Method for producing specific compound from black cumin seed

Abstract

To provide a method for producing a specific compound from black cumin seeds at a low cost.SOLUTION: The present invention provides a production method that includes: preparing residue remaining after extracting black cumin seed oil from black cumin seeds; and performing extraction using the residue as a raw material to create an extract containing one or more specific compounds or salts thereof. The production method yields the specific compounds or salts thereof.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a method for obtaining a specific compound from black cumin seeds, in particular, to a method for obtaining a compound having antibacterial properties, and further to a method for producing food and drink, which includes adding the obtained compound to food and drink. [Background technology]

[0002] Antibacterial agents or preservatives are additives that inhibit the growth of microorganisms that cause spoilage or deterioration of food and beverages, and improve the shelf life. For example, synthetic preservatives or synthetic antibacterial agents such as sodium benzoate and potassium sorbate are commonly used as antibacterial agents or preservatives for beverages. However, the effects of synthetic preservatives on the intestinal flora have been shown in recent years, and there are concerns about their effects on health. In addition to the effects on health, sodium benzoate has a unique astringent taste (Patent Document 1), which may affect the flavor of beverages, and potassium sorbate has a low growth inhibitory effect on certain microorganisms and also has a unique odor. In order to reduce the use of such synthetic preservatives, preservatives derived from natural materials are being considered.

[0003] On the other hand, the seeds of black cumin (Nigella sativa) (black cumin seeds) have been used since ancient times as a remedy for various diseases, and in recent years, the seeds and oil obtained from the seeds (black cumin seed oil) have been used as a medicinal spice, health food, or supplement. Thymoquinone, which is contained in the oil in the seeds, is known as the main active ingredient of black cumin seeds (Patent Document 2). [Prior art documents] [Patent documents]

[0004] [Patent Document 1] JP 2020-171301 A [Patent Document 2] Special Publication No. 2022-523705 Summary of the Invention [Problem to be solved by the invention]

[0005] The present inventors have searched and investigated antibacterial agents or preservatives derived from natural materials that can actually inhibit the growth of microorganisms in foods and beverages and that do not impair the commercial value of the foods and beverages, such as the stability of their appearance. As a result, they have discovered that black cumin seeds, which are natural materials, contain compounds with antibacterial properties. The present invention aims to provide a method for obtaining these compounds (including salts thereof) from black cumin seeds at a lower cost. [Means for solving the problem]

[0006] The present inventors have intensively studied a method for obtaining a compound having antibacterial properties and its salt from black cumin seeds, and as a result, have found that the target compound is contained in a larger amount in the cake (oil cake, residue) remaining after squeezing black cumin seed oil than in black cumin seed oil. And, it has been found that these compounds can be obtained efficiently by subjecting the residue after squeezing black cumin seed oil from black cumin seeds to an extraction process. Thymoquinone, the main active ingredient in black cumin seeds, is known to be mainly contained in the oil, and black cumin seeds are often distributed as black cumin seed oil obtained from seeds using an oil squeezing method such as cold pressing. Until now, the residue (oil cake) after obtaining oil by squeezing oil has hardly been used and has often been discarded. By using the residue after obtaining oil as a raw material, it has been successful in obtaining these compounds efficiently at a lower cost.

[0007] That is, the present invention relates to a method for producing a specific compound or a salt thereof, comprising preparing a residue after squeezing black cumin seed oil from black cumin seeds, and performing extraction using the residue as a raw material to obtain an extract containing one or more of the specific compounds or salts thereof. The specific compound or salt thereof is a compound represented by formula 1 or 2 described below and a salt thereof. This compound or salt thereof has antibacterial properties.

[0008] In the above-mentioned production method, after obtaining an extract containing one or more of the compounds or salts thereof, the extract may be fractionated to select a fraction containing one or more of the compounds or salts thereof. The selection of the fraction may be a selection of a fraction containing the compounds or salts thereof in a total concentration of 0.1% by mass or more.

[0009] In the above-mentioned production method, the residue remaining after squeezing black cumin seed oil from black cumin seeds preferably has a black cumin seed oil content of 35 mass % or less.

