JPS6291161A - Stevia sweetener and production thereof - Google Patents

Abstract

PURPOSE:To produce a stevia sweetener having improved flavor free from bitterness and aftertaste, by decomposing selectively sweetening components except rebaudioside A in stevia sweetening components with a fungus or an enzyme. CONSTITUTION:An aqueous solution containing rebaudioside A and stevia sweetening components except it or an extracted solution of leaves of Stevia rebaudiana BERTONI is treated with a fungus such as Aspergillus nigar ATCC-6275, etc., capable of decomposing selectively the stevia sweetening components except the rebaudioside A or with an enzyme obtained from the fungus, to give a stevia sweetener comprising rebaudiside A, stevioside and decomposition products of the stevia sweetening components except the rebaudioside A, preferably containing >=5.5pts.wt. rebaudioside A based on 1pt.wt. stevioside.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention provides rebaudioside A in a stevia sweetening ingredient.
(1) Rebaudioside A, (2) Stevioside, (3) Rebaudioside A obtained by selectively decomposing sweet ingredients other than stevioside, such as stevioside with strong bitterness or residual stock, using microorganisms or enzymes. The present invention relates to a novel stevia sweetener with excellent taste, which is made of a decomposed product of a stevia sweet component other than the above, and a method for producing the same.

[Conventional technology]

In recent years, artificial sweeteners such as sodium saccharinate, sodium saccharinate, and dultin have been prohibited or restricted from being used in general foods due to safety concerns. As the negative effects of sweeteners have become a problem, there is a strong desire to develop alternative natural sweeteners. Under these circumstances, the demand for stevioside and rebaudioside A is rapidly increasing because, unlike sugar, they are low-calorie sweeteners, and their sweetness ratio is approximately 300 and 360 times higher than that of sugar. ing. Stevioside and rebaudioside A are derived from Stevia rebaudiana bertoni (Stevia rebaudiana), which belongs to the Asteraceae family.
BERTONI) is a sweetening ingredient extracted from A. baudiana BERTONI), and is a β-
It is a glucosyl glycoside. Mixtures of stevioside and rebaudioside A are currently used to flavor foods and drinks. Among these, rebaudioside A has superior sweetness compared to stevioside, but its content is low, and the sweetness of stevioside appears later than that of sugar, and it remains for a long time as a residue. In addition to the disadvantages, it also has disadvantages such as bitterness and unpleasant taste, and the stevia sweetener whose main ingredient is stevioside depends on the amount used.
It is said that there are limits to its uses and that some improvement in sweetness is required.

[Means for solving problems]

As a result of intensive research in order to solve the above-mentioned drawbacks of stevia sweeteners, the present inventors found that among the stevia sweetening components other than rebaudioside A, stevioside, which has a particularly large amount of bitterness, sarcasm, and residual stock, was found. The present inventors have discovered that a sweetener mainly composed of rebaudioside A with excellent taste can be obtained by selectively decomposing it using biological techniques, and have completed the present invention. That is, the present invention provides (1) rebaudioside A, (2) stevioside, (3) a stevia sweetener consisting of a decomposed product of a stevia sweet component other than rebaudioside A, and furthermore, an aqueous solution containing a stevia sweet component. The present invention provides a method for producing a stevia sweetener, characterized in that a microorganism or an enzyme having the ability to selectively decompose stevia sweet components other than rebaudioside A is applied to the stevia sweetener.

In the prior art, there is nothing known about sweeteners such as the sweetener of the present invention, which are obtained by decomposing stevioside other than rebaudioside A, which is a sweet component extracted from stevia. Although the method of decomposing and quantifying stevioside with an enzyme (hesperidinase) is known,
(pp. 507-1511) This hesperidinase differs from the present invention in that it decomposes all stevia sweet components such as stevioside and rebaudioside A.

〔composition〕

The aqueous solution or raw material containing the stevia sweetening component used in the production of the sweetener of the present invention is limited to an aqueous solution of a highly purified or roughly purified stevia sweetener product (mainly consisting of rebaudioside 8 and stevioside). Rather, it may be a solution during extraction from Stevia leaves, a solution after extraction, or any aqueous solution containing Rebaudioside A and other Stevia sweet components.

Preferably, the raw material has a high content of rebaudioside A, and if it is a solution during extraction, it is better in terms of production efficiency.

Therefore, the aqueous solution containing the stevia sweetening component of the present invention is an aqueous solution or extract containing rebaudioside A, rubaudioside C (dulcoside B), D, E, glucoside A1, steviol bioside, and stevioside. . Further, if necessary, the aqueous solution may contain an organic solvent such as ethanol.

