GB2046757A - Process for producing low cariogenic food sweeteners - Google Patents

Abstract

An additive for low-cariogenic and/or anti-caries food or drinks comprises aldosylfructoside obtained by subjecting a substrate solution containing aldose, and sucrose and/or raffinose to the action of levansucrase (E.C.2.4.1.10). Such an aldosylfructoside can be used in the production of any product which is used orally as well as food and drink in general. The levansucrase may be obtained from Actinomyces viscosus, Aerobacter levanicum, Acetobacter suboxydans, Bacillus licheniformin, Bacillus subtilis, Gluconobacter oxydans Streptococcus mutans or Streptococcus salivalius. The aldose used is preferably D-xylose, L- arabinose, D-galactose, D-mannose, xylobiose, cellobiose, maltose, nomaltose, lactose, kojibiose, laminaribiose, nigerose, xylotriose, cellotriose, maltotriose, isomaltotriose, panose or isopanose or a partial hydrolipate obtained from starch, dextran, pullulan, curdlan, pachyman, elsinan, glucomannan, cellulose or xylan.

Description

SPECIFICATION Process for producing foods and drinks The present invention relates to an additive in particular for food and drinks.

Large amounts of sucrose, a typical sweetener with sufficient sweetness and body, are consumed in the production of food and drinks. Recently, it has been demonstrated that sweetened foods and drinks, particularly those sweetened with sucrose, very often cause dental caries. Dental caries are generally caused when sucrose in the foodstuff is converted by oral bacteria into a water-insoluble glucan such as dextran which adheres in layers to the surface of the teeth, further sugar passes through the layers and reaches the tooth surface where it is fermented anaerobically into organic acids, whereupon the acids attack the tooth enamel.

The present invention is based on the discovery that an aldosylfructoside obtained by subjecting a substrate solution containing aldose, and sucrose and/or raffinose to the action of levansucrase (E.C.

2.4.1.10) has an appropriate sweetness and also low-cariogenic and/or anti-caries properties which render the aldosylfructoside a favorable sweetener for prod'ucing sweeteners low-cariogenic foods and drinks.

Accordingly the present invention provides a process for producing an additive for foods and drinks, including the steps of subjecting a substrate solution containing an aldose, and sucrose and/or raffinose to the action of levansucrase (E.C. 2.4.1.10) thereby to promote an enzymatic transfructosylation reaction, and recovering the resultant aldosylfructoside.

The aldoses usable in the present invention are those aldoses which can be converted into aldosylfructosides (with the exception of sucrose and raffinose) by transfructosylation with the said levansucrose. Namely, aldosyl mono-, di- and tri-saccharides excluding glucose and melibiose are preferable: for example one or more of D-xylose, L-arabinose, D-galactose, D-mannose, xylobiose, cellobiose, maltose, isomaltose, lactose, kojibiose, laminaribiose, nigerose, xylotriose, cellotriose, maltotriose, isomaltotriose, panose and isopanose. In addition, sugar mixtures, D. E. (dextrose .equivalent) about 10 to 70, prepared by partial hydrolysis of polysaccharides such as starch, dextran, pullulan, curdlan, pachyman, elsinan, glucomannan, cellulose and xylan, are usable in the present invention.

The substrate solution preferred in the present invention is an aqueous solution containing an aldose which acts as an acceptor for a fructose residue, and sucrose and/or raffinose which act as donors of the fructose residue, when the transfructosylation reaction in the presence of levansucrase takes place.

A mole ratio of acceptor (i.e. the aldose) to the donor (i.e. the sucrose and/or raffinose) of about 1:50 to 50:1, and a substrate concentration of about 5 to 40 w/w %, are preferable.

The levansucrase (E.C. 2.4.1.10) employable in the present invention is that which forms non reducing aldosylfructosides when allowed to act on the above-described substrate solution containing aldose, and sucrose and/or raffinose, to transfer the fructosyl residues of sucrose and/or raffinose to the reducing C1-sites of the aldoses. For example, levansucrase from Actinomyces viscosus, Aerobacter levanicum, Acetobacter sub ox ydans, Bacillus licheniformis, Bacillus subtilis, Gluconobacter oxydans, Streptococcus mutans, Streptococcus salivalius and other micrnorganisrns is advantageously usable in the present invention.

