Method for inhibiting the decline in antibacterial activity of honey and method for producing antibacterial agent

Abstract

To provide a honey having higher antibacterial activity against at least one or more kinds of bacterium selected from a group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis and Prevotella intermedia.SOLUTION: A method for suppressing reduction of antibacterial activity of honey, which includes heating of honey, and in which the heating temperature is 59°C or lower. The antibacterial activity is antibacterial activity against at least one or more kinds of bacterium selected from a group consisting of Fusobacteriumnucleatum, Porphyromonas gingivalis, and Prevotella intermedia.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a method for suppressing the decrease in antibacterial activity of honey and a method for producing an antibacterial agent. [Background technology]

[0002] Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia are known to be the main causative bacteria of periodontal disease. For example, Patent Document 1 discloses an oral antibacterial agent for inhibiting the growth of pathogenic bacteria in the oral cavity. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Japanese Patent Application Laid-Open No. 2016-113460 Summary of the Invention [Problem to be solved by the invention]

[0004] It is generally believed that honey is maintained at around 35°C while stored in a beehive due to the bees' body temperature and other factors. Honey is viscous and can crystallize during storage after harvesting from the hive. Therefore, to reduce viscosity and make it easier to handle, honey products are heated to 60°C or higher before distribution or packaging. In particular, if honey has crystallized during storage, it can be melted by heating it at a high temperature for a long period of time.

[0005] One aspect of the present invention aims to provide honey having higher antibacterial activity against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia. [Means for solving the problem]

[0006] The present inventors have discovered that the antibacterial activity of honey against specific periodontal disease bacteria can be maintained at a high level by maintaining the processing temperature below a certain level during the honey production process.

[0007] The method of the present invention for suppressing a decrease in the antibacterial activity of honey comprises heating honey to a heating temperature of 59°C or less, and the antibacterial activity is antibacterial activity against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia.

[0008] Preferably, the method includes storing the honey at or below 25°C before warming.

[0009] The present invention also provides a method for producing an antibacterial agent against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, which contains honey as an active ingredient, and which comprises heating honey at a heating temperature of 59°C or less.

[0010] The above method preferably includes storing the honey at 25°C or below before heating. [Effects of the Invention]

[0011] The present invention provides honey having higher antibacterial activity against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia. DETAILED DESCRIPTION OF THE INVENTION

[0012] Preferred embodiments of the present invention will be described in detail below, but the present invention is not limited to the following embodiments.

[0013] [Method for suppressing decline in antibacterial activity] The method for suppressing a decrease in the antibacterial activity of honey according to this embodiment includes heating honey to a temperature of 59° C. or less. The method for suppressing a decrease in the antibacterial activity of honey according to this embodiment can suppress a decrease in the antibacterial activity of honey against Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, which are known to cause periodontal disease.

[0014] Honey is produced primarily by bees using nectar collected from plant nectar, tree sap, secretions from insects parasitizing plants, and the like. The method for inhibiting a decrease in the antibacterial activity of honey according to this embodiment can be applied to any type of honey. Examples of honey include manuka honey, acacia honey, honeydew honey, clover honey, orange honey, astragalus honey, rosemary honey, sunflower honey, rapeseed honey, and coffee honey. The honey is preferably acacia honey or honeydew honey, and more preferably acacia honey. The origin of the honey is not particularly limited; for example, acacia honey may be from Romania, and honeydew honey may be from Bulgaria. One type of honey may be used alone, or two or more types may be used in combination.

[0015] The type of bee used to collect honey is not particularly limited. Honey can be obtained, for example, as a beekeeping product in accordance with conventional methods.

[0016] While honey is stored in beehives, it is generally maintained at around 35°C due to the body temperature of the bees. Honey collected from beehives is generally heated at a high temperature of around 60°C to reduce viscosity and facilitate transportation, filling, and other operations, and to melt crystals. In the method for suppressing a decrease in antibacterial activity according to this embodiment, the heating temperature is 59°C or lower. A heating temperature of 59°C or lower can suppress a decrease in the antibacterial activity of honey against Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia. In the method for suppressing a decrease in antibacterial activity according to this embodiment, it is preferable that the processing temperature be 59°C or lower in all processes after honey collection.