The microorganism or enzyme used in the present invention may be any microorganism or enzyme that has the ability to selectively decompose stevia sweet components other than rebaudioside A, but preferably contains a stevia sweet component. Any material can be used as long as it selectively decomposes only stevioside in the aqueous solution and does not use rebaudioside A as a reaction substrate. Furthermore, these microorganisms or enzymes have the ability to cleave stevia sweet ingredients other than rebaudioside A, such as glucose bound in stevioside, one glucose or two or three glucose molecules together, and in particular stevioside. It is preferable to have the ability (glucosidase activity) to decompose the terminal glucose of the sophorose moiety or to release all or part of the glucose bound to the aglycone to produce steviol or steviol-based decomposition products. The stevia sweetness components other than rebaudioside A referred to herein include stevioside, rebaudioside C5DXE, dulcoside A, and steviol bioside. Among these sweet components, it is preferable that rebaudioside D is not decomposed from the viewpoint of taste quality.

In addition, examples of microorganisms or enzymes used in the present invention include, for example, molds, actinomycetes, bacteria, yeasts, and basidiomycetes.
Trichoderma sp., Rhizopus sp., Chaetomium sp., Myriocochieme sp., Thermomyces sp., Aspergillus sp.
Various molds belonging to the genus Penicillium, Mylocesium, Mucor, Sborotrichum, Sphlerotinia, Rhizoctonia, Sclerotium, Bellichielaria, etc., and various molds belonging to the genus Streptomyces, Micropolisvora, Oscobia, etc. Actinomycetes, and Bacillus, Brevibacterium, Arthrobacterium,
Examples include various bacteria belonging to the genus Flavobacterium and the like, yeasts of the genus Schizosatucharomyces, and various basidiomycetes belonging to the genus Trametes, Irpex, Coprinus and the like. Particularly excellent are those belonging to the genus Flavobacterium, Bacillus, Streptomyces, Aspergillus, and the like. The wide distribution of microorganisms that can be used in the present invention is as seen in the results shown in Attached Table 1.

In particular, as a strain of the genus Aspergillus, for example, Aspergillus nigar
) AT CC-6275, Aspergillus niger IFO-6341, etc. are preferably mentioned.

The enzymes used in the present invention and the enzymes obtained from microorganisms do not necessarily need to be purified, and even the culture solution or culture filtrate of microorganisms is sufficient to decompose stevia sweet components other than rebaudioside A, especially stevioside. Any sweetener that can obtain the desired sweetener is fine.

In the reaction of the present invention, the microorganism or an enzyme obtained from the microorganism is allowed to act on an aqueous solution containing the stevia sweet component, and conditions may be selected to decompose the stevia sweet component other than rebaudioside A. However, the reaction temperature may be is 15℃~60℃
, preferably 25°C to 50°C; the pH is preferably 2.0 to 9.0, preferably 4.0 to 7.0, and the reaction time is about 30 minutes to 3 days, preferably 1 to 50°C. 1
2 hours is suitable.

In addition, the concentration of the stevia sweet ingredient used in the reaction is approximately 0.1~
Approximately 10% by weight is suitable.

The stevia sweetener containing rebaudicide A as a main component obtained by this reaction is a sweetener containing more than 5.5 parts by weight of rebaudioside A per 1 part by weight of stevioside. Therefore, when the solid content ratio of each component in the sweetener of the present invention is expressed in parts by weight, for 1 part by weight of stevioside, 5.5 to 70 parts by weight of rebaudioside A (excluding 5.5), The ratio of decomposed sweet components other than Dioside A is preferably 0.01 to 30 parts by weight.

In addition to the above-mentioned components, the sweetener of the present invention also includes rebaudioside C, DXE, etc. derived from stevia sweetening components.
May contain other sweet ingredients such as zurcoside A,
The amount thereof is 0.001 to 10 parts by weight in terms of solid content.

Decomposed products of stevia sweetness components other than rebaudioside A, which is the third component of the present invention, include, for example, stevioside,
Rebaudioside C1 D, same E1 Zurcoside A,
Same B, glucose 1~ from steviol bioside etc.
Steviol glycosides with 5 or rhamnose removed and 1 to 4 glucose or rhamnose added, such as pubusoside, steviol monoside, steviol bioside, and all of these sugars removed It is called steviol. Moreover, many of these decomposition products are derived from stevioside.

The composition of these decomposition products varies depending on the reaction conditions and the microorganisms or enzymes used, but if the decomposition reaction is carried out appropriately and for a sufficient period of time, steviol may be produced.