In order to prepare the levansucrase from the microorganisms, submerged culture is usually employed. The culture broth can be used without pre-treatment, but its supernatant or filtrate is usually used after removing water-insoluble substances by centrifugation or filtration. In some cases, the micro organism can be used intact as an enzyme preparation with extraction, as well as levansucrase extracted from the cells by conventional methods. The levansucrase thus obtained is, if desired, purified further according to conventional methods. In addition, immobilized levansucrases are also usable in continuous or batch-wise operation.

In the production of the aldosylfructoside, the levansucrase is added to a substrate solution containing aldose, and sucrose and/or raffinose to effect the enzymatic transfructosylation reaction. In this case, any reaction temperature and pH can be applied so long as the levansucrase acts on the substrate to produce the corresponding aldosylfructoside; generally a pH range of 3 to 10 and a temperature range of about 20 to 800C are preferable. The amount of levansucrase is usually about 0.01 to 1,000 units per g sucrose and/or raffinose as defined by the following assay method, and the transfructosylation reaction is performed usually for about 0.1 to 100 hours.

Assay of levansucrase activity Two ml of a reaction mixture containing 10 w/v % sucrose, 50 mM phosphate buffer, pH 7.0, and levansucrase is incubated at 30"C for 30 minutes. The reaction mixture is then heated to inactivate the enzyme and the amount of the glucose released is determined by the glucose-oxidase method. One unit of levansucrase activity is defined as the amount of enzyme that produces mole of glucose per minute under the above conditions.

The reaction mixture containing the aldosylfructoside can be, if desired, treated further with other enzymes such as glucose isomerase which isomerizes the glucose released in the reaction mixture into fructose and increases the sweetness of the product.

The transfructosylation reaction is usually suspended by heating and the reaction mixture is filtered. The filtrate is decolorized with activated carbon (charcoal) and deionized with ion exchangers, for example those of H-form and OH-form. The purified solution containing the aldosylfructoside is concentrated into syrup, or dried and pulverised into powder for use in the production of foods and drinks. if necessary, the produced aldosylfructoside can be isolated from the reaction mixture or the purified solution.

In respect to concentration, drying and pulverization, various conventional methods are employable in the present invention, for example evaporation and drying under reduced pressure, and spray-drying.

The sweetness of the sweetener containing aldosylfructoside thus obtained is equal to or slightly higher than that of the substrate solution, and much milder on the basis of dry solid. in addition, the present sugar mixture containing aldosylfructoside offers an advantage that it does not cause crystallization no matter how high the concentration is or how long the period of storage is, although the composition of the substrate solution is very liable to crystallization. Furthermore, the sugar mixture has a sufficient viscosity and moisture-holding capacity, which can be used for imparting a sufficient viscosity, moisture retention capacity, gloss and body to food and drinks, and improving their textures, as well as sweetening them.

For sweetening foods and drinl < s, the sweetener containing aldosylfructoside can be used alone or, if necessary, in combination with or in mixture with other sweeteners such as sucrose, glucose, maltose, corn syrup, isomerized sugar, honey, maple sugar, sorbitol, maltitol, lactitol, dihydrochalcone, Lasparatyl-L-phenylalanine methyl ester, saccharin, glycine, alanine, glycyrrhizin and stevioside; and with fillers such as starch, dextrin and lactose; colouring agents, flavouring agents and seasonings.

Particularly, the sweeteners containing aldosylfructoside differ from sucrose and can be used advantageously as a main- or sub-ingredient to produce foods and drinks because of their lowcariogenic and/or anti-caries properties. Also, since the sweeteners containing aldosylfructoside are compatible with various sour, salty, delicious, astringent, or bitter-tasting substances, they can be used to sweeten foods and drinks in general, and favorite foods, and to improve their tastes: for example together with various seasonings such as soy, powder soy, mayonnaise, dressings, vinegars, sauces, catsups and curry roux;Japanese style confectioneries, western style confectioneries such as breads, biscuits, crackers, pies, puddings, butter creams, cookies, custard creams, waffles, sponge cakes, doughnuts, chocolates, chewing gums, caramels and candies; ice creams and sherbets; syrups; pastes such as flour pastes, peanut pastes and fruit pastes; preserved fruits and vegetables such as jams, marmalades and preserves; pickles and pickle products; meat products such as hams and sausages; fish products such as fish hams and fish sausages; various dainties; daily dishes; bottled foods such as those of fishes, meats, fruits and vegetables; canned foods; liquors such as Japanese Sake, wines, whiskies, brandies and alcohol drinks; drinks such as coffee, cocoa, juices, carbonated beverages, lactic acid drinks and those containing lactic acid-producing microorganisms; and convenience foods such as those of puddings, cakes, juices and coffee.

