Saccharide-depletion agent for saccharide-containing food / beverage

Abstract

Provided is a technique for depleting saccharides in a saccharide-containing food / beverage. The provided technique causes the saccharides in a saccharide-containing food / beverage to be depleted by causing glucose oxidase, α-glucosidase, and / or β-D-fructofuranosidase to act concurrently or in no particular order on the saccharide-containing food / beverage or a feedstock for the saccharide-containing food / beverage. This technique allows the monosaccharides and disaccharides in a saccharide-containing food / beverage, in particular glucose and sucrose, to be depleted.

Description

Sugar-reducing agent for sugar-containing foods and beverages

[0001] The present technology relates to a sugar-reducing agent for sugar-containing foods and beverages. More specifically, the present invention relates to a sugar-reducing agent that can reduce monosaccharides and disaccharides in sugar-containing foods and beverages, a method for producing foods and beverages, and a method for reducing sugar in sugar-containing foods and beverages.

[0002] In recent years, growing health consciousness has led to an increasing demand for low-sugar beverages and reduced-sugar foods. Fruit and vegetable juices, in particular, are popular because of their high nutritional value, but their sugar content has raised concerns. Specifically, excessive sugar intake can pose health risks, increase calories, adversely affect dental health, cause a sudden rise in blood sugar levels, and disrupt the nutritional balance of beverages. Against this backdrop, efforts are underway to develop technologies to reduce the sugar content of sugar-containing foods and beverages.

[0003] For example, Patent Document 1 discloses a technology for producing a reduced-sugar citrus juice beverage by preparing a citrus juice beverage by mixing a citrus juice source having a low Brix value (low sugar content) with additional sinking pulp (or bottom pulp) and a sweetener that enhances the perceived sweetness characteristics of the juice beverage without significantly increasing the calorie content of the beverage.

[0004] Furthermore, Patent Document 2 discloses a technology for reducing the sugar content of a food material by contacting the food material with a glucosyltransferase having a specific sequence, and describes that the total combined content of monosaccharides and disaccharides in the food material can be reduced by 25%.

[0005] JP 2007-504821 A International Publication No. 2016 / 173929

[0006] As mentioned above, low-sugar beverages and reduced-sugar foods have been attracting attention in recent years due to growing health consciousness, but the reality is that further development of technologies to reduce the sugar content of sugar-containing foods and beverages is desired.

[0007] Therefore, the main object of the present technology is to provide a technology for reducing sugars in sugar-containing foods and beverages.

[0008] As a result of intensive research into technologies for reducing sugars in sugar-containing foods and beverages, the inventors of the present application discovered a new technology for reducing sugars in sugar-containing foods and beverages that uses two or more specific enzymes in combination, and have now completed this technology.

[0009] That is, the present technology first provides a sugar-reducing agent for sugar-containing foods and beverages, which contains glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase. The sugar-reducing agent according to the present technology can reduce glucose and sucrose in sugar-containing foods and beverages. The sugar-reducing agent according to the present technology can reduce sugars in sucrose-containing foods and beverages. The sugar-reducing agent according to the present technology can reduce sugars in beverages made from fruits and / or vegetables.

[0010] Next, the present technology provides a food or drink that uses the sugar-reducing agent according to the present technology.

[0011] The present technology further provides a method for producing a food or drink containing sugar, comprising the steps of: allowing glucose oxidase to act on a sugar-containing food or drink or a raw material for the sugar-containing food or drink; and allowing α-glucosidase and / or β-D-fructofuranosidase to act on a sugar-containing food or drink or a raw material for the sugar-containing food or drink; and a method for reducing sugar in a sugar-containing food or drink, comprising the steps of: allowing glucose oxidase to act on a sugar-containing food or drink or a raw material for the sugar-containing food or drink; and allowing α-glucosidase and / or β-D-fructofuranosidase to act on a sugar-containing food or drink or a raw material for the sugar-containing food or drink.

[0012] In this technology, "sugars" is a general term for monosaccharides and disaccharides (excluding sugar alcohols) as defined in the Nutrition Labeling Standards (Ministry of Health, Labour and Welfare Notification No. 176). Monosaccharides are sugars that cannot be further broken down, and include glucose, fructose, and galactose. Disaccharides include sucrose (glucose + fructose), maltose (glucose + glucose), and lactose (glucose + galactose).

[0013] Preferred embodiments for carrying out the present invention will be described below. Note that the embodiment described below is an example of a typical embodiment of the present invention, and the scope of the present invention should not be construed as being narrow.

[0014] 1. Sugar-Reducing Agent The sugar-reducing agent according to the present technology contains glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase as active ingredients. While glucose oxidase, α-glucosidase, and / or β-D-fructofuranosidase are each effective in reducing sugars in sugar-containing foods and beverages when used alone, as shown in the examples described below, the sugar-reducing effect can be significantly improved by using glucose oxidase in combination with α-glucosidase and / or β-D-fructofuranosidase. In particular, the sugar-reducing agent according to the present technology can reduce glucose and sucrose in sugar-containing foods and beverages.

[0015] The reduction rate of sugars (monosaccharides and disaccharides, the same applies below) when food and beverages are produced using the sugar-reducing agent of the present technology is not particularly limited, but is, for example, 30% or more, preferably 35% or more, more preferably 40% or more, and even more preferably 43% or more compared to the sugars in food and beverages produced without using the sugar-reducing agent of the present technology.