Furthermore, these decomposition products themselves are often substances that have low sweetness or are tasteless and odorless, and not only do they not cause unpleasant taste, residual taste, or astringency in the resulting sweetener, There is a feeling that the taste quality of rebaudioside A is improved by the presence of these substances. However, if it is desired to increase the purity of the main components, these decomposed products can be removed after the reaction by a column purification method using an ion exchange resin, an adsorption resin, etc. or a crystallization method, which will be described later.

The sweetener of the present invention can be used as a sweetener as it is. If necessary, after inactivating the microbial cells or enzymes by heating, purification may be performed using a polymerized adsorption resin of styrene and divinylbenzene such as Diaion HP-20 (trade name, manufactured by Mitsubishi Chemical Corporation), Amberlite XAD-2. (trade name, Organo Co., Ltd. product), etc., or by desalting using an ion exchange resin (for example, H-type strong acid ion exchange resin and OH-type weak basic ion exchange resin) and concentrating it to make a syrupy sweetener. Alternatively, it can be dried and powdered to form a powdered sweetener.

The desalted reaction solution was further purified to obtain rebaudioside A.
It can also be purified and collected to be used as a sweetener. At this time, concentration, drying, and powdering are performed using known methods such as vacuum concentration,
Various methods such as membrane concentration, vacuum drying, spray drying etc. are at will. The sweetness of the sweetener of the present invention containing rebaudioside A as a main component obtained in this way is weaker than the sweetness of the raw material before reaction, but the quality of the sweetness is derived from stevioside. It does not have unpleasant tastes such as bitterness or astringency, and has a mellow sweetness that is closer to pure rebaudioside A, similar to sugar, and the remaining leaves are easy to cut.

The sweetener of the present invention is an odorless, white powder with no bitterness, unpleasant taste, or acrid taste, and is soluble in water. can be done. Furthermore, the sweetener of the present invention can be used in combination with well-known synthetic sweetening substances such as saccharin and its salts, sodium cyclamate, dihydrochalcone, and aspartame to effectively utilize their taste characteristics. One or two synthetic sweet substances
By adding the sweetener of the present invention to more than the seeds, it is possible to improve unpleasant tastes such as bitterness and sarcastic taste peculiar to synthetic sweet substances.

In addition, the sweetener of the present invention can be added to well-known sugar sweeteners such as sugar, fructose, glucose, lactose, starch syrup, dextrin, and starch, which are used as excipients, diluents, and adsorbents. It is possible to significantly reduce the usage amount by 1. Furthermore, if the sweetener of the present invention is added to low-calorie sweet substances such as sorbitol, maltitol, mannitol, and xylitol, which have a lower sweetness than sugar, the sweetness can be enhanced without impairing the advantages of the sweet substances, and the sweetness can be increased to produce high-quality products. A low calorie sweetener is obtained.

It goes without saying that the sweetener of the present invention can be used as a sweetening source for general foods, diet foods, medicines, quasi-drugs, cigarettes, feeds, and the like.

For example, soy sauce, powdered soy sauce, miso, powdered miso, mash, mayonnaise, dressing, vinegar, sambai vinegar, powdered sushi vinegar, Chinese soup stock, tempura soup, mentsuyu, sauce, ketchup, yakiniku sauce, curry roux, stew stock, soup stock , dashi stock, compound seasonings, mirin, new mirin, table syrup and other seasonings, rice crackers, arare, okoshi, rice cakes, manju, uiro, red bean paste, yokan,
Various Japanese sweets such as mizu-tekan, jelly, castella, candy, ° bread, biscuits, kura7ka, kutsuki, pie, pudding, butter cream, custard cream, cream puffs, watsuful, sponge cake, donuts, chocolate, chewing gum, caramel, Various Western sweets such as candies, frozen confections such as ice cream, sherbet, and popsicles, fruits preserved in syrup, syrups such as honeydew, pastes such as flower paste, peanut paste, and fuller paste, jams, marmalade, preserved syrups, and sweets. Processed fruits and vegetables such as Fukujinzuke, Senmaizuke, Rakkyōsai, etc., meat products such as ham and sausages, meat products such as meat ham, fish sausage, kamaboko, chikuwa, tempura, etc., sea urchin and squid. Various delicacies such as salted fish, sakisurume, dried puffer fish, seaweed, wild vegetables, dried squid, small fish, fish made from shellfish, vegetable foods such as smoked beans, potato salad, kelp rolls, fish meat, Bottled meat, fruits, and vegetables, canned goods, alcoholic beverages such as synthetic liquor, fruit liquor, and Western liquor, soft drinks such as coffee, cocoa, juice, carbonated drinks, lactic acid drinks, and lactic acid bacteria drinks, pudding mixes, pancake mixes, It can be used for flavoring various foods and beverages such as instant juice, instant coffee, instant shiruko, and other instant foods and beverages.