Also, the sweeteners prepared according to the present invention are usable for improving the taste of feeds and pet foods for domestic animals and fowls, honey bees, silkworms, fishes and other animals. In addition to the above-described uses, the sweeteners can be used for sweetening, improving and altering tastes of various favorites, cosmetics, drugs and medicines in solid, paste and liquid forms; for example tobacco, tooth pastes, lipsticks, lipcreams, medicines for internal administration, troches, drops containing liver oil, refrigerants, cachous and gargles.

The terms food and drinks used throughout this specification are used to mean all foods and drinks in general, favorites, feeds, pet foods, cosmetics, drugs, medicines and all products which used orally.

Any method of incorporating the aldosylfructoside into the foodstuff so long as it is admixed in the foodstuff in a step prior to final processing, for example mixing, kneading, soaking, scattering, applying and injecting. An aldosylfructoside content of about 1% or more per foods and drinks d.s.b. is required when only inhibition of cariogenicity is desired.

The present invention is illustrated further by the following Experiments which describe the preparation and characteristics of aldosylfructosides.

EXPERIMENT 1 Preparation of levansucrase 1-1. Levansucrase of Bacillus Sixty litres of liquid medium, consisting of 3 w/v % defatted soybean, 2 w/v % glucose, 4 w/v % sucrose, 0.6 w/v % (NH4)2HPO4, 0.03 w/v % MgSO4. 7H2O, 0.02 w/v % KCI, 0.02 w/v % calcium acetate, 0.001 w/v % MnSO4. 4H20 and water, was adjusted to pH 7.0, sterilized at 1 200C for 20 minutes, cooled and incoculated with Bacillus subtilis ATCC 6051 and was cultured at 370C for 3 days under agitation 'under aerobic conditions. After completion of the cuitivation, the culture broth was centrifuged to obtain a supernatant. Then the supernatant was added with equivolume of cold ethanol, and the formed percipitate was centrifuged, collected, dissolved in a 20 mM acetate buffer containing 1 mM calcium chloride, pH 5.0, and dialyzed against a fresh buffer of the same composition overnight.

The resultant solution was centrifuged and the obtained supernatant was passed through a DEAE cellulose-packed column to adsorb the levansucrase, and the column was washed with a fresh buffer of the same composition. The adsorbed enzyme was eluted with a fresh buffer of the same composition except that it contained 1 M NaCI additionally. The eluate was saturated to 90% saturation with ammonium sulfate and the formed precipitate was centrifuged, collected and dissolved in 500 ml of the same buffer. The levansucrase activity of the solution was about 120 units per ml.

1-2. Levansucrase of Aerobacter Fifty litres of liquid medium, consisting of 0.2 w/v % pepton, 1 w/v % sucrose, 0.7 w/v % K2HPO4, 0.3 w/v % KH2PO4,0.05 0.05 w/v%MgS04.7H20, 0.05 w/v % KCI, 0.001 w/v % FeSO4,7H20 and water, was sterlized at 1200C for 20 minutes, and inoculated with Aerobacter levancum ATCC 1 5552, and the mixture was cultured at 300C for 3 days under agitation under aerobic conditions. The culture broth was treated similarly as described in Experiment 11 and 600 ml of a levansucrase solution was obtained. The activity of the solution was about 1 50 units per ml.

EXPERIMENT 2 Preparation of aldosylfructoside To a substrate solution, pH 6.0, containing 0.1 M sucrose as donor and 0.5 M of a member of a group comprising L-arabinose, D-xylose, D-mannose, D-galactose, maltose, isomaltose, lactose, cellobiose and maltotriose, was added a levansucrase, 2 units per g sucrose, prepared by the method as described in Experiment 11, and the resulting solution was incubated at 400C for 44 hours to effect the enzymatic transfructosylation reaction.

Analysis of the reaction mixtures according to the oligosaccharide map method described by J. H.