[0010] The compound or a salt thereof obtained by the above-mentioned production method can be used for, but is not limited to, being added to food and drink, and the compound or a salt thereof can be used for inhibiting the growth of one or more microorganisms of the genus Brettanomyces, Zygosaccharomyces, or Saccharomyces in food and drink.

[0011] The present invention also provides a method for producing a food or drink, which comprises producing the compound or a salt thereof by the above-mentioned production method, and adding the produced compound or a salt thereof to a food or drink. Effect of the Invention

[0012] In obtaining compounds having antibacterial properties from black cumin seeds, the cake (oil cake, residue) remaining after squeezing black cumin seed oil is used as a raw material, which allows these compounds to be obtained efficiently at low cost. Thymoquinone, the main active ingredient in black cumin seeds, is known to be mainly contained in the oil, and black cumin seeds have often been distributed as black cumin seed oil obtained from seeds using an oil squeezing method such as cold pressing. The residue (oil cake) obtained after squeezing oil has rarely been used and has often been discarded. The production method of the present invention, which uses the residue after oil squeezing as a raw material, is efficient, low-cost, and environmentally friendly from the viewpoint of reducing waste.

[0013] The obtained compounds (including their salts) can be used as antibacterial agents and preservatives derived from natural materials (black cumin seeds). In particular, in food and beverage applications, they can be used as a substitute for conventional synthetic preservatives such as sodium benzoate and potassium sorbate, or for the purpose of reducing the amount of these used. When synthetic preservatives are used, they are generally added to beverages at about 200 to 400 ppm, but the compounds obtained by the present invention may be able to exhibit a microbial growth inhibitory effect even at concentrations equal to or lower than those of synthetic preservatives. In addition, when these compounds are added to food and beverages, they tend not to change the appearance of the food and beverage and not to impair the commercial value of the food and beverage. When these compounds are used at a concentration at which an antibacterial effect is obtained, they may exhibit a degree of appearance stability that does not impair the commercial value, similar to the case of using sodium benzoate at the same concentration. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] The present invention relates to a method for producing a specific compound from black cumin seeds, specifically, the method includes preparing a residue after squeezing black cumin seed oil from black cumin seeds, and using the residue as a raw material to perform extraction to obtain an extract containing one or more of the compounds represented by formula 1 or 2 below or salts thereof.

[0015] Black cumin seeds are the seeds of black cumin (Nigella sativa). Black cumin, whose Japanese name is Nioi Kurotanesou, is an annual plant of the Nigella genus of the Ranunculaceae family. It is a plant that grows wild in areas such as the Middle East and Southwest Asia, and its seeds (black cumin seeds) and the oil obtained from the seeds (black cumin seed oil, sometimes simply referred to as "oil" in this specification) are distributed as spices, health foods, and supplements with various medicinal properties. Thymoquinone, known as the main physiologically active substance in black cumin seeds, is known to be contained in large amounts in the oil. The residue (oil cake) remaining after obtaining oil by oil extraction has not been used much until now. In the present invention, an antibacterial substance is extracted from this residue.

[0016] In the present invention, first, a residue after squeezing black cumin seed oil from black cumin seeds is prepared. "Preparing" generally means obtaining a residue after oil squeezing. For example, the residue may be obtained by squeezing black cumin seed oil from black cumin seeds, or the residue after oil squeezing may be purchased.

[0017] "Oil extraction" refers to obtaining oil from seeds. The oil extraction method is not particularly limited, and may be any of the following: a pressing method in which pressure is applied to the seeds, an extraction method in which the seeds are placed in a solvent and the oil is extracted into the solvent, and a pressing extraction method in which extraction is performed after pressing. However, among these, the pressing method is most preferable from the viewpoint of cost. When oil extraction is performed by pressing, a hot pressing method in which the seeds are heated and then pressed may be used, or a non-heat pressing method (cold press) in which the seeds are pressed without heating may be used.

[0018] Black cumin has a strong spice-like aroma, but by extracting the oil and using the residue, the aroma components can be removed together with the oil. By using this residue to extract antibacterial substances, it is possible to obtain antibacterial substances that do not affect the flavor when added to food and beverages.