[0017] The honey heating temperature may be 59° C. or lower, 58° C. or lower, 57° C. or lower, 56° C. or lower, 55° C. or lower, 54° C. or lower, 53° C. or lower, 52° C. or lower, 51° C. or lower, 50° C. or lower, 49° C. or lower, 48° C. or lower, 47° C. or lower, 46° C. or lower, 45° C. or lower, 44° C. or lower, 43° C. or lower, 42° C. or lower, 41° C. or lower, 40° C. or lower, 39° C. or lower, 38° C. or lower, 37° C. or lower, or 36° C. or lower. The honey heating temperature may be, for example, 35° C. or higher, 36° C. or higher, or 37° C. or higher. A honey heating temperature of 35° C. or higher is preferable in terms of ease of handling, as it reduces the viscosity of the honey and makes it easier to reduce crystals in the honey.

[0018] The heating time of honey can be appropriately changed depending on the viscosity, degree of crystallization, etc. of the honey. In this specification, the heating time of honey refers to the cumulative time when the honey is heated in multiple stages. The heating time of honey may be, for example, 30 minutes or more, or may be 1 hour or more, 2 hours or more, 3 hours or more, 6 hours or more, 10 hours or more, 12 hours or more, 24 hours or more, 36 hours or more, or 48 hours or more. The heating time of honey may be, for example, 120 hours or less, 96 hours or less, or 72 hours or less. The honey heating step may be, for example, a step of heating at 35 to 52°C for 30 minutes to 96 hours, a step of heating at 35 to 50°C for 30 minutes to 96 hours, or a step of heating at 35 to 45°C for 30 minutes to 96 hours.

[0019] After harvesting, honey before heating is preferably stored at or below 25° C. After heating, honey may be stored again at or below 25° C. The storage temperature of honey before and after heating may be 24° C. or below, 23° C. or below, 22° C. or below, or 21° C. or below, or may be 15° C. or above, 18° C. or above, or 20° C. or above.

[0020] Before or after heating, the collected honey may be subjected to processes such as removal of dirt, beehive residue, filtering, extraction with an organic solvent, purification such as fractionation, and concentration.

[0021] In the method according to this embodiment, even if the viscosity of the honey is high or the honey contains a large amount of crystals, the honey can be transported, conveyed, filled, and other processes while maintaining the honey heating temperature at 59°C or below.

[0022] [Antibacterial agent] The method for producing an antibacterial agent according to this embodiment includes heating honey to a heating temperature of 59°C or less. The antibacterial agent has excellent antibacterial activity against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia. The antibacterial agent obtained by the production method according to this embodiment has higher antibacterial activity against Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia because it uses honey heated to a temperature of 59°C or less as an active ingredient.

[0023] The type of honey used as the active ingredient of the antibacterial agent according to this embodiment and the specific aspects of the treatment of the honey can be similar to those of the method for suppressing a decrease in antibacterial activity described above. Furthermore, the production method according to this embodiment may include suppressing a decrease in the antibacterial activity of honey against the bacteria, as described above in the method for suppressing a decrease in antibacterial activity.

[0024] The concentration of honey in the antibacterial agent may be, for example, 0.1% by weight or more, 1% by weight or more, 3% by weight or more, 5% by weight or more, 7% by weight or more, 10% by weight or more, 15% by weight or more, 20% by weight or more, 25% by weight or more, 30% by weight or more, 35% by weight or more, 40% by weight or more, 45% by weight or more, 50% by weight or more, 55% by weight or more, 60% by weight or more, 65% by weight or more, 70% by weight or more, 75% by weight or more, 80% by weight or more, 85% by weight or more, 90% by weight or more, 95% by weight or more, 97% by weight or more, 98% by weight or more, 99% by weight or more, 99.5% by weight or more, or 100% by weight. The honey concentration in the antibacterial agent is, for example, 99.5% by mass or less, 99% by mass or less, 98% by mass or less, 97% by mass or less, 95% by mass or less, 90% by mass or less, 85% by mass or less, 80% by mass or less, 75% by mass or less, 70% by mass or less, 65% by mass or less, 60% by mass or less, 55% by mass. The following may be 50% by mass or less, 45% by mass or less, 40% by mass or less, 35% by mass or less, 30% by mass or less, 25% by mass or less, 20% by mass or less, 15% by mass or less, 10% by mass or less, 7% by mass or less, 5% by mass or less, 3% by mass or less, 2% by mass or less, or 1% by mass or less.