Pazur and S. Okada, "The Journal of Biological Chemistry", vol. 241, pp 4146-4151(1960), demonstrated in each case the transfer of the fructose residue of sucrose to the reducing group of the acceptor aldose and formation of corresponding non-reducing aldosylfructoside; arabinosylfructoside, xylosylfructoside, mannosylfructoside, galactosylfructoside, maltosylfructoside, isomaltosylfructoside, lactosylfructoside, cel lobiosylfructoside and maltotriosylfructoside.

Isolation of the aldosylfructosides from the reaction mixtures was performed by a conventional method using an activated carbon-packed column including the steps of: passing the reaction mixture through the column to adsorb all the sugars, eluting and fractionating the adsorbed sugars with aqueous alcohol on concentration gradient, and then concentrating and drying the eluate containing aidosylfructoside into powder.

Hydrolysis of the aldosylfructosides using a commercial yeast p-fructofuranosidase (E.C. 3.2.1.26) demonstrated that every aldosylfructoside was hydrolyzed into fructose and starting aldose. Therefore, it was shown that the present aldosylfructosides were aldosyl-,B-fructofuranosides. The yields of the respective aldosylfructosides of high purity were about 15% on a mole basis of material aldoses. The aldosylfructosides were mildly sweet.

EXPERIMENT 3 Low-cariogenic and anti-caries properties of aldosylfructoside In this Experiment the formation of lactic acid and water-insoluble glucan by an oral bacterium was determined using the aldosylfructosides prepared by the method described in Experiment 2.

3-1. Formation of lactic acid 1.8 ml Aliquots of a mixture, containing a 0.1 M phosphate buffer, pH 6.8, 5 mM MgCl2 and a cell suspension of Streptococcus mutans 6715, about 2.5 mg d.s.b., were shaken at 350C for 5 minutes and the reaction was started by an addition of 0.2 ml of a 0.1 M aqueous aldosylfructoside solution to the aliquots, and 20 minutes later, was stopped by an addition of 0.2 ml of a 25 w/v % metaphosphoric acid solution. Then the aliquots were centrifuged and the amount of lactic acid formed in the supernatants was assayed according to the lactate dehydrogenase method. A control experiment was carried out similarly, except that the aldosylfructoside was replaced by sucrose.The results are shown in the Table below, wherein the amount of lactic acid is expressed as a percentage of the lactic acid found in the control.

3-2. Formation of water-insoluble glucan A commercial heart infusion medium containing bovine heart extract, pepton and NaCI was dissolved to give a concentration of about 2.5 w/v %, and respective aldosylfructosides prepared by the method described in Experiment 2 were added thereto to give aldosylfructoside concentrations of about 2 w/v %. Streptococcus mutans 671 5 was inoculated in 4 ml of the above mixtures and the resulting mixtures were incubated at 370C for 1 6 hours. After cultivation, the cultures were centrifuged. 4 ml of a 0.5 N NaOH was added to each of the resultant precipitates and these were then incubated at 370for an additional one hour to dissolve the water-insoluble glucan.The resulting mixtures were centrifuged again and the amount of the water-insoluble glucan formed in the supernatants was assayed by the phenol-sulfuric acid method. A control experiment was carried out similarly, except that the aldosylfructoside was replaced by sucrose. The results are shown in the Table below, wherein the amount of water-insoluble glucan in each sample is expressed as a percentage of the water-insoluble glucan in the control.

3-3. Inhibition of pH-decrease and formation of water-insoluble glucan from sucrose Streptococcus mutans 671 5 was inoculated in a manner similar to that of Experiment 3-2 at 37"C for 1 6 hours in a medium in which 80% of the aldosylfructosides were replaced by sucrose. The pH-level of the mixtures was monitored with a pH-meter and the amount of the formed water-insoluble glucan was assayed as described in Experiment 3-2. A control experiment was carried out similarly, except that sucrose was used in an amount of 2 w/v % and the aldosylfructosides were omitted. The results are shown in the Table below, wherein the amount of water-insoluble glucan is expressed as a percentage of the water-insoluble glucan in the control.

From the results shown in the Table, the results of Experiments 3-1 and 3-2 demonstrate that aldosylfructosides lead to much less formation of lactic acid and water-insoluble glucan than the control, and the results of Experiment 3-3 demonstrate that use of the aldosylfructoside lead to highinhibition of formation of water-insoluble glucan and pH-decrease from sucrose. From the abovedescribed results, it can be concluded that aldosylfructosides are suitable as low-cariogenic and/or anticaries sweeteners.