[0016] The reduction rates of glucose and sucrose when foods and beverages are produced using the sugar-reducing agent of the present technology are not particularly limited, but are, for example, 30% or more, preferably 35% or more, more preferably 40% or more, and even more preferably 43% or more, relative to the glucose and sucrose in foods and beverages produced without the sugar-reducing agent of the present technology. The respective reduction rates are also not particularly limited, but the reduction rate of glucose when foods and beverages are produced using the sugar-reducing agent of the present technology is, for example, 10% or more, 20% or more, preferably 30% or more, and more preferably 40% or more, relative to the glucose in foods and beverages produced without the sugar-reducing agent of the present technology. Furthermore, the reduction rate of sucrose when foods and beverages are produced using the sugar-reducing agent of the present technology is, for example, 40% or more, preferably 50% or more, and more preferably 60% or more, relative to the sucrose in foods and beverages produced without the sugar-reducing agent of the present technology.

[0017] Below, the components that can be used in the sugar-reducing agent according to the present technology will be described in detail.

[0018] (1) Glucose oxidase Glucose oxidase (EC 1.1.3.4) that can be used in the present technology is an enzyme that oxidizes glucose to produce gluconic acid and hydrogen peroxide. Note that the glucose oxidase that can be used in the present technology may also have other functions as long as it has glucose oxidase activity.

[0019] The origin of glucose oxidase that can be used in the present technology is not particularly limited, but examples thereof include glucose oxidases from filamentous fungi (e.g., Acremonium chrysogenum, Aspergillus aculeatus, Aspergillus niger, and Penicillium sp.), etc. Preferably, glucose oxidase is derived from a microorganism of the genus Aspergillus, and more preferably, glucose oxidase is derived from Aspergillus niger.

[0020] Here, "glucose oxidase derived from Aspergillus niger" means a glucose oxidase produced by a microorganism classified as Aspergillus niger (whether a wild-type strain or a mutant strain), or a glucose oxidase obtained by genetic engineering techniques using a glucose oxidase gene. Therefore, a recombinant produced by a host microorganism into which a glucose oxidase gene obtained from Aspergillus niger (or a modified version of said gene) has been introduced also falls under the category of "glucose oxidase derived from Aspergillus niger."

[0021] The glucose oxidase used in the present technology can be prepared from the culture medium of the microorganism from which the glucose oxidase is derived. Specific preparation methods include recovering glucose oxidase from the culture medium or cells of the microorganism. For example, when a glucose oxidase-secreting microorganism is used, the cells can be recovered from the culture medium in advance by filtration, centrifugation, or the like, as necessary, and the enzyme can then be separated and / or purified. When a glucose oxidase-nonsecreting microorganism is used, the cells can be recovered from the culture medium in advance by pressure treatment, ultrasonication, or the like, and the enzyme can then be extracted and / or separated and / or purified. The enzyme can be separated and / or purified by any known protein separation and / or purification method, without any particular limitation. Examples of the enzyme separation and / or purification method include centrifugation, UF concentration, salting out, and various chromatography methods using ion exchange resins, etc. The separated and / or purified enzyme can be powdered by drying methods such as lyophilization and vacuum drying, or by using an appropriate excipient and / or drying aid in the drying method. The isolated and / or purified enzyme can also be liquefied by adding an appropriate additive and sterilizing by filtration.

[0022] In the present technology, commercially available glucose oxidase can also be used, and preferred examples of commercially available glucose oxidases include those derived from Aspergillus niger and Acremonium chrysogenum, manufactured by Amano Enzyme Inc.

[0023] The content of glucose oxidase in the sugar-reducing agent according to the present technology can be freely set as long as it does not impair the effects of the present technology. The content of glucose oxidase can be set to, for example, 20 U or more per gram of sugar contained in the sugar-containing food or beverage or per gram of sugar contained in the raw materials of the sugar-containing food or beverage, and from the viewpoint of further enhancing the sugar-reducing effect, the content can be set to preferably 50 U or more, more preferably 100 U or more, 150 U or more, and even more preferably 180 U or more.

[0024] The upper limit of the glucose oxidase content is not particularly limited as long as it does not impair the effects of the present technology, but can be set to, for example, 25,000 U or less, 20,000 U or less, 10,000 U or less, 5,000 U or less, 3,000 U or less, 2,500 U or less, 2,000 U or less, 1,400 U or less, 1,200 U or less, 1,050 U or less, 900 U or less, 800 U or less, or 770 U or less per 1 g of sugars contained in the sugar-containing food or beverage or per 1 g of sugars contained in the raw materials of the sugar-containing food or beverage.

[0025] In this technique, the activity of glucose oxidase is defined as 1 unit (1 U) of the enzyme amount required to oxidize 1 μmol of glucose per minute.

[0026] (2) α-Glucosidase The α-glucosidase (EC.3.2.1.20) that can be used in the present technology is an enzyme that mainly hydrolyzes the α-1,4-glucosidic bond at the non-reducing end of maltooligosaccharides. Some α-glucosidases have strong glycosyltransferase activity, and some enzymes use maltose as a substrate to produce isomaltooligosaccharides with α-1,6-glucosidic bonds. Note that the α-glucosidase that can be used in the present technology may also have other functions as long as it has α-glucosidase activity.