[0025] The antibacterial agent obtained by the production method of this embodiment has high antibacterial activity against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, and can therefore be used, for example, as an agent for preventing or treating periodontal disease.

[0026] Volatile sulfur compounds produced by periodontal disease bacteria are known to be involved in bad breath. Therefore, the antibacterial agent of this embodiment can also be used as an agent for preventing bad breath. Periodontal disease bacteria such as Fusobacterium nucleatum are also known to be involved in the growth of colon cancer. Therefore, it is believed that the antibacterial agent of this embodiment can also be suitably used for the prevention or treatment of colon cancer.

[0027] The antibacterial agent obtained by the production method according to this embodiment can be used as a pharmaceutical, quasi-drug, cosmetic, or food composition, or can also be used as an ingredient in a pharmaceutical, quasi-drug, cosmetic, or food composition. Honey, which is an active ingredient in the antibacterial agent, is also suitable for use as a food, so the antibacterial agent is suitable for oral administration. The antibacterial agent may be applied to the affected area, such as the gums. When applying the antibacterial agent, it may be diluted appropriately before use. The antibacterial agent according to this embodiment may be administered once a day, or in multiple doses, such as twice or three times a day.

[0028] The subject to which the antibacterial agent is administered may be, for example, a patient with periodontal disease or a person at risk of periodontal disease, or a healthy person.

[0029] The antibacterial agent according to this embodiment may further contain other components, such as pharmaceutically acceptable components (e.g., excipients, binders, lubricants, disintegrants, emulsifiers, surfactants, bases, solubilizers, and suspending agents) and food acceptable components (e.g., minerals, vitamins, flavonoids, quinones, polyphenols, amino acids, nucleic acids, essential fatty acids, cooling agents, binders, sweeteners, disintegrants, lubricants, colorants, flavorings, stabilizers, preservatives, sustained-release regulators, surfactants, solubilizers, and wetting agents).

[0030] The pharmaceuticals, quasi-drugs, and cosmetics may be in the form of, for example, a solution, emulsion, ointment, cream, paste, jelly, aerosol, powder, oil, solid, foam, etc. Examples of quasi-drugs and cosmetics include mouth rinse, mouthwash, oral cosmetics, dentifrices, etc. These various preparations can be prepared, for example, by mixing the antibacterial agent obtained by the above-mentioned method with other ingredients as necessary and shaping the mixture into the above-mentioned dosage form.

[0031] When the antibacterial agent is used as a food composition or as one of its components, the food composition preferably emphasizes the tertiary function of food, i.e., the function of regulating physical condition. Examples of products that emphasize the tertiary function of food include health foods, functional foods, foods with nutrient claims, nutritional supplements, dietary supplements, and foods for specified health uses.

[0032] Examples of foods include the following, and foods used for the above purposes can be obtained by mixing the honey or antibacterial agent with intermediate products in the manufacturing process or with final products: beverages such as soft drinks such as coffee, juice, and tea drinks, milk drinks, lactic acid bacteria drinks, yogurt drinks, carbonated drinks, and alcoholic beverages such as sake, Western liquor, fruit wine, and mead; spreads such as custard cream; pastes such as fruit paste; sweets such as chocolate, donuts, pies, cream puffs, gum, jelly, candy, cookies, cakes, and puddings; Japanese sweets such as daifuku, mochi, manju, castella, anmitsu, and yokan; frozen desserts such as ice cream, popsicles, and sorbet; cooked foods such as curry, beef bowls, rice porridge, miso soup, soup, meat sauce, pasta, pickles, and jam; and seasonings such as dressings, furikake, umami seasonings, and soup bases.

[0033] When the antibacterial agent obtained by the method according to this embodiment is used as a pharmaceutical, quasi-drug, cosmetic, or food composition, or as one of their components, it is preferable that the processing temperature of these products is 59°C or less throughout all manufacturing steps after the antibacterial agent is incorporated. [Example]

[0034] The present invention will be explained in more detail below with reference to examples, but the present invention is not limited to these examples.

[0035] The antibacterial activity of honey against several periodontal pathogens was investigated in relation to the honey processing temperature.

[0036] The honey used was Romanian acacia honey and Bulgarian honeydew honey stored at 20°C. Approximately 2g of the acacia honey stored at 20°C was taken and left to stand for 1 hour in a thermostatic water bath set at 35°C, 45°C, or 60°C. Approximately 2g of the honeydew honey stored at 20°C was taken and left to stand for 1 hour in a thermostatic water bath set at 45°C or 60°C. After heating, the honey was stored at room temperature in the dark.

