GB2105338A

GB2105338A - A low-calorie sweetener and a process for preparation of low- calorie food and drink containing the same

Info

Publication number: GB2105338A
Application number: GB08224919A
Authority: GB
Inventors: Ueto Takeda
Original assignee: Meiji Seika Kaisha Ltd
Current assignee: Meiji Seika Kaisha Ltd
Priority date: 1981-09-01
Filing date: 1982-09-01
Publication date: 1983-03-23
Also published as: DE3232531C2; JPS5840065A; DE3232531A1

Abstract

The low-calorie sweetener contains oligosaccharides consisting of from 1 to 4 molecules of fructose bound to sucrose. The oligosaccharides are produced by causing fructosyl transferase to act on sucrose. The sweetener may also contain glucose, sucrose and fructose and is used by incorporating into a food or drink as a low-calorie sweetener therefor.

Description

SPECIFICATION A low-calorie sweetener and a process for preparation of low-calorie food and drink containing the same The present invention relates to a low-calorie sweetener containing oligosaccharides which are obtained by causing fructosyl transferase to act on sucrose and which are composed of from 1 to 4 molecules of fructose bound to sucrose, and further relates to a process for preparation of low-calorie food and drink which comprises using said sweetener.

Sucrose has heretofore been widely used in confectionery and food by virtue of its excellent characteristics such as good sweetness, body taste, and crystallinity. Sucrose, however, constitutes a substrate for dextransucrase produced by intraoral microorganisms, and, as a result, consecutive intake of sucrose leads to formation of large amounts of insoluble dextran in the mouth; thus formation of dental plaque is accelerated. Therefore, sucrose is said to possess cariogenicity. Recently, there is a trend of reducing the calorie intake for prevention of obesity, and a low-calorie sweetener, instead of a high-calorie one such as sucrose, has been demanded.

The present inventors have intensively investigated to develop low-calorie sucrose-related sugars which still have excellent properties of sucrose. As a result, it has now been found that oligosaccharides obtained by causing fructosyl transferase to act on sucrose, namely, an oligosaccharide in which one molecule of frustose is bound to sucrose (hereinafter referred to as GF2), an oligosaccharide in which two molecules of fructose are bound to sucrose (hereinafter referred to as GF3), and an oligosaccharide in which three molecules of fructose are bound to sucrose (hereinafter referred to as GF4) are low-calorie sugars.Oligosaccharides herein used, such as GF2, GF3, GF4 and GF5, can be isolated and purified, from the transferred sugar mixture obtained by causing fructosyl transferase to act on sucrose, by means of, for example, carbon chromatography, ion-exchange chromatography, or the like. From the practical point of view, however, the oligosaccharide mixture is preferably used per se. These oligosaccharide compositions can further be used with addition of sugar alcohols such as sorbitol, mannitol, and multitol, and artificial sweeteners such as dehydrochalcone and stevioside.The sugar compositions obtained by causing fructosyl transferase to act on sucrose can also be used in the form in which monosaccharides therein, that is, glucose and fructose alone, have been selectively converted into sorbitol and mannitol by catalytically reducing an aqueous slution of said sugar compositions (adjusted to pH 7 to 9) at a temperature of 50 to 1 30 C and a hydrogen pressure of 50 to 1 20 kg/cm3 in the presence of a nickel catalyst in an amount of 3 to 10% relative to the solid material.

Further, the composition of the present invention possesses excellent properties as a sweetener, such as good sweetness, proper body taste, good moisture retention, etc., in addition to being low-calorie.

As described above, the composition which contains as its component oligosaccharides consisting of from 1 to 4 molecules of fructose bound to sucrose (hereinafter referred to as lowcalorie sweetener) is a low-calorie sweetener composition and is obtained by causing fructosyl transferase to act on sucrose.