[0019] The residual oil amount (i.e., black cumin seed oil content) of the "residue after oil extraction" (oil cake) is preferably 35% by mass or less, may be 30% by mass or less, or may be 25% by mass or less. The content of black cumin seed oil in the residue can be measured by the Soxhlet extraction method.

[0020] Next, the residue prepared above is used as a raw material to perform extraction, and an extract containing one or more compounds represented by formula 1 or formula 2 (including salts thereof) described later is obtained. The extraction can be performed by a conventional method, for example, by immersing the residue in an extraction solvent such as methanol for a certain period of time and then filtering the residue. Before the immersion, the residue may be crushed using a mill or crusher to increase the extraction efficiency, but is not limited thereto. When such a pretreatment is performed, a known method may be used. Examples of the solvent used for the immersion include, but are not limited to, organic solvents such as methanol, ethanol, propanol, and hexane. Of these, it is preferable to use methanol or ethanol, and in consideration of the final use as a food raw material, it is preferable to extract with ethanol. In addition, these organic solvents may be diluted with water to a concentration of 20 to 80 mass%, preferably 40 to 60 mass%, before use. The immersion time in the extraction solvent is preferably 1 hour or more. During the immersion, physical stimulation such as ultrasonic treatment or stirring may be applied. For the filtration after immersion, a known filter or the like can be used, but it is desirable to use a PTFE membrane filter with a pore size of about 0.4 to 2.0 μm, preferably about 0.5 to 1.4 μm. When filtering, it is desirable to perform pressure filtration. In addition, in order to improve filterability, rough filtration may be performed as a pretreatment using absorbent cotton or the like.

[0021] The compounds obtained by the present invention are one or more of compounds represented by the following formula 1 or 2 or salts thereof. Hereinafter, the compound represented by formula 1 will be referred to as "compound 1," and the compound represented by formula 2 will be referred to as "compound 2." In addition, "compound 1, compound 2, or one or more salts thereof" may be simply referred to as "compounds 1 and 2."

[0022] Compound 1 (CAS No. 27013-91-8) is a compound having the following formula 1:

[0023] [ka]

[0024] Compound 2 is a compound having the following formula 2:

[0025] [ka]

[0026] Compounds 1 and 2 may be in the form of any salt.

[0027] Compounds 1 and 2 have the effect of inhibiting the growth of microorganisms, that is, antibacterial properties. For example, when a microbial growth test is performed in which microorganisms are added to a food or drink and stored at room temperature for two weeks, it can be confirmed that the growth of microorganisms is inhibited when these compounds are added compared to when these compounds are not added. "The growth of microorganisms is inhibited" includes the fact that the growth of microorganisms is less than that of a comparative sample to which these compounds are not added. When microorganisms are added and stored at room temperature for two weeks, the fact that the growth of microorganisms is less than that of a comparative sample indicates that the growth of the added microorganisms is inhibited due to the presence of a compound having antibacterial properties in the sample. The room temperature is 10 to 30°C, preferably about 25 to 28°C. An example of a method for testing the growth of microorganisms is the method described in the Examples.

[0028] The production method of the present invention may include obtaining an extract containing compounds 1 and 2 from the residue, and then fractionating the extract to select a fraction containing compounds 1 and 2. Fractionation can be performed using a conventional method, such as using a known fractionation means such as column chromatography. As a packing material used in column chromatography, for example, a synthetic adsorbent made of styrene or divinylbenzene or an alkyl group-bonded silica gel-based adsorbent can be used, and for each packing material, a separation (development) solvent such as methanol / water, ethanol / water, or acetonitrile / water can be used. The fact that the fraction contains compounds 1 and 2 can be confirmed by using a conventional method used in the analysis of triterpene compounds. The obtained fraction may be further fractionated using high performance liquid chromatography (HPLC, etc.). When selecting a fraction, for example, a fraction containing a large amount of compounds 1 and 2 can be selected. For example, a fraction containing compounds 1 and 2 at a concentration of 0.1% by mass or more in total may be selected. This concentration may be 0.5% by mass or more, 1.0% by mass or more, 2.0% by mass or more, or 5.0% by mass or more. There is no particular limitation on the method for measuring Compounds 1 and 2. For example, a standard sample of each compound is prepared, and the compound can be measured using high performance liquid chromatography (HPLC) or the like.