[0037] The heated honey was dissolved in a broth medium to prepare a honey-containing medium with a honey concentration of 20% by mass. The honey-containing culture solution was passed through a 0.2 μm filter and then further diluted with broth medium to prepare honey-containing media with honey concentrations of 2.5%, 5%, and 10% by mass.

[0038] The bacteria used in the study were as follows: Fusobacterium nucleatum CCUG9126 and Porphyromonas gingivalis CCUG25893 were obtained from the CCUG (Culture Collection, University of Gothenburg). Prevotella intermedia NCTC13070 was obtained from the NCTC (National Collection of Type Cultures). These bacteria were stored at -80°C until testing.

[0039] The anaerobic bacteria Porphyromonas gingivalis, Prevotella intermedia, and Fusobacterium nucleatum were cultured on blood agar at 35–37°C for 2–5 days in an anaerobic chamber (80% N2 + 10% CO2 + 10% H2).

[0040] After culturing, pure colonies were picked from the agar medium and inoculated into the following broth media to obtain approximately 10 8 Bacterial suspensions were prepared in CFU / mL in enriched Clostridium broth for Fusobacterium nucleatum and Brucella broth (containing hemin and vitamin K1) for Porphyromonas gingivalis and Prevotella intermedia.

[0041] For Porphyromonas gingivalis and Fusobacterium nucleatum, 10 μL of the culture medium containing the bacteria prepared above was mixed with 290 μL of honey-containing medium and incubated under static anaerobic conditions at 35-37°C for 24 hours. Each test was performed five times, and 0.1 mL of the serial dilutions were plated on agar medium, cultured, and the resulting viable bacterial counts (CFU / mL) were counted. As a negative control, the test was performed under the same conditions except for the absence of honey, and the viable bacterial counts (CFU / mL) were also counted.

[0042] The growth inhibitory activity of each honey against each bacterium was calculated using the following formula: The growth inhibitory activity of each honey against Fusobacterium nucleatum is shown in Table 1, and the growth inhibitory activity of each honey against Porphyromonas gingivalis is shown in Table 2. Growth inhibition activity (%) = 100 - [(number of bacteria growing on honey-containing medium / number of bacteria growing on negative control) x 100]

[0043] [Table 1]

[0044] [Table 2]

[0045] Both 35°C-treated acacia honey and 45°C-treated honeydew showed higher growth inhibitory activity against Fusobacterium nucleatum than the same honeys treated at 60°C at each concentration. 20% acacia honey treated at 35°C and 10% honeydew honey treated at 45°C also showed higher growth inhibitory activity against Porphyromonas gingivalis than honeys treated at 60°C.

[0046] For Prevotella intermedia, 40 μL of heated honey was mixed with 140 μL of broth medium and then further diluted two-fold with broth medium to prepare honey-containing medium at concentrations ranging from 22% to 2.75%. After passing the honey-containing culture medium through a 0.2 μm filter, 20 μL of honey-containing medium was mixed with 180 μL of the culture medium containing the bacteria prepared above and incubated at 35–37°C for 48 hours to determine the minimum honey concentration required to completely inhibit bacterial growth. The minimum inhibitory concentrations of acacia honey for Prevotella intermedia are shown in Table 3.

[0047] [Table 3]

[0048] Acacia honey treated at 35°C showed growth inhibitory activity against Prevotella intermedia, a periodontal pathogen, at lower concentrations than acacia honey treated at 60°C.

Claims

1. A method for suppressing a decrease in antibacterial activity of honey, comprising: This involves warming the honey. The heating temperature is 59°C or less, the honey processing temperature is 59°C or less, and the honey includes at least one of honeydew honey and acacia honey; The method, wherein the antibacterial activity is against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia.

2. 10. The method of claim 1, comprising storing the honey at or below 25°C prior to warming.

3. A method for producing an antibacterial agent against at least one bacterium selected from the group consisting of Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, which comprises using honey as an active ingredient, This involves warming the honey. The method, wherein the warming temperature is 59°C or less, the honey processing temperature is 59°C or less, and the honey comprises at least one of honeydew honey and acacia honey.

4. 4. The method of claim 3, comprising storing the honey at or below 25°C prior to warming.

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