JPH0341085A - Novel sugar alcohol, its production and use - Google Patents

Abstract

NEW MATERIAL:The compound of formula (Gal is galactose residue; Sor is sorbitol residue; (n) is 1-4). EXAMPLE:O-beta-D-galactopyranosyl-(1 6)-O-beta-D-galactopyranosyl-(1 4)-D-sorbitol. USE:A constipation ameliorating agent. PREPARATION:The objective compound can be produced by reacting lactose or lactose-containing material with beta-galactosidase or beta-glucosidase to effect the rearrangement reaction of the galactose residue in the lactose molecule and reducing the glucose residue of the resultant oligosaccharide.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a novel sugar alcohol, a method for producing the same, and further its use.

This sugar alcohol has an excellent proliferative effect on Bifidobacterium in the body, and when taken in combination with dietary fiber, lactose, and lactitol, it shows a remarkable effect on improving constipation. It is expected to be useful as a bioactive agent.

Many studies have been conducted on substances that promote the growth of bifidobacteria (hereinafter referred to as bifidobacterial growth factors), and oligosaccharides have attracted attention as growth factors. Such an oligosaccharide is obtained by reacting β-galactosidase produced by Aspergillus oryzae with lactose or a lactose-containing substance, in the form % formula % (where Gal represents a galactose residue, Glc
indicates a glucose residue, and n indicates an integer from 1 to 4) has been proposed to be used as a Bifidobacterium growth factor [JP-A-55104885].

The present inventor has demonstrated that by reducing glucose residues in oligosaccharides obtained by treating lactose or lactose-containing substances with β-galactosidase or β-glucosidase, and by treating lactitol or lactitol-containing substances with β-galactosidase or β-glucosidase. - Obtained the knowledge that a new sugar alcohol can be obtained by the action of glucosidase, and that this sugar alcohol has a proliferating effect on bifidobacteria and a remarkable effect on improving constipation when taken in combination with lactose and dietary fiber. The present invention has been accomplished.

Therefore, the present invention provides a novel sugar alcohol exhibiting the above-mentioned physiological effects, a method for producing the sugar alcohol,
Furthermore, it is an object of the present invention to provide a bifidobacteria growth-promoting composition and a constipation-improving composition containing this sugar alcohol as an active ingredient.

Did you decide on the assignment? )? The sugar alcohol according to the present invention has the following general formula (I)% formula% () (However, in the formula, Gal represents a galactose residue, Sor
each represents a sorbitol residue, and n represents an integer of 1 to 4.) The sugar alcohol represented by the above general formula (I) is represented by the following formula (
It includes each sugar alcohol represented by 1) to (3).

Formula (+) 0-β-D-galactopyranosyl-(1→6)-〇-β
-D-galactopyranosyl-(1→4)-〇-sorbitol (1) Formula (2) 0-β-D-galactopyranosyl-(1→3)-〇-β
-D-galactopyranosyl-<1→4)-〇-sorbitol (2) Formula (3) 0-β-D-galactopyranosyl-(1→4)-[0-
β-D-galactopyranosyl-(1→6))-D-sorbitol (3) The physical properties and constituent sugars and sugar alcohols of these sugar alcohols are as follows.

(a) Molecular weight It has a molecular weight of 506 as measured by a mass spectrometer.

(b) Color When dried and powdered, all exhibit a white color. (c) Distinction between acidic, basic and neutral, all indicate neutrality.

(d) Solubility All of them are soluble in water, but they are soluble in benzene, chloroform, and acetone.

(e) Color reaction Silver nitrate reaction Anthrone sulfuric acid reaction Juninhydrin reaction Biulet reaction (f) tR constituent sugar sugar alcohol The sugar alcohol according to the present invention was hydrolyzed with 0.5N-11CI at a temperature of too'c for 4 hours. Since the molar ratio of the resulting sugar and sugar alcohol was galactose:sorbitol=2:1, it was confirmed that the sugar alcohol according to the present invention consisted of two molecules of galactose and one molecule of sorbitol.

(g) Bonding mode of constituent sugars and sugar alcohols The sugar alcohols according to the present invention were each subjected to high performance liquid chromatography (
After separation and purification by HPLC), methylation analysis was performed using a conventional method, and the binding mode shown in Table 1 was confirmed.