[0029] Alternatively, when selecting fractions, each obtained fraction may be selected based on its antibacterial activity. Specifically, a fraction with high antibacterial activity may be selected. The method for testing the antibacterial activity (the ability to inhibit the growth of microorganisms) of each fraction is not particularly limited. As an example, a test such as that described in the Examples can be performed.

[0030] Compounds 1 and 2 obtained by the present invention may be in a state contained in an extract of the residue after squeezing black cumin seed oil, or may be in a concentrate obtained by concentrating the extract using a conventional means such as vacuum filtration, or may be in a state contained in a fraction obtained by fractionating the extract as described above. Compounds 1 and 2 may also be obtained by purifying the extract or the above fraction. From the viewpoint of cost and effectiveness, a fraction containing one or more of compounds 1 and 2 obtained by fractionating the extract is preferable.

[0031] In addition, the above-mentioned extract, concentrate, fraction, or compounds 1 and 2 obtained by purification may be formulated by a conventional method. For example, they may be dried, pulverized, and optionally molded into solid preparations such as powder, granules, and tablets, or may be dissolved or suspended in any solvent to form a liquid or paste preparation. When formulating, for example, various solvents, excipients, flavors, pH adjusters, antioxidants, stabilizers, specific gravity adjusters, and the like may be added depending on the application.

[0032] Compounds 1 and 2 obtained by the method of the present invention can be used for adding to food and beverages, but are not limited thereto. Compounds 1 and 2, when added to food and beverages, can exhibit the effect of suppressing the growth of microorganisms in the food and beverages (antibacterial properties). The microorganisms referred to here are microorganisms that cause the deterioration or decay of food and beverages, such as yeast. Typical examples of yeasts that cause the deterioration or decay of food and beverages include the genus Brettanomyces, the genus Zygosaccharomyces, and the genus Saccharomyces.

[0033] The type of food and drink to which the compounds 1 and 2 of the present invention are added is not particularly limited. For example, when the food and drink is a beverage, examples of the beverage include carbonated beverages, beverages containing fruit juice, beverages containing vegetable juice, coffee beverages, tea beverages, cocoa beverages, jelly beverages, dairy beverages, lactic acid bacteria beverages, soy milk, soups, nutritional drinks, sports beverages, alcoholic beverages, and concentrated beverages to be diluted and consumed. Among them, acidic beverages with a pH of 2 to 5 (preferably pH 2.5 to 4.5) are beverages in which synthetic preservatives have been used frequently in the past, and are therefore preferred as the type of beverage to which the compounds of the present invention are added. Examples of acidic beverages include carbonated beverages, beverages containing fruit juice, beverages containing vegetable juice, lactic acid bacteria beverages, nutritional drinks, and sports beverages. The beverages may be ready-to-drink beverages, or may be concentrated beverages to be diluted and consumed. The type of fruit juice or vegetable juice is not particularly limited, and examples thereof include citrus fruit juice (lemon, grapefruit, lime, oranges, Satsuma mandarins, tangor, summer mandarins, sweet summer, hassaku citrus, hyuganatsu, shekwasha, sudachi, yuzu, kabosu, daidai, iyokan, ponkan, kumquat, sanbokan, oroblanco, and pomegranate, etc.), black currant juice, apple juice, grape juice, peach juice, tropical fruit juice (pineapple, guava, banana, mango, acerola, papaya, passion fruit, lychee, etc.), plum juice, pear juice, apricot juice, plum juice, berry juice, kiwi fruit juice, cherry juice, chestnut juice, watermelon juice, tomato juice, carrot juice, strawberry juice, and melon juice.

[0034] Furthermore, when the food or drink is a food other than a beverage, the type is not particularly limited, and examples include sauces, sweets, bread, meat, minced fish, frozen foods, retort foods, brewed foods, seasonings, dairy products, and prepared foods.