Fructosyl transferase herein used has such activities as to act mainly on sucrose to sever the P-1-2- between fructose and glucose, then transfer the resulting fructose to sucrose to yield GF2, and further transfer fuctose to GF2 to yield GF3. It is different from inulosucrose (2.4.1.9] and levansucrase [2.4.1. 10] described in Enzyme Nomenclature (Academic Press, 1978) in that the reaction products are oligosaccharides wherein fructose is bound to sucrose, as GF2, GF3, etc.

As a source for the enzyme, there are microorganisms such as fungi, for example, the genus Aspergillus (Aspergillus niger ACE-2- 1, FERM-P5886, etc.), the genus Penicillium (Penicillium nigricans, etc.), the genus Fusarium (Fusarium lini IAM 5008, etc.), the genus Gloeosporium (Gloeosporium kaki IAM 5011, etc.), and the genus Aureobasidium (Aureobasidium pullulans var. melanigenum, A-8, ACTT 20612, etc.) and yeasts, for example, the genus Saccharomyces (Saccharomyces cerevisiae, etc.), the genus Phodotorulla (Rhodotorulla glutinis, etc.), the genus Pichia (Pichia miso, etc.), the genus Hansenula (Hansenula miso, etc.), and the genus Candida (Candida tropicalis, etc.) and as asparagus and Jerusalem artichoke.Fructosyl transferase of microorganism origin can be obtained by cultivating each microorganism at an optimal temperature for the microorganism, i.e., from 25"C to 30"C, for from 24 to 96 hours in a suitable medium, for example, a medium containing 5.0% sucrose, 1.0% peptone, 0.7% meat extract, and 0.3% NaCI and, after completion of cultivation, removing cells of the microorganism by filtration or centrifugation and so forth to obtain a culture filtrate. There can be used the filtrate itself or an enzyme obtained by purifying the filtrate according to a conventional process known for the purification of enzyme, such as ultrafiltration, salting out with ammonium sulfate, solvent precipitation, gel filtration, or ion-exchange chromatography.The enzyme of plant origin can be obtained by destroying vegetable tissue through physical means such as grinding, and then extracting the enzyme. The crude extract itself or an enzyme obtained by purifying the extract in a conventional manner can be used.

The desired low-calorie sweetener can then be obtained by subjecting sucrose to the action of the enzyme thus obtained. As a result of various investigations on suitable conditions for the industrial transfer reaction, the following conditions have been found preferable. The sucrose concentration in the transfer reaction is adjusted to from 5% to 70%, preferably from 30% to 60%. The reaction pH and the reaction temperature, varying with the origin of the enzyme, are from 4.0 to 7.0 and from 25"C to 65 C, preferably 50"C to 60 C, respectively. As to the enzyme quantity used, from 5 to 200 units, preferably from 20 to 80 units, of the enzyme is used per g of sucrose.The enzyme quantity is herein indicated in terms of "units" by defining, as one unit, an enzyme quantity having the activity of yielding 1 ymole of glucose per 2.5 ml of the reaction solution when the reaction is conducted by adding 0.5 ml of an enzyme solution to 1.0 ml of a 5% sucrose solution and 1.0 ml of a buffer solution having the pH 5.0 followed by warming at 40"C for 60 minutes.

After completion of the transfer reaction, the reaction mixture is heated to inactivate the enzyme, decolorized with activated carbon, desalted with an ion-exchange resin, and concentrated to obtain the end product. Analysis of the composition resulting from the transfer reaction can be conducted by high-speed liquid chromatography using Microbondapack CH column (manufactured by Waters, Ltd.) and a solvent system of acetonitrile/water 0: 20 (v/v)].

The low-calorie sweetener thus obtained has a composition of, for example, 30% glucose, 11% sucrose, 28% GF2, 25% GF3, 5% GF4, and 1% GF5. This constituent sugar composition, however, can greatly vary depending upon the reaction conditions.