Table (H) The sugar alcohol of the present invention was confirmed to be β-coordinated since galactose and sorbitol were produced by the action of β-galactosidase.

In view of the properties shown in (a) and (f) to (h) above, the sugar alcohols according to the present invention can be identified as those represented by the above formulas (1) to (3), respectively.

Next, the method for producing sugar alcohol according to the present invention will be explained.

The sugar alcohol according to the invention has β
- Obtained by reacting galactose residues in lactose molecules with galac-1-sidase or β-glucosidase to generate oligosaccharides, and reducing the glucose residues of the resulting oligosaccharides. . Moreover, this sugar alcohol can also be obtained by allowing β-galactosidase or β-glucosidase to act on lactitol or a lactitol-containing substance.

The origin of galactosidase or glucosidase used here does not need to be particularly limited, and it does not need to be highly purified, and can be used in the form of a crude enzyme.

In order to cause the above enzyme to act on the starting material lactose or a lactose-containing substance, use the starting material whose lactose concentration has been adjusted to 5 to 50% as a substrate, and add the enzyme concentration to 0.5% with pif 2 to 8.
It is appropriate to allow the enzyme to act at a temperature of 10 to 60'C at a concentration of 1 to 280 units/TR1.

The enzymatic reaction causes a galactose transfer reaction to occur in the starting material, producing an oligosaccharide mixture.

Next, the reaction solution containing the oligosaccharide mixture is heated at a temperature of 90' or higher for 30 seconds to 3 minutes to inactivate the enzyme, and then
The oligosaccharide in the reaction solution is adsorbed by contacting with an adsorbent containing activated carbon (usually passed through a column packed with the adsorbent).

Next, the adsorbed oligosaccharides are eluted using a 15-50% ethanol solution to obtain an eluate containing oligosaccharides.

The obtained eluate is concentrated under reduced pressure and further dried if necessary.

The thus obtained galacto-oligo mass is treated with sodium borohydride in an aqueous or alcoholic solution to reduce its glucose residue and convert it into sorbitol. At this time, the concentration of galacto-oligo is 1 to 50% (W
/V), it is preferable to adjust the concentration of sodium borohydride used in the reduction reaction to 0.1 to 5M. In addition, the above reduction reaction is carried out at a temperature of 0 to 50°C for 30 minutes to
It is appropriate to do this for 3 hours.

After the reaction is completed, excess sodium borohydride in the reaction solution is decomposed with acetic acid or acetone, etc., and then the reaction solution is desalted using a desalting resin, electrodialysis, ultrafiltration, etc., and the resulting tank is The alcohol is concentrated under reduced pressure and, if necessary, dried by freeze-drying or the like to obtain a powdered target sugar alcohol.

Furthermore, in the present invention, the target sugar alcohol can also be produced by reducing glucose residues of oligosaccharides by the following method.

The oligosaccharide-containing eluate obtained in the above manufacturing process was heated under reduced pressure.
The a-condensed product is reacted with hydrogen at high temperature and high pressure using a nickel catalyst such as Raney nickel, thereby reducing glucose residues and converting them into sorbitol.

At this time, the concentration of galactooligosaccharide is 40-70% (n/
v), the amount of nickel used in the catalyst is 0.02 to 5% (w
/v). In addition, the above reduction reaction is carried out at a temperature of 100 to 200"C (7) and a hydrogen pressure of 5
It is appropriate to carry out the treatment for 20 to 5 hours at 0 to 17 hours.

After the reaction is completed, impurities in the reaction solution can be removed by passing the solution through activated carbon, ion exchange resin, or the like.

The obtained aphrodisiac alcohol is concentrated under reduced pressure and, if necessary, dried by freeze-drying or the like to obtain a powdered target sugar alcohol.

Furthermore, in the present invention, the target sugar alcohol can also be produced by the following method.

The target sugar alcohol (galactosyl lactitol) is obtained by using lactitol or a lactitol-containing material as a starting material and allowing β-galactosidase or β-glucosidase to act on it to cause a transfer reaction to the galactose residues of lactitol.