[0035] The present invention also provides a method for producing a food or drink, which comprises adding the obtained compounds 1 and 2 to the food or drink. In the method for producing a food or drink of the present invention, "adding compounds 1 and 2 to the food or drink" preferably includes intentionally adding compounds 1 and 2 obtained by the present invention separately from the usual components of various foods and drinks, for the purpose of enhancing the microbial growth inhibitory effect of the food or drink. For example, it includes adding compounds 1 and 2 obtained by the present invention to foods and drinks that have conventionally used synthetic preservatives, instead of the synthetic preservatives or for the purpose of reducing the amount of synthetic preservatives used.

[0036] The timing of adding compounds 1 and 2 is not particularly limited, and they may be added at any stage of the manufacturing process of food and drink. For example, the antibacterial substance obtained by the present invention may be added and mixed when adding and mixing normal ingredients of food and drink, or the antibacterial substance obtained by the present invention may be added and mixed after normal blending of food and drink. The method of addition and mixing is not particularly limited. For example, the method of adding and mixing ingredients in normal manufacturing of food and drink may be used.

[0037] The amount of compounds 1 and 2 added to food and drink is not particularly limited as long as it is an amount that can obtain the microbial growth inhibitory effect in the food and drink. It can be appropriately set according to the type of food and drink and the degree of the desired microbial growth inhibitory effect, and also considering the influence on the flavor of the food and drink. For example, but not limited to, the total concentration of compounds 1 and 2 in the food and drink may be 0.1 ppm or more, 0.5 ppm or more, 0.8 ppm or more, 3 ppm or more, 10 ppm or more, or 20 ppm or more. In addition, in order to minimize the influence on the flavor and appearance of the food and drink, it is preferable to set the amount of addition to the food and drink as small as possible within the range in which the desired effect can be obtained. For example, the total concentration of compounds 1 and 2 in the food and drink is preferably 400 ppm or less, more preferably 300 ppm or less, more preferably 250 ppm or less, more preferably 125 ppm or less, more preferably 100 ppm or less, and more preferably 50 ppm or less. For example, when the food or drink is a concentrated beverage to be diluted and consumed, an amount that provides a microbial growth inhibitory effect in the concentrated beverage may be added with the intention of obtaining a microbial growth inhibitory effect during storage in a concentrated state. Alternatively, with the intention of obtaining a microbial growth inhibitory effect during storage of a diluted concentrated beverage, an amount obtained by multiplying the above amount by a predetermined dilution degree of the beverage may be added to the concentrated beverage so that the amount provides a microbial growth inhibitory effect in the diluted state according to a predetermined dilution degree of the concentrated beverage.

[0038] When the compounds 1 and 2 of the present invention are added to food and beverages, as described above, a fraction containing the compounds 1 and 2 obtained by fractionating an extract from the residue may be added to the food and beverage. As shown in the examples described below, the present inventors confirmed that when the fraction containing the compounds 1 and 2 obtained by the present invention is used in a beverage, a growth inhibitory effect against microorganisms (yeasts) of the genera Zygosaccharomyces, Saccharomyces, and Brettanomyces can be obtained at a very low concentration. The concentration of the fraction when added to a beverage is not particularly limited as long as it is an amount that can obtain a microbial growth inhibitory effect. It can be appropriately set taking into consideration the concentration of the compounds 1 and 2 contained in the fraction, the type of beverage, the degree of the desired microbial growth inhibitory effect, and the effect on the flavor of the beverage. For example, when a fraction containing compounds 1 and 2 at a total concentration of about 10% by mass is used, the concentration of the fraction in the beverage may be, but is not limited to, 1 ppm or more, 5 ppm or more, 8 ppm or more, 30 ppm or more, or 100 ppm or more. In addition, in order to minimize the influence on the flavor and appearance of the beverage, it is preferable to set the amount of the compound added to the beverage as small as possible within the range in which the desired effect can be obtained. For example, the concentration of the above fraction in the beverage is preferably 500 ppm or less, and may be 200 ppm or less.