[0027] The origin of α-glucosidase that can be used in the present technology is not particularly limited, and examples include α-glucosidases from filamentous fungi (e.g., the genera Absidia, Acremonium, and Aspergillus), yeasts (e.g., the genus Saccharomyces), actinomycetes (e.g., Streptomyces avermitilis, Streptomyces griseus, and Streptomyces violaceoruber), and bacteria (e.g., the genera Bacillus, Burkholderia ginsengisoli, Halomonas aquamarina, and Pseudomonas). Preferred examples include α-glucosidases derived from microorganisms of the genus Aspergillus, and more preferably α-glucosidases derived from Aspergillus niger.

[0028] Here, "α-glucosidase derived from Aspergillus niger" means an α-glucosidase produced by a microorganism classified as Aspergillus niger (whether a wild-type strain or a mutant strain), or an α-glucosidase obtained by genetic engineering techniques using an α-glucosidase gene. Therefore, a recombinant produced by a host microorganism into which an α-glucosidase gene obtained from Aspergillus niger (or a gene obtained by modifying said gene) has been introduced also falls under the category of "α-glucosidase derived from Aspergillus niger."

[0029] The α-glucosidase used in the present technology can be prepared from the culture medium of the microorganism from which the α-glucosidase is derived. Specific preparation methods include methods of recovering α-glucosidase from the culture medium or cells of the microorganism. For example, when an α-glucosidase-secreting microorganism is used, the cells can be recovered from the culture medium in advance by filtration, centrifugation, or the like, as necessary, and the enzyme can then be separated and / or purified. Furthermore, when an α-glucosidase-non-secreting microorganism is used, the cells can be recovered from the culture medium in advance by pressure treatment, ultrasonic treatment, or the like, as necessary, and the enzyme can then be extracted and / or separated and / or purified. The enzyme separation and / or purification method can be any known protein separation and / or purification method, without any particular limitation. Examples of such methods include centrifugation, UF concentration, salting out, and various chromatography methods using ion exchange resins, etc. The isolated and / or purified enzyme can be powdered by a drying method such as freeze-drying or vacuum drying, or can be powdered using an appropriate excipient and / or drying aid in the drying method. The isolated and / or purified enzyme can also be liquefied by adding an appropriate additive and sterilizing by filtration.

[0030] In the present technology, commercially available α-glucosidase can also be used, and a preferred example of a commercially available product is α-glucosidase derived from Aspergillus niger manufactured by Amano Enzyme Inc.

[0031] The content of α-glucosidase in the sugar-reducing agent according to the present technology can be freely set as long as it does not impair the effects of the present technology. The content of α-glucosidase can be set to, for example, 2000 U or more per gram of sugars contained in the sugar-containing food or drink or per gram of sugars contained in the raw materials of the sugar-containing food or drink, and from the viewpoint of further enhancing the sugar-reducing effect, the content can be set to preferably 4000 U or more, 6000 U or more, 8000 U or more, more preferably 10000 U or more, 15000 U or more, and even more preferably 18000 U or more.

[0032] The upper limit of the α-glucosidase content is not particularly limited as long as it does not impair the effects of the present technology, and can be set to, for example, 1,600,000 U or less, 1,300,000 U or less, 1,000,000 U or less, 500,000 U or less, 320,000 U or less, 240,000 U or less, 200,000 U or less, 170,000 U or less, 130,000 U or less, 100,000 U or less, 70,000 U or less, or 50,000 U or less per 1 g of sugars contained in the sugar-containing food or drink or per 1 g of sugars contained in the raw materials of the sugar-containing food or drink.

[0033] In the present technology, the activity of α-glucosidase is defined as one unit (1 U) of the enzyme that produces 1 μg of glucose per minute using α-methyl-D-glucoside as a substrate.

[0034] (3) β-D-fructofuranosidase The β-D-fructofuranosidase that can be used in the present technology is an enzyme that hydrolyzes the residue at the non-reducing end of β-D-fructofuranoside. Note that the β-D-fructofuranosidase that can be used in the present technology may also have other functions as long as it has β-D-fructofuranosidase activity.

[0035] The origin of β-D-fructofuranosidase that can be used in the present technology is not particularly limited, and examples include β-D-fructofuranosidases from filamentous fungi (e.g., Aspergillus aculeatus, Aspergillus awamori, Aspergillus niger, and Aspergillus japonicus), yeasts (e.g., Kluyveromyces lactis and Saccharomyces cerevisiae), and bacteria (e.g., the genus Arthrobacter and the genus Bacillus).

[0036] Here, "β-D-fructofuranosidase derived from Saccharomyces cerevisiae" means a β-D-fructofuranosidase produced by a microorganism classified as yeast (whether a wild-type strain or a mutant strain), or a β-D-fructofuranosidase obtained by genetic engineering techniques using a β-D-fructofuranosidase gene. Therefore, a recombinant produced by a host microorganism into which a β-D-fructofuranosidase gene obtained from Saccharomyces cerevisiae (or a modified version of said gene) has been introduced also falls under the category of "β-D-fructofuranosidase derived from Saccharomyces cerevisiae."