TABLE

EXPERIMENT 3 - 1 3 - 2 3 - 3 Lactic acid Water-insoluble Water-insoluble pH Aldosylfructoside (%) glucan (%) glucan (%) Sucrose (control) 100 100 100 4.1 Arabinosylfructoside 2.3 3.3 23.6 5.2 Xylosylfructoside 1.5 2.1 16.8 5.4 Mannosylfructoside 10.2 12.4 36.0 5.0 Galactosylfructoside 13.6 15.7 38.1 5.2 Maltosylfructoside 4.5 8.2 33.7 5.5 Isomaltosylfructoside 1.8 3.2 16.4 6.0 Lactosylfructoside 3.6 7.7 44.2 6.2 Cellobiosylfructoside 1.2 8.9 22.5 6.1 Maltotriosylfructoside 2.6 10.4 40.4 5.6 The present invention wiil be further illustrated by following Examples.

EXAMPLE 1 Sweetener A solution was prepared by dissolving 3 kg of sucrose and 1 kg of xylose in 1 0 1 of water. A levansucrase, 5 units per g sucrose, prepared by the method described in Experiment 1-1 was added thereto and the solution was then incubated at 400C and pH 6.0 for 1 6 hours to effect an enzymatic transfructosylation reaction. Then the enzyme was inactivated by heating and the reaction mixture was filtered. The filtrate was decolorized with activated carbon and deionized with ion exchangers of H-form -and OH-form according to conventional methods, and concentrated to give a syrup having a water content of about 20 w/w %, relatively low viscosity and high sweetness. The yield of the strup sweetener was about 95% against material sugars d.s.b.

The syrup which has a xylosylfructoside content about 30 w/w % d.s.b. greatly inhibits the occurrence of dental caries when used alone and when used as a mixture with sucrose; it is thus an effective iow-cariogenic and/or anti-caries sweetener.

EXAMPLE 2 Sweetener A solution was prepared by dissolving 40 kg of sucrose and 20 kg of maltose in 100 1 of water. A levansucrase, 2 units per g sucrose, prepared by the method described in Experiment 1-1 was added thereto and the solution was then incubated at 400C and pH 6.0 for 44 hours to effect the transfructosylation reaction. The reaction mixture was purified and concentrated in a manner similar to that described in Example 1 to give a water content of about 30 w/w %, and then spray-dried into white powder having a relatively high sweetness. The yield was about 90% against material sugars d.s.b.

The resultant sweetener, having a maltosylfructoside content about 30% d.s.b., is feasible as a low-cariogenic sweetener.

EXAMPLE 3 Sweetener A solution was prepared by dissolving 1 kg of sucrose and 5 kg of a high maltose syrup, water content 25 w/w % and D.E. about 60, in 7 1 of water. A levansucrase, 10 units per g sucrose, prepared by the method described in Experiment 1-2 was added thereto and the solution was incubated at 350C and pH 5.5 for 14 hours to effect the enzymatic reaction. Then the reaction mixture was purified and concentrated in a manner similar to that described in Example 1 to give a syrup having a relatively high sweetness and a water content of about 20 w/w %. The yield was about 93% against material sugars d.s.b.

The syrup, containing about 10% of aldosylfructosides excluding sucrose d.s.b., is suitable as a low-cariogenic sweetener and for imparting a proper moisture retention capacity and gloss to foods and drinks.

EXAMPLE 4 Sweetener A solution was prepared by dissolving 1 kg of sucrose and 2 kg of a partially-hydrolyzed powder dextran, D.E. about 30, in 4 1 of water. A levansucrase, 2 units per g sucrose, prepared by the method described in Experiment 1-2 was added thereto, and the solution was then incubated at 350C and pH 5.5 for 40 hours to effect the enzymatic reaction. The reaction mixture was purified and concentrated in a manner similar to that described in Example 1 to give a syrup sweetener having a water content of about 20 w/w % and a moderate sweetness. The yield was about 96% against starting materials d.s.b.

The syrup which was a content of aldosylfructosides excluding sucrose of about 25% d.s.b., greatly inhibits the occurrence of dental caries when used alone or as a mixture with sucrose. It is an excellent low-cariogenic and/or anti-caries sweetener. In addition, the sweetener is suitable for imparting a proper sweetness, moisture retention capacity and gloss to foods and drinks.