[0039] Conventional synthetic preservatives such as sodium benzoate and potassium sorbate are generally used in beverages at a concentration of about 200 to 400 ppm. Compounds 1 and 2 obtained by the present invention can obtain a microbial growth inhibitory effect in beverages at a concentration equal to or lower than that of conventional synthetic preservatives. Compounds 1 and 2 also have the property of not impairing the commercial value of beverage products, such as the appearance stability and flavor.

[0040] The method for measuring the contents of Compounds 1 and 2 in foods and beverages is not particularly limited. For example, the contents can be measured by preparing a standard sample of each compound and setting conditions according to the type of food and beverage using HPLC or the like.

[0041] The food and drink obtained by the method of the present invention is preferably a food and drink that shows very little microbial growth when added with microorganisms and stored at room temperature for 2 weeks, and therefore does not deteriorate or spoil. More preferably, the food and drink does not show microbial growth.

[0042] The food and drink obtained by the method of the present invention preferably has an appearance stability to such an extent that the commercial value of the food and drink is not impaired. Appearance stability refers to the property that the appearance of the food and drink is unlikely to change over time. For example, it refers to the fact that the appearance of the food and drink does not change much when the food and drink is stored sealed in a packaging container or the like at room temperature for about two weeks.

[0043] The food and beverage obtained by the method of the present invention preferably contains compounds 1 and 2, but exhibits little change in color or taste from the original color or taste of the food and beverage (the color or taste of a food or beverage having the same composition except that the compounds are not added).

[0044] Compounds 1 and 2 obtained by the present invention can be preferably used to inhibit the growth of microorganisms in the above-mentioned foods and beverages, but may also be used in products other than foods and beverages. Examples of products in which compounds 1 and 2 can be used include kitchen detergents, laundry detergents, antibacterial sprays, toothpastes, cosmetics, household detergents, soaps, shampoos, and other household products, as well as pet foods.

[0045] Compounds 1 and 2 obtained by the present invention may be contained in foods and beverages or products other than foods and beverages as a flavor or as part of a flavor composition. EXAMPLES

[0046] Hereinafter, the details of the present invention will be specifically described with reference to experimental examples, but the present invention is not limited thereto. In addition, in this specification, unless otherwise specified, numerical ranges are described as including their endpoints.

[0047] Example 1: Extraction of antibacterial substances from black cumin seeds 1014 g of black cumin (Nigella sativa) seeds were crushed, heated to 200°C, and pressed with an oil press. As a result, 223 g of oil and 766 g of oil cake (residue after oil pressing) were obtained. The oil was filtered to obtain 147 g of oil filtrate and 29 g of oil residue. The total amount of triterpene glycosides including compounds 1 and 2 contained in the oil filtrate was measured by HPLC and found to be 124 mg. This corresponds to 0.0122% by mass of the initial amount of seeds (black cumin seeds).

[0048] On the other hand, the oil cake (residue) was immersed in an extraction solvent consisting of 3.15 L of methanol and 350 mL of water, and extraction was performed for 18 hours under ultrasonic treatment. The obtained extract was subjected to column chromatography (packing material: MCI GEL (trademark) CHP20P (75-150 μ), separation solvent: 50% methanol to 100% methanol) to obtain a triterpene glycoside-containing fraction. This fraction was further fractionated using HPLC to obtain fraction A containing compounds 1 and 2. The total amount of triterpene glycosides containing compounds 1 and 2 contained in fraction A was measured using HPLC and was found to be 2.17 g. This corresponds to 0.214 mass % relative to the amount of the initial seeds (black cumin seeds).

[0049] From the above, it was found that using oil cake (residue) made it possible to obtain larger amounts of triterpene glycosides, including compounds 1 and 2, than using black cumin seed oil for extraction.

[0050] In addition, the contents of compounds 1 and 2 in fraction A were measured by HPLC. The contents of compound 1 and its salts in fraction A were 9.34% by mass, and the contents of compound 2 and its salts in fraction A were 0.34% by mass.

[0051] <Example 2: Microbial growth test of fraction A containing compounds 1 and 2 in a carbonated drink containing lemon juice> A commercially available carbonated beverage containing 1% lemon juice (containing sugars (fructose glucose liquid sugar, sugar), lemon juice, carbonic acid, flavoring, vitamin C, acidulant, safflower color, calcium pantothenate, vitamin B6, and carotene color) was used to confirm the growth inhibitory effect of various yeasts in the beverage of fraction A obtained in Example 1 and sodium benzoate for comparison. The specific procedure is as follows.