In addition, as pharmaceuticals and quasi-drugs, it can be freely used as a sweetener in line brushes, lipsticks, lip balms, oral medicines, troches, cod liver oil drops, mouth fresheners, mouthwashes, gargles, etc.

[Effects] Since the sweetener of the present invention contains a large amount of rebaudioside A, it has excellent sweetness quality comparable to that of pure rebaudioside A. Compared to conventional sweeteners containing less than 550 parts of rebaudioside A per 100 parts of stevioside, its sweetness is more rebaudioside in terms of astringency, persistence, bitterness, and sarcastic taste. It is an excellent sweetener with a taste quality similar to Side A.

The method of the present invention and the sweetener obtained thereby will be specifically explained below using Examples.
is based on the heavy N standard.

Example 1 The presence or absence of stevioside decomposition activity and its selectivity were investigated for the microorganisms shown in Table-1. The results are shown in Table-1. The microorganisms shown in Table 1 were grown in a medium containing the sweetening component of Stevia, or the culture solution of the microorganism was added to an aqueous solution containing the sweetening component of Stevia and reacted, and the results were determined by liquid chromatography.

Table 1 ◎: Strongly present O: Present Δ: Slightly present ×: Not present Stv: Stevioside Example 2 Potassium phosphate 0.4%, ammonium sulfate 0.5%, magnesium sulfate 0.06%, yeast extract 0.1% ,
Zinc sulfate 0.001%, ferrous sulfate o, o o s%
, (hereinafter referred to as Medius A).
H6,0) with 1% glucose added to 100ml of medium for 500mj! Dispense into Yosaka Roof flask, and then add purified stevia sweetener (composition: 20% stevioside, 41.6% rebaudioside A, 8.5% rebaudioside C).
%), add 1g, dissolve, and autoclave to 120%
℃×15min bacteria. After cooling, one platinum loop of Aspergillus niger ATCC 6275 was inoculated and incubated at 34°C for 44 hours.
A decomposition reaction was carried out by shaking culture for hours.

After the reaction, the obtained supernatant synovial fluid was filtered with a Buchner filter, and the resulting supernatant fluid was coated with adsorption resin Diaion HP-20 (manufactured by Mitsubishi Kasei) with S, V,
= 2 to adsorb steviosides. after that,
After desorption with 95% ethanol and distilling off the ethanol in the desorption solution under reduced pressure, strongly acidic ion exchange resin Amberlite I was used.
R-120B (H type, product name, Rohm and Haas Company)
, Weakly basic ion exchange resin Amberly 1-IRA-9
3 (OH type, product name, Rohm and Haas Company) with S and V
, -2 to desalt. This was then concentrated under reduced pressure at 70°C or lower and vacuum dried. As a result, HPLC analysis revealed that the sweet components were 40.0% rebaudioside A, 7.2% rebaudioside C, and 0.5% stevioside.
% or less, and a stevia sweetener having a composition of 20.2% decomposition products of sweet components other than rebaudioside A was obtained.

Example 3 Completely the same basal medium composition as in Example 2 (MedlumA
) and 1% glucose was added to a 101 volume jar fermenter and sterilized at 110°C for 30 minutes. After cooling, 0.2j of Aspergillus niger IFO6341 seed culture cultured in the same medium composition! was added and cultured at 32°C for 30 hours. When entering the stationary phase,
Separately sterilized purified stevia sweetener (composition: stevioside 2). 2%, rebaudioside A 55.1%, rebaudioside C 6.3%) solution (40g10.3g
was added, aeration rate was 4 j/win, and stirring was performed at 400 rpm.
The 0 reaction solution subjected to the decomposition reaction at 34° C. for 14 hours was purified in exactly the same manner as in Example 2. HPLC analysis of the obtained stevia sweetener revealed that it contained 53.3% rebaudioside A and 5.8% rebaudioside C.
%, stevioside 1.2%, and decomposition products of sweet components other than rebaudioside A 22.2%.

Example 4 A medium 3.51 containing the same basal medium composition (MediaA) as in Example 2 and 1% RAF-su was added to an IOA jar fermenter and sterilized at 110°C for 30 minutes.

After cooling, 0.2) of Aspergillus niger ATCC6275 seed culture cultured with the same medium composition was added, and the mixture was incubated at 32°C.
The cells were cultured for 40 hours.