Of the oligosaccharides, as GF2, there are illustrated O-ss-D-fructofuranosyl-(2#1)-O-ss-fructofu- ranosyl-(2 1 )-a-D-glucopyranoside, O-ss-D-fructofuranosyl-(2 < 6)-O-ss-glucopyranosyl-( 1 o2)-ss-D- fructofuranoside, O-ss-D-fructofuranosyl-(2 > 6)-O-ss-fructofuranosyl-(2e 1 )-a-D-glucopyranoside, etc.; as G F3, there are illustrated O-ss-D-fructofuranosyí-(2o[1-O-ss-D-fructofuranosyl-2]2o1)-a-D- glucopyranoside, O-ss-D-fructofuranosyl-(2#6)-O-[ss-D-fructofuranosyl-(2#2)]-O-α-D-glucopyrano- syl-( 1 2)-ss-D-fructofuranoside, etc.; and as G F4, there are illustrated 0-fl-D4ructofuranosyl- (2#[1-O-ss-D-fructofuranosyl-2]3#1)-α-D-glucopyranoside, etc.

Of G F2 0--D4ructofuranosyI-(2 1 )-O-P-f ructof u ra nosyl-(2-, 1 )-a-D-glucopyranoside (hereinafter referred to as 1 -ketose), of GF3 O-ss-D-fructofuranosyí-(2 < [1-O-ss-fructofuranosyl-2]241)-a- D-glucopyranoside (hereinafter referred to as nystose), and of GF4 O-ss-D-fructofuranosyl-(2#[1- O-P-D-fructofuranosy1-2],1 )-a-D-glucopyranoside are low-calorie as shown in Test Example 1 and Test Example 2 described hereinafter; therefore, oligosaccharide compositions containing these are necessarily low-calorie sweeteners.

The sweetness of the low-calorie sweetener, obtained by causing fructosyl transferase to act on sucrose, which is composed of glucose, sucrose, and fructooligosaccharides such as GF2, GF3, and GF4 is 60 to 80 when the sweetness of sucrose is defined as 100, whereas a lowcalorie sweetener comprising only fructooligosaccharides is 30 to 60 by the same standard.

Since these sweeteners are a mixture ranging from monosaccharides to pentasaccharides or a mixture ranging from trisaccharides to pentasaccharides, they have viscosity close to that of sucrose, and their physical and chemical properties necessary for food processing such as formretention and moisture-retention fall within the range of various physical properties possessed by sugars and sugar alcohols that have heretofore been used in food processing; therefore, the sweeteners can also impart various physical properties required for food processing as well as sweetness to food and drink.Accordingly, the food and drink containing such sweeteners increases calorie less than does the food and drink containing sugars and sugar alcohols that have heretofore been used; thus these sweeteners can provide a low-calorie or non-calorie food and drink suited to a therapeutic diet or a beauty diet for diabetics and obese persons.

Further, the main ingredients of the sweetener according to the present invention, i.e., GF;, GF3 and GF4, are not susceptible to decomposition due to a disaccharide decomposing enzyme of rabbit intestine so that an increase in blood sugar is greatly suppressed when compared with cases of using sucrose or glucose. This means that the sweetener composition of the present invention is a low-calorie sweetener.

These facts are demonstrated by Test Examples 1 and 2. In tests. as GF2, GF3 and GF4, O-ss- D-fructofuranosyl-(2#1)-O-ss-D-fructofuranosyl-(2#1)-α-D-glucopyranoside (hereinafter referred to as 1-ketose), O-ss-D-fructofuranosyl-(2#[1-O-ss-D-fructofuranosyl-2]2#1)-α-D-glucopyranoside (hereinafter referred to as nystose), and O-b-D-fructofuranosy1-(2-[1 -O-P-D-fructofuranosyl- 2]3o1)-a-D-glucopyranoside (hereinafter referred to as 1 F-fructofuranosyl-nystose), respectively.

Test Example 1 Intestinal disaccharide decomposing enzyme was prepared from the intestinal mucous membrane of a rabbit (body weight: 3 kg) according to the method of Y. Takesue (Journal of Biochemistry, Vol. 65, 545 (1969)). The crude enzyme system showed 280 U/ml of sucrose activity, 540 U/ml of maltose activity and 8 U/ml of trehalase.