[0037] The β-D-fructofuranosidase used in the present technology can be prepared from the culture medium of the microorganism from which the β-D-fructofuranosidase is derived. Specific preparation methods include recovering the β-D-fructofuranosidase from the culture medium or cells of the microorganism. For example, when a β-D-fructofuranosidase-secreting microorganism is used, the cells can be recovered from the culture medium in advance by filtration, centrifugation, or the like, as necessary, and the enzyme can then be separated and / or purified. Furthermore, when a β-D-fructofuranosidase-non-secreting microorganism is used, the cells can be recovered from the culture medium in advance by pressure treatment, ultrasonic treatment, or the like, and the enzyme can then be extracted and / or separated and / or purified. The enzyme can be separated and / or purified by any known protein separation and / or purification method, without any particular limitation, and examples thereof include centrifugation, UF concentration, salting out, and various chromatography methods using ion exchange resins, etc. The isolated and / or purified enzyme can be powdered by a drying method such as freeze-drying or vacuum drying, or can be powdered using an appropriate excipient and / or drying aid in the drying method. The isolated and / or purified enzyme can also be liquefied by adding an appropriate additive and sterilizing by filtration.

[0038] In the present technology, commercially available β-D-fructofuranosidase can also be used, and a preferred example of a commercially available β-D-fructofuranosidase is β-D-fructofuranosidase derived from Saccharomyces cerevisiae manufactured by Sigma-Aldrich.

[0039] The content of β-D-fructofuranosidase in the sugar-reducing agent according to the present technology can be freely set as long as it does not impair the effects of the present technology. The content of β-D-fructofuranosidase can be set to, for example, 0.15 U or more per gram of sugar contained in the sugar-containing food or beverage or per gram of sugar contained in the raw materials of the sugar-containing food or beverage. From the viewpoint of further enhancing the sugar-reducing effect, the content can be set to preferably 7 U or more, 8 U or more, 9 U or more, 10 U or more, more preferably 15 U or more, 30 U or more, 50 U or more, 100 U or more, 200 U or more, 300 U or more, 400 U or more, 500 U or more, 600 U or more, 700 U or more, 800 U or more, 900 U or more, 1000 U or more, 1500 U or more, 2000 U or more, 2500 U or more, and even more preferably 2700 U or more.

[0040] The upper limit of the β-D-fructofuranosidase content is not particularly limited as long as it does not impair the effects of the present technology, and can be set to, for example, 3,000,000 U or less, 1,000,000 U or less, 600,000 U or less, 300,000 U or less, 100,000 U or less, 50,000 U or less, 10,000 U or less, 6,000 U or less, 4,500 U or less, or 3,300 U or less per gram of sugars contained in the sugar-containing food or beverage or per gram of sugars contained in the raw materials of the sugar-containing food or beverage.

[0041] In this technology, the activity of β-D-fructofuranosidase is defined as one unit (1 U) of the enzyme amount required to hydrolyze 1 μmol of sucrose into invert sugar in 1 minute at pH 4.5 and 55°C using 1.0% w / v sucrose as a substrate.

[0042] (4) Others The sugar-reducing agent according to the present technology may be composed of the glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase described above, as long as it contains the glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase described above. Alternatively, one or more other components may be freely selected and contained as long as the effects of the present technology are not impaired. Examples of other components that can be used include excipients, pH adjusters, colorants, flavoring agents, disintegrants, lubricants, stabilizers, and other components commonly used in formulations. Furthermore, components or enzymes with known or future functions can also be used in combination, as appropriate, depending on the purpose.

[0043] 2. Food and drink The food and drink according to the present technology is produced using the sugar-reducing agent described above.

[0044] (1) Raw materials The raw materials that can be used in the sugar-containing food and beverage used in the present technology or the food and beverage related to the present technology are not particularly limited in origin, type, etc. of the raw materials, and can be freely selected depending on the target food and beverage, as long as the effects of the present technology are not impaired. In the present technology, raw materials containing sucrose (sucrose-containing raw materials) are particularly preferred.

[0045] The origin of the raw material is not particularly limited and can be freely selected as long as it does not produce the function or effect of the present technology. Examples of raw material juices used for fruit juice drinks include orange (including mandarin orange), pineapple, grapefruit, apple, grape, peach, strawberry, banana, mango, melon, apricot, lemon, kiwi, blueberry, pear, grape, pomegranate, lime juice, and other fruit juices listed in the Fruit Drink Quality Labeling Standards.

[0046] Furthermore, examples of vegetable ingredients used in the vegetable juice beverage of the present invention include vegetable juices from carrots, spinach, onions, tomatoes, celery, paprika, pumpkin, corn, carrots, cucumbers, beets, parsley, lettuce, spinach, cauliflower, broccoli, kale, red bell peppers, ginger, green peppers, red cabbage, and the like.

[0047] Examples of the form of the raw material include liquid, paste, and slurry, and more preferably liquid. Furthermore, the raw material that can be used in the sugar-containing food and drink used in the present technology or the food and drink according to the present technology can be used as is or in the form of liquid, paste, or slurry, and can also be preferably used in the form of squeezed juice (fresh juice), concentrated juice, paste juice, powdered juice, etc.

[0048] (2) Amount of sugars contained in food and beverages The amount of sugars (monosaccharides and disaccharides, the same applies hereinafter) contained in the food and beverages according to the present technology is not particularly limited, but is preferably reduced relative to the sugar content in the food and beverages produced without using the sugar-reducing agent according to the present technology. Specifically, the total sugar content in the food and beverages according to the present technology is, for example, 70% or less, preferably 65% ​​or less, more preferably 60% or less, and even more preferably 57% or less, 50% or less, or 40% or less of the total sugar content in the food and beverages produced without using the sugar-reducing agent according to the present technology.