Thereafter, 0.3 A of Stevia leaf extract (&l composition: adult stevioside 9.9%, Baudioside A 3.8%, Rebaudioside C 1.6%) was added in exactly the same manner as in Example 3. After 300 hours of decomposition reaction under the same conditions as in Example 3, the product was purified in the same manner as in Example 2 to obtain a dry powder.

The sweetener obtained was rebaudioside A 44.4%,
Rebaudioside CI 4.3%, Stevioside 5
．． It had a composition of 2% steviol and 11.8% steviol.

Example 5 (Sweetness level test of sweetener) Sample size 1 of the stevia sweetener obtained in Example 2 (stevioside (Stν) 20%, rebaudioside A
(RabA) 41.6% aqueous solution (purified stevia sweetener), Reb2 (RebA 40%, 5tv0.5%)
The following 0.02% and 0.05% aqueous solutions of the present invention sweetener) were prepared, and three types of standard solutions were prepared using 1 to 7% aqueous sugar solutions at 0.5% concentration levels. A sweetness test was conducted. The test was conducted by 20 panelists at a room temperature of 20°C using a two-point comparison method between a sample solution and a standard solution, and the results are shown in Table 2.

Table 2 A 0.05% aqueous solution corresponds to a sand Iff level of 4.5% (sweetness level 225 times), and a 0.05% aqueous solution corresponds to a sugar concentration of 9.0% (sweetness level 1).
80 times). Similarly, the sweetness of Sample-2 corresponds to 4.0% and 8.5% of the sugar concentration, respectively, so the sweetness of the sweetener of the present invention is slightly lower than that commensurate with the raw material used. It is judged that.

Example 6 (Taste quality test of sweetener) In the same manner as in Example 5, the control product of Sample 1 and the sweetener of the present invention of Sample 2 were used to compare the difference in sweetness quality.

Calculated from the sweetness determined in the sweetness test, each sample was adjusted to an aqueous solution with a sweetness corresponding to 3%, 6%, and 10% sugar aqueous solutions. And 1 sample fish for each sweetness level, sample! l
The taste quality of the h2 sample solutions was compared.

The test was conducted at room temperature of 20° C. by 20 panel members.

The results are shown in Table-3.

From the results in Table 3, it is clear that the sweetness quality of the sweetener of the present invention in Sample No. 2 is superior to the control product in Sample No. 1 at all sweetness levels.

Application example 1 A sample of powdered juice was prepared using the following formulation.

Granule 1! 950 g 1li 230 g sodium citrate 20 g malic acid
14g fragrance
10g β-carotene (1,5
%) 3g sweetener of the present invention
2.0g As a comparative example, Sample No. 1 of Example 5 was added to 2.0g instead of the sweetener of the present invention in the above formulation. A prototype was made with ogg added.

As a result of a taste test conducted by 20 panel members, the product of the present invention had no bitterness or unpleasant taste, while the comparative product had a bitter aftertaste.

Application Example 2 A trial production of 20% orange juice was made using the following formulation.

Orange ioo% natural fruit juice 430g top white
Sugar Log
High fructose isomerization vM2) 0 g Citric acid 3.5 g Malic acid 2 g Sodium citrate 0.5 g Flavoring
2g β-carotene (1,
5% > 0.5g Invention sweetener
The total amount of 0.5g was made up to 22 with water.As a comparative example, a sample was prepared in which 0.5g of sample 11hl of Example 5 was added instead of the sweetener of the present invention in the above formulation.

As a result of a taste test conducted by 20 panel members, the product of the present invention had a mild sweet taste and flavor, while the comparative product had an unpleasant aftertaste.

Application example 3 A trial cider was produced using the following formulation.

Granulated sugar 30g High fructose sugar 170g Citric acid
3g Sodium citrate
0.2 g cider essence
0.2g Invention sweetener
The total amount of 0.22 g was adjusted to 2) with carbonated water.As a comparative example, a sample was prepared in which 0.2) g of Sample Level 1 of Example 5 was added instead of the sweetener of the present invention in the above formulation.

As a result of a taste test conducted by 20 panel members, the product of the present invention had a light sweetness and had a good residual taste, while the comparative product had a deceptive bitterness and astringency.

Claims (1)

[Claims] 1. A stevia sweetener comprising (1) rebaudioside A, (2) stevioside, and (3) a decomposition product of a stevia sweet component other than rebaudioside A. 2. A patent claim characterized in that the ratio of (1) rebaudioside A and (2) stevioside is more than 5.5 parts by weight per 1 part by weight of (2). Stevia sweetener according to scope 1. 3. A method for producing a stevia sweetener, which comprises reacting an aqueous solution containing stevia sweeteners with microorganisms or enzymes capable of selectively decomposing stevia sweeteners other than rebaudioside A.