A mixture of 1.0 ml of a 5% substrate, 1.0 ml of a 0.25 M phosphate buffer solution (pH 6.5) and 0.5 ml of the above prepared crude enzyme system was allowed to react at 37"C for 24 hours. The resulting reaction mixture was subjected to high pressure liquid chromatography using a column of IL Bondapack CH (a registered trademark manufactured by Waters, Ltd.) and acetonitrile-water (75: 25) as an eluent to obtain a rate of decomposition. The results obtained are shown in Table 1. The rate of decomposition was calculated according to the following formula.

wt% of unreacted substrate based on Rate of Decomposition (%) = 100 -- total solid weight of the reaction mixture after 24 hrs' reaction Table 1 Decomposition by Rabbit Intestinal Disaccharide Decomposing Enzyme Rate of Substrate Decomposition Sucrose 100 Maltose 100 1-Kestose 0 Nystose 0 1 F-Fructofuranosyl-Nystose 0 As is shown in Table 1 above, 1-kestose, nystose and 1 F-fructofuranosyl-nystose are not at all decomposed by the rabbit intestinal disaccharide decomposing enzyme.

Test Example 2 Wistar male rats (body weight: 1 70 g; 30 rats per group) were fasted for 1 7 hours. Each of the indicated saccharides was orally administered to the rats at a dose of 3 g/kg. After 30, 60, 90, 1 20 or 1 80 minutes from the administration, the blood was taken from 6 rats in each group, and the glucose level in blood was determined by a glucose oxidase method. The results obtained are shown in Table 2.

Table 2 Changes in Blood Sugar Level in Fasted Rats 30 Min. 60 Min. 90 Min. 120 Min. 180 Min.

Control 100 100 100 100 100 (no administration) Sucrose 287 254 221 163 177 Glucose 341 286 185 163 163 Fructose 236 252 246 232 183 1-Kestose 139 122 121 123 123 Nystose 123 109 106 108 119 1 F-Fructofuranosyl-Nystose 1 20 109 104 110 109 The values given in Table 2 above were represented as compared with the blood sugar level (mg/dl) of the control group which was taken as 1 00. The blood sugar levels of the control group after 30, 60, 90, 120 and 180 minutes were 61 j43, 65 + 5.0, 67 + 2.3, 73 j 7.0, and 64 + 3.7, respectively.

As can be seen from Table 2, no increase in blood sugar was observed in 1-kestose, nystose and 1 F-fructofuranosyl-nystose groups. This indicates that the fructooligosaccharides of the present invention are not absorbed into bodies and, therefore, do not furnish substantial calories.

The low-calorie sweetener and the process for preparation of a low-calorie food and drink with the use thereof will be explained below in detail by citing illustrative examples.

Example I 10 ml portions of the BS medium containing 5.0% sucrose, 1.0% peptone, 0.7% meat extract, and 0.3% NcCI were each poured into two test tubes. After sterilizing at 120"C for 30 minutes, each of the media was inoculated with a platinum wire loop of Aspergillus nigerACE-2 1, FERM-P5886, and cultivation was conducted at 28"C for 24 hours.

10 ml portions of the culture broth obtained were each inoculated into two Erlenmeyer flasks containing each 200 ml of the BS medium (sterilized at 120"C for 30 minutes), and shaking culture was conducted at 28"C for 24 hours as precultivation.

In a 30 I jar fermentor was charged 20 1 of the BS medium. After sterilizing at 120"C for 30 minutes, the BS medium was cooled and inoculated with 400 ml of the foregoing preculture broth. Cultivation was conducted at 300 rpm at 28"C for 72 hours. After completion of the cultivation, cells were removed by filtration to obtain 20 1 of a culture filtrate. 20 1 of this culture filtrate was concentrated and purified by ultrafiltration to obtain 2 1 of an enzyme solution having an enzyme activity of 240 units/ml.

