, the NLRP3 suppression effect was observed when longan fruit and pyrroloquinoline quinone were administered alone. However, this effect was significantly enhanced by the combination of longan and pyrroloquinoline quinone. In particular, it was a very surprising finding that the combination of longan and pyrroloquinoline quinone, even when the total amount was low, exhibited a better NLRP3 inhibitory effect than the addition of higher amounts of longan or pyrroloquinoline quinone alone.

[0157] <Test using human skin fibroblasts-1> Various evaluations were carried out under various addition conditions using human skin fibroblasts according to the following method.

[0158] (1) Sample preparation As in the above "Test using a melanin-containing three-dimensional skin model," a longan berry sample and a pyrroloquinoline quinone sample were prepared. The standards for expressing the concentration of each sample are the same as those in the "Test using a melanin-containing three-dimensional skin model" above.

[0159] (2) Culture test In this example, the following culture method (system using human skin fibroblasts) was commonly adopted.

[0160] (2-1) Preparation of human dermal fibroblasts Human dermal fibroblasts were subcultured in DMEM medium containing 5% FBS, and the cells that reached the population doubling level (PDL) determined for each test were used below.

[0161] Hereinafter, the PDLs in the present examples were all determined by the following method. At the end of each subculture, the total number of cells in the culture was counted, and the cell proliferation rate and population doubling time were then calculated using the least squares method. From these values, the PDL was estimated for each cell passage. The cell proliferation rate and population doubling time were calculated based on the following formulas.

[0162] Growth rate μ=log(N(t))-log(N(t0)) / (t-t0) Population doubling time x=log2 / μ (t: most recent subculture time t0: Culture time before passage 1 t-t0: culture time)

[0163] Human dermal fibroblasts in a normal state (cultured at approximately PDL20, with no signs of cellular senescence) proliferated from seeded cells of 13,000 to recovered cells of 55,000 after 72 hours of culture. Based on this, the following formula was used to define the proliferation rate from the early to mid-stages of culture as "34.6 hours," and the PDL (population doubling number) was calculated based on that culture time. μ=log(55000)-log(N(13000) / (72-0)) x=log2 / 0.0087=34.6 hours

[0164] Furthermore, it was confirmed that the population doubling time was delayed with increasing passage number, and that the cell proliferation rate decreased with increasing passage number, which was the cause of cellular senescence.

[0165] (2-2) Tracking the expression levels of Per2 and Sirt1 A test was conducted to track the expression levels of Per2 and Sirt1 at PDL50-60.

[0166] Per2 is known as a clock gene, and its gene expression cycle usually fluctuates on a 24- to 25-hour cycle. It is thought that the amount of Per2 protein expression is also affected by this gene expression cycle. Therefore, in this study, before adding PQQ or longan nikomi, cells were treated with high serum culture conditions (40% horse serum, Heat-Inactivated Horse Serum, Gibco HI Horse Serum) for 2 hours to synchronize the expression cycle of clock genes. PQQ or longan nikomi was then added, and changes in Per2 production were monitored.

[0167] Specifically, human dermal fibroblasts were subcultured, and when they reached PDL50, 100,000 cells / well were seeded onto a 6-well plate. After the cells settled, they were treated with 40% horse serum to synchronize circadian rhythm-related expression. After 2 hours, the horse serum was removed and replaced with serum-free medium (Gibco D-MEM, Fujifilm Wako Pure Chemical Industries), to which PQQ and longan nutrient were added. After adding PQQ or longan nut, the cells were cultured for 72 hours at 37°C in a 5% CO2 incubator. After the culture was completed, a portion of the NAD +The culture medium was collected to measure production levels. The medium was removed from the remaining culture medium, washed with PBS(-), and the cells were collected with a scraper. The cell suspension was centrifuged to prepare a cell pellet, which was then solubilized in T-PER buffer (Tissue Protein Extraction Reagent (Thermofisher)) to prepare a cell lysate. A portion of the cell extract (20 μL) was used to quantify protein (total protein amount) using the standard BCA protein assay method (measured at OD562 nm), and the total protein concentration was quantified by relative comparison with BSA (bovine serum albumin). The remaining cell extract (100 μL) was subjected to quantification of the protein expression levels of Per2 and Sirt1 using a commercially available ELISA kit. The ELISA kits used are as follows: ·Per2:human Per2 ELISA kit Human PER2(Period circadian protein homolog 2) ELISA Kit(Assay Genie) ·Sirt1:human Sirt1 ELISA kit Human SIRT1 ELISA Kit (ab171573)(abcam)

[0168] The conditions for adding ingredients to the medium are as follows. Control: Medium only Longan (100): Longan sample 100 μg / ml Longan (300): Longan sample 300 μg / ml PQQ(3): Pyrroloquinoline quinone sample 3 μg / ml PQQ(10): Pyrroloquinoline quinone sample 10 μg / ml PQQ(30): Pyrroloquinoline quinone sample 30μg / ml Longan (150) + PQQ (5): Longan sample 150 μg / ml and pyrroloquinoline quinone sample 5 μg / ml

[0169] Four wells were set up for each sample (n=4).