[0049] In particular, the amounts of glucose and sucrose contained in the food and beverage products according to the present technology are preferably reduced relative to the glucose and sucrose content in the food and beverage products produced without the sugar-reducing agent according to the present technology. Specifically, the glucose and sucrose content in the food and beverage products according to the present technology is, for example, 70% or less, preferably 65% ​​or less, more preferably 60% or less, and even more preferably 57% or less, relative to the glucose and sucrose content in the food and beverage products produced without the sugar-reducing agent according to the present technology. The glucose content in the food and beverage products according to the present technology is, for example, 95% or less, preferably 93% or less, more preferably 90% or less, 80% or less, 70% or less, or 60% or less, relative to the glucose content in the food and beverage products produced without the sugar-reducing agent according to the present technology. Furthermore, the sucrose content in the food and beverage products according to the present technology is, for example, 60% or less, preferably 50% or less, and more preferably 40% or less, relative to the sucrose content in the food and beverage products produced without the sugar-reducing agent according to the present technology.

[0050] (3) Types of Food and Drink The types of food and drink according to the present technology are not particularly limited as long as they do not impair the effects of the present technology, but beverages are preferred. Examples of beverages include vegetable drinks and fruit juice drinks. Examples of foods other than beverages include Japanese sweets (rice cakes, manju, daifuku, castella, bean paste, etc.), Western sweets (cream, cake, ice cream, etc.), soups, sauces, dressings, etc.

[0051] 3. Method for producing food and drink products, and method for reducing sugar content in sugar-containing food and drink products The method for producing food and drink products, and method for reducing sugar content in sugar-containing food and drink products according to the present technology, are methods comprising a step of allowing glucose oxidase to act on a sugar-containing food or drink product or a raw material for the sugar-containing food or drink product (hereinafter also referred to as the "glucose oxidase action step"), and a step of allowing α-glucosidase and / or β-D-fructofuranosidase to act on a sugar-containing food or drink product or a raw material for the sugar-containing food or drink product (hereinafter also referred to as the "α-glucosidase action step" and / or "β-D-fructofuranosidase action step").

[0052] The glucose oxidase reaction step and the α-glucosidase reaction step and / or β-D-fructofuranosidase reaction step can be performed simultaneously or in any order. That is, glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase may be reacted simultaneously or separately on a sugar-containing food or drink or a raw material for the sugar-containing food or drink. When acting separately, the order is not particularly limited, and an enzyme inactivation step can be performed between the actions of each enzyme, as necessary.

[0053] In addition, depending on the type of food or beverage, general food manufacturing processes such as a recovery process may be performed before, after, or simultaneously with each process, as long as the effects of the present technology are not impaired. Each process will be described in detail below.

[0054] (1) Glucose oxidase action step The glucose oxidase action step is a step of allowing glucose oxidase to act on a sugar-containing food or beverage or a raw material for the sugar-containing food or beverage. The amount of glucose oxidase added in the glucose oxidase action step can be freely set as long as it does not impair the effects of the present technology. The specific amount of glucose oxidase added in the present technology is the same as the content of glucose oxidase in the sugar-reducing agent described above, and therefore will not be described here.

[0055] Various conditions for the glucose oxidase reaction step can be freely set as long as they do not impair the effects of the present technology. For example, the pH, temperature, reaction time, etc. can be set depending on the physicochemical properties of the glucose oxidase used, such as the optimal pH, stable pH range, optimal temperature, and temperature stability. The pH can be set, for example, to 2.0 to 11.0, preferably 2.5 to 10.0, and more preferably 3.0 to 5.5. The temperature can be set, for example, to 10°C to 70°C, preferably 20°C to 50°C, and more preferably 25°C to 45°C. The reaction time can be set, for example, to 15 minutes to 120 hours, preferably 30 minutes to 72 hours, and more preferably 12 hours to 48 hours. Optimal reaction conditions can be determined through preliminary experiments.

[0056] (2) α-glucosidase action step The α-glucosidase action step is a step in which α-glucosidase is allowed to act on a sugar-containing food or beverage or a raw material for the sugar-containing food or beverage. The amount of α-glucosidase added in the α-glucosidase action step can be freely set as long as it does not impair the effects of the present technology. The specific amount of α-glucosidase added in the present technology is the same as the α-glucosidase content in the sugar-reducing agent described above, and therefore will not be described here.

[0057] Various conditions for the α-glucosidase reaction step can be freely set as long as they do not impair the effects of the present technology. For example, the pH, temperature, reaction time, etc. can be set depending on the physicochemical properties of the α-glucosidase used, such as the optimal pH, stable pH range, optimal temperature, and temperature stability. The pH can be set, for example, to 2.0 to 11.0, preferably 2.5 to 10.0, and more preferably 3.0 to 5.5. The temperature can be set, for example, to 10°C to 70°C, preferably 20°C to 50°C, and more preferably 25°C to 45°C. The reaction time can be set, for example, to 15 minutes to 120 hours, preferably 30 minutes to 72 hours, and more preferably 12 hours to 48 hours. The optimal reaction conditions can be determined through preliminary experiments.