10 kg of sucrose was dissolved by addition of 6.7 1 of water. After the solution had been adjusted to pH 6.0, the enzyme was added thereto in an amount of 48 units per g of sucrose, and the transfer reaction was conducted at 50"C for 72 hours. After completion of the transfer reaction, the reaction mixture was heated at 100or for 1 5 minutes to inactivate the enzyme followed by adding cultivated carbon in a proportion of 0.5% based on solids to decolorize the reaction mixture. After removal of the activated carbon, the reaction mixture was treated with ion-exchange resins of Amberlite IR'120B and Amberiite IRA411 and then concentrated to a 75% w/w concentration to obtain 1 2 kg of a low caloric sweetener.

The sweetener thus obtained had a sugar composition of 33% glucose; 4% fructose; 10% sucrose; 29% GF2; 20% GF3; and 4% GF4.

There will next be described examples in which the low-calorie sweetener of the present invention was applied to the preparation of food and drink. The compositions of the low-calorie sweeteners used in the examples are as follows: Sweetener A: 36% glucose, 2% fructose, 10% sucrose, 23% 1-kestose, 24% nystose, 6% 1 F4ructofuranosyl-nystose Sweetener B: 45% 1-kestose, 46% nystose, 9% 1 F-fructofuranosyl-nystose Example 2 Preparation of sweet paste of steamed beans (also referred to as ''neri-yokan''): 1 2 g of agar-agar was soaked in water for 3 hours, and ground after removing water. Then 260 ml of water was added thereto and the mixture was heated to provide a solution.

Sweetener A (960 g) (water content: 25% w/w) was added thereto, and the mixture filtered after the agar-agar had been completely dissolved. The agar-agar was placed over a fire, and 500 g of raw bean-jam was added thereto, and the mixture kneaded. After it had been boiled down until the sugar content had reached 70 to 71%, the mixture was put into a box and hardened to prepare sweet paste of steamed beans (neri-yokan). As a control, sweet paste of steamed beans (neri-yokan) was prepared in the same manner with the use of 720 g of sugar in place of Sweetener A.

The calorific value of 100 g of the sweet paste of steamed beams containing Sweetener A was 1 55 cal, whereas that of the control was 266 cal.

Example 3 Preparation of chewing gum.

75 parts of the low-calorie Sweetener B powder and 22 parts of chicle rubber were dissolved and mixed. Then, a flavor and menthol were added thereto followed by mixing and kneading.

The mixture was rolled into a given thickness by a rolling mill, and then the rolled product was cut and dried to prepare plate gum. As a control, plate gum was prepared in the same manner with the use of 50 parts of sugar and 25 parts of glucose in place of Sweetener B.

The gum prepared with the use of Sweetener B is essentially non-calorie, the calorific value per 10 g of the control gum being 30 cal.

Claims

1. A low-calorie sweetener containing at least one oligosaccharide consisting of from one to four molecules of fructose bound to sucrose.

2. A sweetener as claimed in claim 1, containing a mixture of said oligosaccharides.

3. A sweetener as claimed in claim 1 or 2, consisting solely of said at least one oligosaccharide.

4. A sweetener as claimed in claim 1 or 2, containing one or more other sweeteners.

5. A sweetener as claimed in claim 4, containing glucose and sucrose.

6. A sweetener as claimed in claim 5, containing up to 36% glucose and up to 11% sucrose.

7. A sweetener as claimed in claimed in any preceding claim, wherein said at least one oligosaccharide is one which has been produced by causing fructosyl transferase to act on sucrose.

8. A sweetener as claimed in claim 1, substantially as hereinbefore described in Example 1.

9. A process for the preparation of a low-calorie food or drink comprising incorporating into the food or brink a low-calorie sweetener as claimed in claim 1.

10. A process as claimed in claim 9, substantially as herein before described in Example 2.