[0170] Statistical analysis of the results was performed using the Dunnett test as a multiple comparison test to determine whether or not there was a significant improvement compared to the control group, and the results were expressed as *p<0.05, **p<0.01. Note that a significant trend was determined to exist when 0.05≦p<0.1, and the p-value was expressed accordingly.

[0171] The results for Sirt1 are shown in Figure 7. The results for Per2 are shown in Figure 8. The higher the amount of Sirt1 produced, the greater the anti-aging effect is expected to be. It is also known that Per2 expression level changes in response to fluctuations in Sirt1, and can form circadian rhythms. 7 and 8, administration of pyrroloquinoline quinone alone increased Sirt1 and decreased Per2, but these effects were more pronounced when administered in combination with longan fruit and pyrroloquinoline quinone.

[0172] (2-3) NAD + Measurement of production amount The culture medium collected in (2-2) above was subjected to protein removal using a spin column (10 kD Spin Columns, manufactured by Abcam), and the eluate of low molecular weight fractions was collected. The obtained low molecular weight fraction was analyzed for NAD using the "NAD / NADH Assay Kit" (Colorimetric). + The production amount was quantified.

[0173] Statistical analysis of the results was performed using the Dunnett test as a multiple comparison test to determine whether or not there was a significant improvement compared to the control group, and the results were expressed as *p<0.05, **p<0.01. Note that a significant trend was determined to exist when 0.05≦p<0.1, and the p-value was expressed accordingly.

[0174] NAD + The results are shown in Figure 9. +The higher the production amount, the greater the anti-aging effect can be expected. As shown in Figure 9, administration of longan and pyrroloquinoline quinone alone significantly increased NAD + Almost no change in production amount was observed. However, the combination of longan and pyrroloquinoline quinone increases NAD + The production yield was significantly increased.

[0175] <Test using human skin fibroblasts-2> Various evaluations were carried out under various addition conditions using human skin fibroblasts according to the following method.

[0176] (1) Sample preparation As in the above "Test using a melanin-containing three-dimensional skin model," a longan berry sample and a pyrroloquinoline quinone sample were prepared. The standards for expressing the concentration of each sample are the same as those in the "Test using a melanin-containing three-dimensional skin model" above.

[0177] (2) Culture test In this example, the following culture method (system using human skin fibroblasts) was commonly adopted.

[0178] (2-1) Preparation of human dermal fibroblasts Human dermal fibroblasts were subcultured in DMEM medium containing 5% FBS, and the cells that reached the population doubling level (PDL) determined for each test were used below.

[0179] (2-2) Tracking Cellular Metabolism We conducted a study to track cell metabolism at PDL 80-90. ATP, lactate, cell viability, oxygen consumption rate, and cell senescence (SPiDER-βGal) were selected as indicators of cell metabolism. Cells with a PDL of 80 to 90 are considered to be in a state of cellular senescence (also called replicative senescence) in which cell proliferation is significantly reduced.

[0180] Specifically, human dermal fibroblasts that had reached PDL80-90 were seeded at 20,000 cells / well on a 96-well plate, and PQQ and longan nut were added after the cells had settled. After adding PQQ or longan nut, the cells were cultured in a 5% CO2 incubator at 37°C for the incubation time set for each measurement item.

[0181] The conditions for adding ingredients to the medium are as follows. Control: Medium only Longan (100): Longan sample 100 μg / ml PQQ(10): Pyrroloquinoline quinone sample 10 μg / ml Longan (50) + PQQ (0.5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 0.5 μg / ml Longan (50) + PQQ (2.5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 2.5 μg / ml Longan (50) + PQQ (5): Longan sample 50 μg / ml and pyrroloquinoline quinone sample 5 μg / ml Longan (150) + PQQ (5): Longan sample 150 μg / ml and pyrroloquinoline quinone sample 5 μg / ml

[0182] Four wells were set up for each sample (n=4).

[0183] Statistical analysis of the results was performed using the Dunnett test as a multiple comparison test to determine whether or not there was a significant improvement compared to the control group, and the results were expressed as *p<0.05, **p<0.01. Note that a significant trend was determined to exist when 0.05≦p<0.1, and the p-value was expressed accordingly.

[0184] (2-2-1) ATP production After adding PQQ or longan nutmeg, the cells were cultured for 2 hours, the culture medium was removed, and the cells were washed with PBS(-). Next, 50 μL / well of ATP chemiluminescence reagent ("Cellular" ATP Measurement Reagent Ver. 2 (Toyo Viewnet)) was added to the cells in each well, and the reaction was allowed to proceed for 10 minutes at room temperature in the dark, and the chemiluminescence was quantified using a plate reader (SYNERGY H1, BioTek).

[0185] The results are shown in Figure 10. It can be expected that the higher the amount of ATP produced, the greater the anti-aging effect. As shown in FIG. 10, the combination of longan fruit and pyrroloquinoline quinone was found to have an effect of improving ATP production.