[0058] (3) β-D-fructofuranosidase Reaction Step The β-D-fructofuranosidase reaction step is a step in which β-D-fructofuranosidase is allowed to act on a sugar-containing food or beverage or a raw material for the sugar-containing food or beverage. The amount of β-D-fructofuranosidase added in the β-D-fructofuranosidase reaction step can be freely set as long as it does not impair the effects of the present technology. The specific amount of β-D-fructofuranosidase added in the present technology is the same as the content of β-D-fructofuranosidase in the sugar-reducing agent described above, and therefore will not be described here.

[0059] Various conditions for the β-D-fructofuranosidase reaction step can be freely set as long as they do not impair the effects of the present technology. For example, the pH, temperature, reaction time, etc. can be set depending on the physicochemical properties of the β-D-fructofuranosidase used, such as the optimal pH, stable pH range, optimal temperature, and temperature stability. The pH can be set, for example, to 2.0 to 11.0, preferably 2.5 to 10.0, and more preferably 3.0 to 5.5. The temperature can be set, for example, to 10°C to 70°C, preferably 20°C to 65°C, and more preferably 25°C to 60°C. The reaction time can be set, for example, to 15 minutes to 120 hours, preferably 30 minutes to 72 hours, and more preferably 12 hours to 48 hours. Optimal reaction conditions can be determined through preliminary experiments.

[0060] (4) Recovery Step The recovery step is a step of recovering the food or beverage that has been subjected to the glucose oxidase reaction step and the α-glucosidase reaction step and / or the β-D-fructofuranosidase reaction step. Specific recovery methods can be selected from recovery methods commonly used in the production of foods or beverages, and can be used in combination of one or more of these, depending on the type of food or beverage to be produced.

[0061] Foods and beverages produced through a glucose oxidase action step, an α-glucosidase action step, and / or a β-D-fructofuranosidase action step are characterized by having a reduced amount of sugars (monosaccharides + disaccharides). That is, the present technology can perform a recovery step to recover foods and beverages having a reduced sugar (monosaccharides + disaccharides) content compared to foods and beverages produced without the glucose oxidase action step, the α-glucosidase action step, and / or the β-D-fructofuranosidase action step.

[0062] One embodiment of the method for producing a food or beverage product and the method for reducing sugars in a sugar-containing food or beverage product of the present technology includes the following steps (1), (2), (3), and / or (4), and (5). Note that an enzyme deactivation step may be added as needed after one or more steps selected from steps (2), (3), and (4). Steps (2), (3), and (4) can be performed simultaneously or in any order. (1) A step of preparing a sugar-containing food or drink or raw materials for the sugar-containing food or drink; (2) A step of allowing glucose oxidase to act on the prepared sugar-containing food or drink or raw materials for the sugar-containing food or drink; (3) A step of allowing α-glucosidase to act on the prepared sugar-containing food or drink or raw materials for the sugar-containing food or drink; (4) A step of allowing β-D-fructofuranosidase to act on the prepared sugar-containing food or drink or raw materials for the sugar-containing food or drink; (5) A step of recovering the food or drink produced through steps (1), (2), and (3) and / or (4).

[0063] The present invention will be described in more detail below with reference to examples. Note that the examples described below are representative examples of the present invention and should not be construed as narrowing the scope of the present invention.

[0064] 1. Raw materials and enzymes used The raw materials and enzymes used in the examples are shown in Table 1 below.

[0065]

[0066] 2. Enzyme Activity Measurement Method [Glucose Oxidase Activity Measurement Method] An appropriate amount of enzyme was weighed and dissolved or uniformly dispersed in chilled potassium phosphate-sodium hydroxide buffer (0.1 mol / L) at pH 7.0 to prepare a 50 mL sample solution. 2.50 g of D(+)-glucose was weighed and dissolved in water to prepare a 25 mL substrate solution. 0.5 mL of the substrate solution, 2 mL of potassium phosphate-sodium hydroxide buffer (0.1 mol / L, pH 7.0, containing phenol), 0.5 mL of peroxidase test solution (25 units / mL), and 0.1 mL of 4-aminoantipyrine solution (1→250) were placed in a quartz cell and heated at 37°C for 10 minutes. 0.1 mL of the sample solution was added to this solution, mixed well, and heated at 37°C to prepare the test solution. Separately, a comparison solution was prepared in the same manner as in the preparation of the test solution, except that a pH 7.0 potassium phosphate-sodium hydroxide buffer solution (0.1 mol / L) or water was used instead of the sample solution. The absorbance of the test solution and comparison solution at a wavelength of 500 nm was measured 2 minutes and 5 minutes after the addition of the sample solution. The amount of glucose oxidized was quantified from the molar extinction coefficient of the quinoneimine dye produced. The amount of enzyme required to oxidize 1 μmol of glucose per minute was defined as 1 unit (1 U).