EXAMPLE 5 Sweetener A solution was prepared by dissolving 50 kg of sucrose and 10 kg of lactose in 70 1 of water. A levansucrase, 1 unit per g sucrose, prepared by the method as described in Experiment 1-2 was added to the solution and the solution was then incubated at 400C and pH 6.0 for 40 hours to effect the enzymatic reaction and then incubated at 600C for an additional 5 hours after addition of a commercial glucose isomerase, 20 units per g sucrose. The reaction mixture was purified and concentrated in a manner similar to that described in Example 2, and then spray-dried to give a powder sweetener having a relatively high sweetness. The yield was about 92% against material sugars d.s.b.

The powder product, containing about 20% of lactosylfructoside d.s.b., is suitable as a lowcariogenic sweetener.

EXAMPLE 6 Sweetener A solution was prepared by dissolving 2 kg of raffinose and 1 kg of a powder maltodextrin, D.E.

about 20, in 4 1 of water. A levansucrase, 3 units per g raffinose, prepared by the method as described in Experiment 1-1 was added thereto and the solution was then incubated at 400C and pH 6.0 for 20 hours to effect the enzymatic reaction. Then the reaction mixture was purified and concentrated in a manner similar to that described in Example 1 to give a syrup sweetener having a low sweetness, high viscosity and water content of about 20 w/w %. The yield was about 94% against starting materials d.s.b.

The syrup thus obtained, having a content of aldosylfructosides (excluding raffinose) about 25% d.s.b., is suitable as a low-cariogenic sweetener and can be used to impart a proper viscosity, moisture retention capacity and gloss to foods including drinks.

EXAMPLE 7 Sweetener A syrup sweetener mixture was prepared by dissolving 250 g of a powder sweetener as prepared by the method described in Example 2 in 1 kg of a hydrogenated maltose syrup, water content about 25 wlw %.

The mixture has the same sweetness as sucrose, and therefore is suitable as a diet sweetener for diabetics and obese persons as well as a low-cariogenic sweetener. Also, the sweetener offers an advantage that foods can be prepared with much less browning when the sweetener is used because it tolerates heating and is thus less liable to impart undesired colour to foods. In addition, the sweetener is also suitable for imparting a sufficient moisture retention capacity and gloss to foods including drinks.

EXAMPLE 8 Hard candy Ten kg of a syrup prepared by the method described in Example 7 was heated and evaporated under reduced pressure to a moisture content of below 2 w/w %, mixed with 100 g of citric acid, small amounts of a lemon flavouring and a colouring agent and shaped into hard candy using a conventional method.

The hard candy is low-cariogenic.

EXAMPLE 9 Chewing gum Two kg of a gum base was heated and softened, and mixed with 7 kg of a powder sweetener prepared by the method described in Example 5, small amounts of peppermint and colouring matter.

Then the mixture was kneaded with a roller and shaped into chewing gums using a conventional method.

The chewing gum has a good taste and texture, and low-cariogenicity.

EXAMPLE 10 Chocolate Forty kg of cacao, 10 kg of cacao butter, 1 5 kg of a powder sweetener prepared by the method described in Example 2 and 20 kg of a whole milk were mixed and then passed through a refiner to reduce the solid particles. Then 500 g of lecithin was added to the mixture and the mixture was kneaded at 500C for 2 days in a conche. The resulting mixture was moulded and solidified according to a conventional method.

The low-cariogenic chocolate thus obtained is less liable to fat and sugar blooms, and also very tasty and appealing to the palate.

EXAMPLE 11 Lactic acid drink Then kg of skimmed milk was pasteurized by heating at 800C for 20 minutes and cooled to 4COOC.

To the milk was then added 300 g of a starter and the milk was then fermented at a temperature of about 35 to 370C for 10 hours. The obtained mixture was homogenized, and 7 kg of a syrup prepared by the method described in Example 3 was added thereto. The mixture was pasteurized by heating at a temperature of about 60 to 650C while suppressing decomposition of the aldosylfructoside contents.

The mixture was mixed with a small amount of flavouring agent and bottled after cooling.

The drink is suitable as a low-cariogenic drink, and its sweetness and flavour are compatible with its sour taste.