[0052] The beverage was opened and left to stand until the carbon dioxide gas escaped, and then the pH of the beverage was adjusted to 3.3 using an aqueous sodium hydroxide solution. Fraction A was dissolved in dimethyl sulfoxide (DMSO), and the resulting solution was added to the beverage so that the final concentration of Fraction A was the concentration shown in Table 1. For comparison, an aqueous solution of sodium benzoate was added to the beverage so that the final sodium benzoate concentration was 400 ppm. A 96-well microplate was prepared, and 240 μL of the beverages to which Fraction A had been added at various concentrations and the beverage to which 400 ppm of sodium benzoate had been added were dispensed into each well.

[0053] Bacterial solutions of 10,000 cfu / mL were prepared for each of three types of yeast (Brettanomyces sp., Zygosaccharomyces bailii, and Saccharomyces cerevisiae).

[0054] 10 μL of each bacterial solution was added to each well to which the beverage containing fraction A or sodium benzoate was added. As a blank, 10 μL of saline was added to each well to which the beverage containing fraction A or sodium benzoate was added. In addition, as a group without antibacterial substance, 10 μL of each bacterial solution was added to 240 μL of beverage containing neither fraction A nor sodium benzoate.

[0055] The microplates prepared as described above were stored at 28°C for 2 weeks. After storage, it was confirmed that yeast had grown in the antibacterial substance-free area, and then yeast growth was visually confirmed in the beverages containing Fraction A or sodium benzoate, in comparison with the blank. Specifically, the presence or absence of yeast precipitation was visually confirmed, and those for which precipitation was confirmed were judged to have yeast growth (+), and those for which no precipitation was confirmed were judged to have no yeast growth (-). The results are shown in Table 1.

[0056] [Table 1]

[0057] From the results in Table 1, it can be seen that fraction A, which contains compounds 1 and 2 and was obtained by extraction and fractionation from oil cake (residue) of black cumin seeds, exhibits the effect of inhibiting the growth of various yeasts in existing carbonated drinks containing lemon juice at a lower concentration than sodium benzoate, which is known to have antibacterial properties. In particular, it shows the growth inhibitory effect against microorganisms of the genus Brettanomyces at a very low concentration.

[0058] Example 3: Evaluation of appearance stability in beverages containing fraction A The commercially available carbonated beverage containing 1% lemon juice used in Example 2 was used to evaluate the effects of Fraction A and sodium benzoate for comparison on the appearance of the beverage. The specific procedure is as follows.

[0059] The beverage was opened and left to stand until the carbon dioxide gas escaped, and then the pH of the beverage was adjusted to 3.3 using an aqueous sodium hydroxide solution. Fraction A was dissolved in DMSO, and the resulting solution was added to the beverage so that the concentration of Fraction A was the concentration shown in Table 2. For comparison, an aqueous solution of sodium benzoate was added to the beverage to a concentration of 400 ppm. The beverages to which Fraction A had been added at various concentrations and the beverage to which sodium benzoate had been added at 400 ppm were dispensed at 240 μL per well on a microplate.

[0060] Separately from the above, 240 μL of a beverage containing neither Fraction A nor sodium benzoate (additive-free group) was prepared as a control.

[0061] The centrifuge tubes prepared as described above were stored for 2 weeks at 28° C. After storage, the appearance of the beverages containing Fraction A or sodium benzoate was compared with that of the control and scored according to the following criteria. 0 points: No difference compared to the compound-free group 1 point: There is a slight difference compared to the compound-free area. 2 points: There is a clear difference compared to the compound-free area. 3 points: Significantly different from the compound-free group, and no commercial value as a beverage product. In the above criteria, a score of 0 to 2 means that the product is suitable for use as a beverage product (has commercial beverage value), and a score of 3 means that the product is not suitable for use as a beverage product (has no commercial beverage value). The results are shown in Table 2 ("-" in Table 2 means that the product has not been evaluated).