[0067] [Method for measuring α-glucosidase activity] Measurements were performed using Method 2 of the α-glucosidase activity test in the 9th edition of the Japanese Standards for Food Additives. (1) 2.0 g of α-methyl-D-glucoside was weighed out and dissolved in water to make 100 mL, preparing an α-methyl-D-glucoside solution. 1 mL of the α-methyl-D-glucoside solution and 1 mL of 0.02 mol / L acetic acid / sodium acetate buffer (pH 5.0) were weighed into a test tube and left at 40°C for 10-15 minutes. After this, 0.5 mL of an aqueous solution containing transglucosidase (enzyme solution) was added and shook well. This was left at 40°C for exactly 60 minutes. After exactly 60 minutes, the tube was placed in a boiling water bath, heated for exactly 5 minutes, and cooled under running water. (2) 3 mL of the color-developing solution from the glucose measurement kit (Fujifilm Wako Pure Chemical Industries, Ltd.; hereafter the same) was weighed into a test tube, and 0.2 mL of the reaction solution obtained in (1) was added. The mixture was shaken well and then left at 40°C for exactly 5 minutes. The absorbance (E60) of this solution at a wavelength of 505 nm was measured using water as a control. Separately, as a blank, 1 mL of 0.02 mol / L acetic acid / sodium acetate buffer (pH 5.0) and 0.5 mL of an aqueous solution containing transglucosidase (enzyme solution) were weighed into a test tube, heated in a boiling water bath for exactly 5 minutes, and then cooled under running water. After cooling, 1 mL of α-methyl-D-glucoside solution was added, and the absorbance (E0) was measured in the same manner as above. (3) Glucose standard solution I or II from the glucose measurement kit was diluted with water to the specified concentration (20 mg / dL, 40 mg / dL). Three mL of the color-developing solution from the glucose assay kit was weighed into a test tube, and 0.2 mL of the glucose solution was added. The mixture was shaken well and then allowed to stand at 40°C for exactly 5 minutes. The absorbance (ES) of this solution was measured at a wavelength of 505 nm using water as a control. Separately, the absorbance (EB) was measured in the same manner as above using 0.2 mL of water instead of the glucose solution as a blank. A glucose calibration curve was created from the obtained absorbance, and the amount of glucose (μg) (G) at an absorbance difference of 1.000 was calculated. (4) The amount of enzyme required to produce 1 μg of glucose in 60 minutes was defined as 1 unit (1 U), and calculated using the following formula:α-Glucosidase activity (U / g, U / mL) = (E60 - E0) × G × 2.5 / 0.1 × n / 0.5 E60: absorbance of reaction solution E0: absorbance of blank solution G: amount of glucose (μg) when the absorbance difference is 1.000 (calculated from the glucose calibration curve) 2.5: volume of reaction system solution (mL) 0.1: volume of reaction solution collected (mL) 0.5: volume of enzyme solution collected (mL) n: dilution factor per 1 g or 1 mL of sample.

[0068] [Method for measuring β-D-fructofuranosidase activity] (1) Preparation of reagents: A 100 mM sodium acetate solution (sodium acetate trihydrate (Sigma-Aldrich #S8625) dissolved in purified water) adjusted to pH 4.5 with 1 M HCl at 55°C was used as a buffer solution. Sucrose (Sigma-Aldrich #S7903) was dissolved in 100 mM sodium acetate buffer (pH 4.5) to prepare a 1.0% w / v sucrose substrate solution. Immediately before use, a solution containing 0.3-0.4 U / mL of β-D-fructofuranosidase was dissolved in cold purified water to prepare the enzyme solution. 1 N NaOH (Supelco #S2567) was diluted 1:2 with purified water to prepare a 0.5 M NaOH dilution. p-Hydroxybenzhydrazide (PAHBAH) (Sigma-Aldrich #H9882) was dissolved in 0.5 M NaOH to prepare a 0.5% w / v PAHBAH reagent. Glucose (Sigma-Aldrich #G8270) was dissolved in purified water to prepare a 5.56 mM glucose standard solution. (2) Measurement Method (2-1) Preparation of Glucose Standard Solution: 1.0 mL of purified water was used as a glucose standard solution blank. Purified water (0.95 mL, 0.90 mL, 0.80 mL, 0.70 mL, 0.60 mL, 0.50 mL) was mixed with the glucose standard solution prepared above (0.05 mL, 0.10 mL, 0.20 mL, 0.30 mL, 0.40 mL, 0.50 mL), and the glucose standard solutions were equilibrated at 55°C. (2-2) Preparation of Blank Test Solution: 0.10 mL of purified water and 0.90 mL of the sucrose substrate solution prepared above were mixed to prepare a blank test solution, which was then equilibrated at 55°C. (2-3) Preparation of Test Solution: 0.90 mL of the sucrose substrate solution prepared above and 0.10 mL of the enzyme solution prepared above were mixed to prepare a test solution, which was then precisely incubated at 55°C for 20 minutes. (2-4) Measurement of Absorbance: 0.10 mL of each of the glucose standard solutions, blank test solution, and test solution prepared above was measured and added to a tube containing 2.90 mL of the 0.5% w / v PAHBAH reagent prepared above. The mixture was quickly mixed, placed in a boiling water bath, heated for 5 minutes, and allowed to cool to room temperature. After cooling, the solution was mixed and transferred to an appropriate cuvette.All reaction mixtures were measured for absorbance at 410 nm using a suitable spectrophotometer. The amount of enzyme required to hydrolyze 1 μmol of sucrose to invert sugar per minute was defined as 1 unit (1 U).

[0069] 3. Experimental Example <Experimental Example 1> (1) Experimental Method 50 mL of commercially available fruit juice (V8 Pomegranate Blueberry Juice) was pre-incubated at 35°C for 10 minutes with stirring. Next, each enzyme shown in Table 3 below was added, and the mixture was treated with enzymes at 35°C for 24 hours with stirring. After the enzyme reaction was completed, the enzyme activity was inactivated by treating at 90°C for 20 minutes, and the mixture was cooled to room temperature to obtain Fruit Juice 1.