EXAMPLE 12 Tsukudani (Japanese style preserved food boiled down in soy) After removing sand from 250 g of tangle, the tangle was treated with an acid, cut into small squares according to conventional method and soaked in a solution comprising 21 2 ml of soy, 31 8 ml of amino acid solution and 100 g of a syrup prepared by the method as described in Example 4. The mixture was added additionally with 12 g of sodium glutamate, 8 g of caramel and 21 ml of Mirin (sweet Sake) while boiling the mixture. Then the mixture was boiled down to Tsukudani.

The product was an appetizing and appealing Tsukudani in colour and gloss as well as in flavour.

EXAMPLE 13 Pickled scallion Five kg of fresh scallions were soaked in 2.5 1 of a 20 w/v %'saline solution for 3 weeks according to a conventional method and the solution was drained off. The salted scallions were pickled for one month in an acetic acid solution containing 80 g of sodium chloride, 80 ml of glacial acetic acid and 2.0 1 of water. The pickled scallions thus obtained were then picl < led in a fresh pickle solution consisting of 800 ml of vinegar, 200 ml of Mirin, 0.5 g of stevioside, 10 g of red pepper and 1 50 g of a sweetener prepared by the method described in Example 6 for an additional 10 days to obtain tasty pickled scallions.

EXAMPLE 14 Tablet 50 g of aspirin as admixed homogeneously with 4 g of còrnstarch and 14 g of a powder sweetener prepared by the method described in Example 2. A tabletting machine equipped with a pestle having a diameter of 12 mm and a curvature of 20 R was used to form each tablet, having a weight of 680 mg, a thickness of 5.25 mm and a hardness of 8 + 1 kg.

The tablets are less liable to crack and deform during long periods of storage and can be taken easily because of their pleasant sweetness.

EXAMPLE 1 5 Tooth paste The materials described below were mixed according to a conventional method to prepare a tooth paste.

Formulation Dipotassium hydrogenphosphate 45 % Pullulan 2.95% Sodium lauryl sulfate 1.5 % Glycerin 20 % Polyoxyethylene sorbitan laurate 0.5 % Antiseptic agent 0.05% A liquid sweetener prepared by the method described in Example 1 18 % Water 12 % The tooth paste is suitable for children owing to its appropriate sweetness.

Claims (38)

1. A process for producing an additive for foods or drinks, including the steps of subjecting a substrate solution containing an aldose, and sucrose and/or raffinose to the action of levansucrase (E.C.

2.4.1.10) thereby to promote an enzymatic transfructosylation reaction, and recovering the resultant a Idosylfructoside.

2. A process according to Claim 1, in which the levansucrase is one obtained from Actinomyces viscosus, Aerobacter levanicum, Acetobacter suboxydans, Bacillus licheniformis, Bacillus subtilis, Gluconobacter oxydans, Streptococcus mutans or Streptococcus salivalius.

3. A process according to Claim 1 or 2, in which the aldose is one or more of D-xylose, Larabinose, D-galactose, D-mannose, xylobiose, cellobiose, maltose, isomaltose, lactose, kojibiose, laminaribiose, nigerose, xylotriose, cellotriose, maltotriose, isomaltotriose, panose and isopanose.

4. A process according to Claim 1 or 2, in which the aldose is a partial-hydrolysate obtained from starch, dextran, pullulan, curdlan, pachyman, elsinan, glucomannan, cellulose or xylan.

5. A process according to Claim 4, in which the dextrose equivalent (D.E.) of the hydrolysate is from 10 to 70.

6. A process according to any one of Claims 1 to 5, in which the substrate solution is subjected to the action of the levansucrase for 0.1 to 100 hours.

7. A process according to any one of Claims 1 to 6, in which the enzymatic transfructosylation reaction is carried out at a temperature of from 20 to 800C.

8. A process according to any one of Claims 1 to 7, in which the enzymatic transfructosylation reaction is carried out at a pH of from 3 to 1 0.

9. A process according to any one of Claims 1 to 8, in which the amount of levansucrase is from 0.01 to 1,000 units per g sucrose and/or raffinose.

1 0. A process according to any one of Claims 1 to 9, in which the mole ratio of aldose to sucrose and/or raffinose is from 1: 50 to 50 1.

11. A process according to Claim 1 substantially as hereinbefore described in the Experiments and in any one of Examples 1 to 6.