[0186] (2-2-2) Lactic acid level After adding PQQ or longan nutmeg, the cells were cultured for 2 hours and the culture medium was collected. The resulting culture medium was added to a new 96-well plate at 20 μL / well, and the absorbance (OD450 nm) was quantified using a 96-well plate reader with the "Lactate Assay Kit-WST" (Dojin). Based on the quantification results, the lactate concentration in the culture medium was quantified using a calibration curve.

[0187] The results are shown in Figure 11. The lower the lactic acid level, the greater the expected anti-aging effect. As shown in FIG. 11, the combination of longan fruit and pyrroloquinoline quinone was found to have a lactic acid level lowering effect.

[0188] (2-2-3) Oxygen consumption rate Immediately after the addition of PQQ or longan nut, the oxygen consumption rate derived from cellular respiration of oxidative phosphorylation, i.e., mitochondrial reactions, was measured over time using the "Extracellular OCR Plate Assay Kit" (Dojin). From 0 to 3 hours after addition, the fluorescence intensity at Ex=500 nm / Em=650 nm was analyzed kinetically using a fluorescence plate reader (SYNERGY H1, BioTek) and quantified as the extracellular oxygen consumption rate OCR (pmol / min).

[0189] The results are shown in Figure 12. The higher the oxygen consumption rate, the stronger the anti-aging effect can be expected. As shown in FIG. 12, the administration of longan berry or pyrroloquinoline quinone alone did not decrease the oxygen consumption rate. However, the combination of longan and pyrroloquinoline quinone significantly increased the oxygen consumption rate.

[0190] (2-2-4) Cell viability (MTT measurement) After adding PQQ or longan nutmeg, the cells were cultured for 72 hours, and the culture medium was removed and then washed with PBS(-). Next, 0.5 mg / ml MTT (Nacalai Tesque) was added, and the formazon reaction was carried out at 37°C for 2 hours. After the reaction was completed, the solution was removed, the cells were washed with PBS(-), and 100 μL / well of 2-propanol was added. The formazon formed by the reaction of MTT was quantified at OD570nm using an absorbance plate reader, and the cell viability was calculated as a ratio to the control value.

[0191] The results are shown in Figure 13. The higher the cell viability, the stronger the anti-aging effect can be expected. As shown in FIG. 13, the administration of longan berry or pyrroloquinoline quinone alone did not significantly change the cell viability. However, the combination of longan and pyrroloquinoline quinone significantly enhanced cell viability.

[0192] (2-2-5) Cell aging (SPiDER-βGal measurement) After 72 hours of culture following the addition of PQQ or longan nut, the degree of cellular senescence was quantified using the "Cellular Senescence Plate Assay Kit - SPiDER-βGal" (Dojin).

[0193] Specifically, first, the following two types of reagents were prepared in advance according to the kit instructions. "SPiDER-βGal DMSO stock solution": DMSO (125 μL) was added to a tube containing SPiDER-βGal and dissolved by stirring using a vortex mixer. "SPiDER-βGal working solution": "SPiDER-βGal DMSO stock solution" diluted 10 times with "Assay Buffer."

[0194] Next, after 72 hours of culture, the supernatant was removed by suction, and the plate was washed once with PBS. Then, 50 μL / well of "Lysis Buffer" was added, and the plate was incubated at room temperature for 10 minutes. The obtained lysate was added to each well of a 96-well clear bottom plate, and then 50 μL of "SPiDER-βGal working solution" was added to each well, followed by incubation at 37° C. for 30 minutes. Next, 100 μL of "Stop Solution" was added to each well, and measurements were taken using a fluorescent plate reader under conditions of Ex: 530 nm, Em: 580 nm to quantify the amount of β-galactosidase derived from the senescent cells.

[0195] The results are shown in Figure 14. The lower the SPiDER-βGal value, the stronger the anti-aging effect can be expected. As shown in FIG. 14, the administration of longan berry or pyrroloquinoline quinone alone caused almost no change in SPiDER-βGal. However, the combination of longan and pyrroloquinoline quinone significantly reduced the level of SPiDER-βGal.

Claims

1. A food and drink composition comprising longan fruit and pyrroloquinoline quinone.

2. The food and beverage composition according to claim 1, wherein the mass ratio of the component derived from longan berry to the pyrroloquinoline quinone is 5:1 to 30:

1.

3. An anti-aging agent comprising longan fruit and pyrroloquinoline quinone.

4. A mitochondrial dysfunction improving agent comprising longan berry and pyrroloquinoline quinone.

5. A nicotinamide phosphoribosyltransferase production promoter comprising longan berry and pyrroloquinoline quinone.

6. A nicotinamide adenine dinucleotide production promoter comprising longan berry and pyrroloquinoline quinone.

7. A sirtuin 1 production promoter comprising longan berry and pyrroloquinoline quinone.

8. A melanin production inhibitor comprising longan fruit and pyrroloquinoline quinone.

Images