[0070] (2) Sugar Composition Analysis The resulting fruit juice 1 was diluted 5-fold with deionized water and centrifuged at 13,300 G for 10 minutes. The supernatant was diluted 2-fold with ethanol (final ethanol concentration: 50% or more), passed through a membrane filter (0.22 μm), and analyzed by high-performance liquid chromatography (Refractive Index Detector (RI) "RID-20A" Shimadzu Corporation). Column details and conditions are shown in Table 2 below. The sugar composition was analyzed and quantified using standard sugars (Sigma). The ratio of glucose to sucrose relative to the total sugar content (sugar composition) was calculated from the obtained sugar composition.

[0071]

[0072] (3) Results The results are shown in Table 3 below.

[0073]

[0074] (4) Discussion As shown in Comparative Examples 1 to 3 and 6, when glucose oxidase, catalase, β-D-fructofuranosidase, or α-glucosidase was used alone, the sugar reduction rate was less than 30%. Furthermore, as shown in Comparative Examples 4 and 5, when glucose oxidase and catalase were used in combination, the sugar reduction rate was less than 35%. On the other hand, Example 1, in which glucose oxidase and α-glucosidase were used in combination, and Example 2, in which glucose oxidase and β-D-fructofuranosidase were used in combination, showed a high sugar reduction rate of more than 40%.

[0075] Experimental Example 2 (1) Experimental Method 50 mL of commercially available fruit juice (Ceres Peach Juice) was pre-incubated at 35°C for 10 minutes under stirring. Each enzyme shown in Table 4 below was added, and the mixture was treated at 35°C for 24 hours. After the enzyme reaction was completed, the enzyme activity was inactivated by treating at 90°C for 20 minutes, and the mixture was cooled to room temperature to obtain Fruit Juice 2.

[0076] (2) Analysis of Sugar Composition The sugar composition of the obtained fruit juice 2 was analyzed in the same manner as in Experimental Example 1.

[0077] (3) Results The results are shown in Table 4 below.

[0078]

[0079] (4) Discussion As shown in Table 4, Example 3, in which glucose oxidase and α-glucosidase were used in combination, showed a higher reduction rate than Comparative Examples 7 to 10, in which glucose oxidase or α-glucosidase was used alone, and Comparative Examples 11 to 13, in which glucose oxidase and catalase were used in combination.

[0080] Experimental Example 3 (1) Experimental Method 50 mL of commercially available fruit juice (TRADERJOE'S Orange Peach Mango Juice) was pre-incubated at 35°C for 10 minutes with stirring. Each enzyme shown in Table 5 below was added, and the mixture was treated at 35°C for 24 hours. After the enzyme reaction was completed, the enzyme activity was inactivated by treating at 90°C for 20 minutes, and the mixture was cooled to room temperature to obtain Fruit Juice 3.

[0081] (2) Analysis of Sugar Composition The obtained fruit juice 3 was subjected to analysis of sugar composition in the same manner as in Experimental Example 1.

[0082] (3) Results The results are shown in Table 5 below.

[0083]

[0084] (4) Discussion As shown in Table 5, Examples 4 to 6, in which glucose oxidase and α-glucosidase were used in combination, showed a higher reduction rate than Comparative Examples 14 and 15, in which glucose oxidase or α-glucosidase was used alone.

[0085] Experimental Example 4 (1) Experimental Method 50 mL of commercially available fruit juice (V8 Vegetable Juice) was pre-incubated at 35°C for 10 minutes under stirring. Each enzyme shown in Table 6 below was added, and the mixture was treated at 35°C for 24 hours. After the enzyme reaction was completed, the enzyme activity was inactivated by treating at 90°C for 20 minutes, and the mixture was cooled to room temperature to obtain Fruit Juice 4.

[0086] (2) Analysis of Sugar Composition The sugar composition of the obtained fruit juice 4 was analyzed in the same manner as in Experimental Example 1.

[0087] (3) Results The results are shown in Table 6 below.

[0088]

[0089] (4) Discussion As shown in Table 6, it was found that the reduction rate of sugars improved as the amount of glucose oxidase used in combination with α-glucosidase increased.

Claims

1. A sugar-reducing agent for sugar-containing foods and beverages, comprising glucose oxidase and α-glucosidase and / or β-D-fructofuranosidase.

2. A sugar-reducing agent according to claim 1, which reduces glucose and sucrose in sugar-containing foods and beverages.

3. The sugar-reducing agent according to claim 1, wherein the sugar-containing food or beverage is a sucrose-containing food or beverage.

4. The sugar-reducing agent according to claim 1, wherein the sugar-containing food or beverage is a beverage made from fruits and / or vegetables.

5. A food or drink containing the sugar-reducing agent according to any one of claims 1 to 4.

6. A method for producing a food or drink, comprising: a step of allowing glucose oxidase to act on a sugar-containing food or drink or a raw material for the sugar-containing food or drink; and a step of allowing α-glucosidase and / or β-D-fructofuranosidase to act on the sugar-containing food or drink or a raw material for the sugar-containing food or drink.

7. A method for reducing sugars in sugar-containing foods and beverages, comprising the steps of: allowing glucose oxidase to act on sugar-containing foods and beverages or raw materials for the sugar-containing foods and beverages; and allowing α-glucosidase and / or β-D-fructofuranosidase to act on sugar-containing foods and beverages or raw materials for the sugar-containing foods and beverages.

Images