β-galactosidase or β-glucosidase used here does not need to be particularly limited in its origin, and does not need to be highly purified, as in the case where lactose or its containing substance is used as a starting material, as described above. The crude enzyme may also be used.

To cause the above enzyme to act on the starting material lactitol or β-lactitol-containing substance, use the starting material adjusted to a lactitol concentration of 5 to 75% as a substrate, and add all 2 to 8 to an enzyme concentration of 0.1 to 75%. 280 units/R1] It is appropriate to allow the enzyme to act at a temperature of 0 to 60°C.

By allowing the above enzyme to act, a galactose transfer reaction occurs on lactitol and a target alcohol mixture is produced, so the reaction mixture containing the sugar alcohol is
After inactivating the enzyme by heating at a temperature of 0°C or higher for 30 seconds to 3 minutes, the enzyme is brought into contact with an adsorbent containing activated carbon (usually,
The adsorbent is passed through a packed column to adsorb the sugar alcohol in the reaction mixture.

Next, the adsorbed target alcohol is eluted using an ethanol solution with a concentration of 15 to 50%, the obtained eluate is concentrated under reduced pressure, and if necessary, dried by freeze-drying etc. to obtain a powdered target sugar alcohol. obtain.

As mentioned above, the sugar alcohol obtained as above has an excellent proliferative effect on Bifidobacterium in the living body, and also has a remarkable constipation-improving effect when taken in combination with dietary fiber, lactose, etc. Therefore, it can be used as an active ingredient of a bifidobacteria growth promoter and a constipation improving agent.

Next, the results of testing the growth promoting effect on bifidobacteria and the constipation improving effect of the sugar alcohol according to the present invention will be shown.

■Proliferation test test method for Bifidobacterium: Cynomolgus monkeys (groups of 10 animals) were used as test animals, and after 5 weeks of feeding commercially available powdered milk for infants with 5% by weight of lactose added to each group, sugar alcohol according to the present invention was added to each group of cynomolgus monkeys. Powdered milk for infants containing 5% by weight of (powdered form) was given for an additional 3 weeks, during which time the feces of each monkey was collected and the proportion of bifidobacteria in the feces was measured. The results are shown in the attached drawings.

As seen in the figure, the proportion of bifidobacteria in feces during the period when the sugar alcohol of the present invention was added and fed was significantly higher than that during the period when lactose was added and fed. It is rising.

In other words, the Tang alcohol according to the present invention has an extremely excellent effect on promoting the growth of Bifidobacteria in vivo, and is also effective against various Bifidobacteria that grow in the human intestinal tract, such as Bifidobacterium longum and Bifidobacterium longum. Fydobacterium breed, Bifidobacterium
It is recognized that it exhibits high growth activity against a wide range of types of bifidobacteria, such as Bifidum, Bifidobacterium adressentis, and Bifidobacterium infantis.

Therefore, the sugar alcohol according to the present invention can be blended into dairy products such as powdered milk and fermented milk, or added as a component of drugs such as intestinal preparations.

Incidentally, when a milk drink containing 2% by weight of sugar alcohol according to the present invention was prepared and administered to 30 healthy adults, the increase in the ratio of bifidobacteria to the total number of bacteria in their feces was due to lactose. It was observed that the amount was approximately twice as high as when a milk drink containing 2% by weight was similarly administered.

■Constipation improvement test test method: The sugar alcohol powder obtained above was divided into 5 groups of 4 people each to 20 healthy elderly people who complain of constipation on a daily basis, and sugar alcohol powder with the following combination was used as test groups Nal to Na5. were administered orally.

Cutting plate k Sugar alcohol dosage (g) kl
5 Na2 4 Hiroshi 33 Hiroshi 42 Hyo 50 In each test group, the above amount of sugar alcohol was added to 1
Administer once a day for 2 weeks, 2i before administration! The number of defecations during the two-week period during which the rats were administered was sensory evaluated for the hardness of the feces at each defecation. The results are shown in Table 2.

table 2 note) The number of defecation times is the average for 2 weeks per person.

The hardness of feces is expressed as the average of each defecation per person, and is expressed in the following 5 levels.