12. An additive for food and drink whenever prepared by a process as claimed in any one of Claims 1 to 11.

1 3. An additive according to Claim 12, in which the aldosyfructoside is arabinosylfructoside, xylosylfructoside, mannosylfructoside, galactosylfructoside, maltosylfructoside, isomaltosylfructoside, lactosylfructoside, cel lobiosylfructoside or ma Itotriosylfructoside.

14. An additive according to Claim 1 2 substantially as hereinbefore described in any one of Examples 1 to 7.

1 5. A sweetening composition including an additive as claimed in any one of Claims 12 to 14.

1 6. A food or drink incorporating an additive as claimed in Claim 1 2, 1 3 or 14 or a composition as claimed in Claim 1 5.

1 7. A food or drink according to Claim 16, in which the aldosylfructoside content of the food or drink is at least 1% based on the dry solid.

1 8. A food or drink according to Claim 1 6, or 1 7, which is a food or drink in general, a favorite food or a sweetener.

1 9. A food or drink according to Claim 1 6 or 17, which is a feed or pet food for domestic animals and fowls, honey bees, silkworms and fishes; a favorite, a cosmetic, a drug or a medicine.

20. A food or drink according to Claim 1 6 or 1 7, which is a tobacco product, tooth paste, lipstick, lipcream, a medicine for internal administration, troches, drops containing liver oil, a refrigerant, cachou or a gargle.

21. A food or drink according to any one of Claims 1 6 to 20, which is low-cariogenic and/or anticaries.

22. A food or drink according to Claim 1 6 substantially as hereinbefore described in any one of Examples 8 to 1 5.

23. A process for producing food or drink characterised in using aldosylfructoside obtained by subjecting a substrate solution containing aldose, and sucrose and/or raffinose to the action of levansucrase (E.C. 2.4.1.10).

24. A process according to Claim 23, in which the levansucrase is one obtained from Actinomyces viscosus, Aerobacter levanicum, Acetobacter sub ox ydans, Bacillus licheniformis, Bacillus subtilis, Gluconobacter oxydans, Streptococcus mutans or Streptococcus salivalius.

25. A process according to Claim 23 or 24, in which the aldoses are one of more of D-xylose, Larabinose, D-galactose, D-mannose, xylobiose, cellobiose, maltose, isomaltose, lactose, kojibiose, laminaribiose, nigerose, xylotriose, cellotriose, maltotriose, isomaltotriose, panose and isopanose.

26. A process according to Claim 25 or 26, in which the aldoses are partial-hydrolysates obtained from starch, dextran, pullulan, curdlan, pachyman, elsinan, glucomannan, cellulose or xylan.

27. A process according to Claim 26, in which D.E. of the hydrolysates are in the range from 10 to 70.

28. A process according to any of Claims 23 to 27, in which the substrate solution is subjected to the action of levansucrose for 0.1 to 100 hours.

29. A process according to any of Claims 23 to 28, in which the enzymatic transfructosylation reaction is carried out in the temperature range from 20 to 800C.

30. A process according to any of Claims 23 to 29, in which the enzymatic reaction is carried out in the pH range from 3 to 10.

31. A process according to any of Claims 23 to 30, in which the levansucrase is used in the range from 0.01 to 1,000 units per g sucrose and/or raffinose.

32. A process according to any of Claims 23 to 31, in which the mole ratio of aldose vs. sucrose and/or raffinose is in the range from 1 : 50 to 50:1.

33. A process according to any of Claims 23 to 32, in which the aldosylfructoside contents of said foods and drinks are at least 1% on the basis of dry solid.

34. A process according to any of Claims 23 to 33, in which the aldosylfructosides are arabinosylfructoside, xylosylfructoside, mannosylfructoside, galactosyifructoside, maltosylfructoside, isomaltosylfructoside, lactosylfructoside, cel lobiosylfructoside and maltotriosylfructoside.

35. A process according to any of Claims 23 to 34, in which the foods and drinks are foods and drinks in general, favorite foods and sweeteners.

36. A process according to any of Claims 23 to 35, in which the foods and drinks include feeds and pet foods for domestic animals and fowls, honey bees, silkworms, and fishes, favorites, cosmetics, drugs and medicines.

37. A process according to any of Claims 23 to 36, in which the foods and drinks are tobaccos, tooth pastes, lipsticks, lipcreams, medicines for internal administration, troches, drops containing liver oil, refrigerants, cachous and gargles.

38. A process according to any of Claims 23 to 37, in which the foods and drinks are lowcariogenic and/or anti-caries.