[0062] [Table 2]

[0063] The results in Table 2 show that fraction A, which contains compounds 1 and 2 and was obtained by extraction and fractionation from the oil cake (residue) of black cumin seeds, maintains the appearance stability without impairing the commercial value of the beverage even when used in a beverage at a concentration that exerts antibacterial activity.

[0064] <Reference Example: Microbial growth test in culture medium for Compounds 1 and 2> In order to confirm that each of Compounds 1 and 2 has antibacterial activity, the following tests were carried out.

[0065] Commercially available potato dextrose medium powder was dissolved in water and sterilized. After sterilization, the pH of the medium was adjusted to 3.8 using sulfuric acid. Compounds 1 and 2 were each dissolved in dimethyl sulfoxide (DMSO), and the resulting solutions of each compound were added in triplicate to the medium so that the final concentrations of each compound were 7.8 ppm, 31 ppm, and 125 ppm. A 96-well microplate was prepared, and 240 μL of the medium to which compounds 1 and 2 had been added at each concentration was dispensed into each well.

[0066] Bacterial solutions of 10,000 cfu / mL were prepared for each of three types of yeast (Brettanomyces sp., Zygosaccharomyces bailii, and Saccharomyces cerevisiae).

[0067] 10 μL of each yeast cell solution was added to wells to which media containing compounds 1 and 2 were added. As a blank, 10 μL of saline was added to wells to which media containing compounds 1 and 2 were added. As a compound-free group, 10 μL of each yeast cell solution was added to 240 μL of media to which neither compound 1 nor 2 was added.

[0068] These were stored at 28°C for 2 weeks. After storage, it was confirmed that yeast had grown in the compound-free area, and then the growth of yeast was visually confirmed in the medium to which each compound had been added, compared with the blank. The results are shown in Table 3. The numbers in Table 3 indicate the lowest concentration at which the yeast growth inhibitory effect was obtained among the concentrations of 7.8 ppm, 31 ppm, and 125 ppm prepared for each compound. For example, "31" indicates that the yeast growth inhibitory effect was not observed at a compound concentration of 7.8 ppm, but was obtained at 31 ppm and 125 ppm, and "7.8" indicates that the growth inhibitory effect was obtained at all of 7.8 ppm, 31 ppm, and 125 ppm. In addition, ">125" indicates that the yeast growth inhibitory effect was not obtained at all of 7.8 ppm, 31 ppm, and 125 ppm.

[0069] [Table 3]

[0070] The results in Table 3 show that both compounds 1 and 2 exhibit a growth inhibitory effect against certain yeasts and have antibacterial properties.

Claims

1. preparing a residue after expression of black cumin seed oil from black cumin seeds; and The residue was used as a raw material to perform extraction, and the resulting mixture was reacted with the following formula 1: 【Chemistry 1】 or a salt thereof, or a compound represented by the following formula 2: 【Chemistry 2】 Obtaining an extract containing one or more of the compounds represented by the formula: A method for producing the compound or a salt thereof, comprising:

2. The method according to claim 1 , wherein the compound or a salt thereof has antibacterial properties.

3. The method according to claim 1 or 2, further comprising, after obtaining an extract containing one or more of the compounds or salts thereof, fractionating the extract to select a fraction containing one or more of the compounds or salts thereof.

4. The method according to claim 3 , wherein the selection of the fraction comprises selecting a fraction containing the compound or a salt thereof at a total concentration of 0.1% by mass or more.

5. The method according to claim 1 or 2, wherein the residue after the oil expression has a black cumin seed oil content of 35% by mass or less.

6. The method according to claim 1 or 2, wherein the compound or a salt thereof is a compound or a salt thereof for addition to a food or drink.

7. The method according to claim 6, wherein the compound or its salt is a compound or its salt for inhibiting the growth of one or more microorganisms of the genus Brettanomyces, the genus Zygosaccharomyces, or the genus Saccharomyces in food or drink.

8. Producing a compound or a salt thereof as specified in claim 1 by carrying out the production method as described in claim 1 or 2; and adding the produced compound or a salt thereof to a food or drink; A method for producing a food or beverage comprising the steps of:

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