Very soft 5 points Soft 4 points Average 3 points Hard 2 points Very hard 1 point As shown in the table, the group administered the sugar alcohol of the present invention showed a significant increase in the frequency of defecation when 3 g or more was administered. Softening of feces was observed. In the control group, no increase in the frequency of defecation or softening of feces was observed in the prisoners.

As mentioned above, the single administration of the sugar alcohol according to the present invention has a remarkable effect of improving constipation.

This effect is due to the large amount of β-gauctosidase present in human small intestinal microvilli, so the sugar alcohol of the present invention reaches the large intestine without being decomposed in the small intestine, and as a result, the osmotic pressure in the large intestine increases. It is presumed that this increases in concentration and softens feces to improve constipation.

As mentioned above, the sugar alcohol according to the present invention exhibits an excellent growth-promoting effect on Bifidobacterium, and furthermore,
It also shows excellent constipation-improving effects, increasing the frequency of defecation and softening feces.

The present invention will be specifically described below with reference to Examples.

Example 1 Preparation of sugar alcohol: 10 kg of lactose (commercially available) was dissolved in 15 kg of warm water. Add citric acid to the 8 solution to adjust the pH to 4.5, add 50,000 units of β-galactosidase, and heat at 40°C.
The mixture was allowed to react for 10 hours. The resulting reaction mixture was heated to 105°C.
The enzyme was inactivated by heating for 2 seconds.

Next, 2 kg of this liquid was passed through an activated carbon-Celite (2:1) column (φ50 cm x 50 c11) to adsorb some of the lactose and transferred galacto-oligosaccharides. Then 1
5 kg of water was passed through the column to elute and remove monosaccharides and unadsorbed lactose, and then 20 kg of 5% ethanol was passed through the column to completely elute and remove the adsorbed lactose. Next, 40%
The eluate obtained by passing 10 kg of ethanol through the column was concentrated under reduced pressure and then lyophilized to obtain galacto-oligo tJ! 95
150 g of powder containing more than % by weight was obtained.

Next, sodium borohydride aqueous solution (10% W/V)
Galactooligosaccharide powder 1 obtained by the above method in 3e
50 g was dissolved and a reduction reaction was performed at room temperature for 3 hours. After the reaction was completed, the mixture was neutralized using acetic acid until the pH reached 7.0. This makes it possible to decompose unreacted sodium borohydride present in excess in the reaction solution.

Next, this material was distilled off under reduced pressure and added to methanol in step 31 again!
l! It became cloudy and was distilled off again under reduced pressure. After repeating this operation twice, the solution dissolved in water 11 was passed through a desalting resin to perform desalting. The resulting solution was reduced in size under reduced pressure and freeze-dried to obtain 150 g of sugar alcohol powder containing the sugar alcohols represented by formulas (1), (2), and (3).

Example 2 150 g of a powder containing at least 95% by weight of galacto-oligosaccharides was obtained according to the same procedure as described in Example 1, the powder was then dissolved to a concentration of 50% (n/v), and Raney nickel 1 [L% and increase the temperature to 90-1 while stirring.
After raising the temperature to 25°C and raising the hydrogen pressure to 20 to 100 kg/cJ to complete hydrogenation, Raney nickel was removed, and the purified product was concentrated under reduced pressure using activated carbon and ion exchange resin according to a conventional method. , further freeze-dried to obtain the formula (1),
150 g of sugar alcohol powder containing sugar alcohols represented by (2) and (3) was obtained.

Example 3 β-
53 units of galactosidase were added and reacted at 40°C for 10 hours.

The resulting reaction mixture was heated at 105°C for 2 seconds to inactivate the enzyme.

Next, 2 kg of this liquid was passed through an activated carbon-Celite (2:1) column (φ50 cm x 50 c+w) to adsorb lactitol and transferred sugar alcohol.
Kg of water was passed through the column to elute and remove monosaccharides and unadsorbed lactitol, and then 5% ethanol 20
kg was passed through the column to completely elute and remove the adsorbed lactitol. Next, the eluate obtained by passing 10 kg of 40% ethanol through the column was concentrated once under reduced pressure, and then lyophilized to obtain 150 g of powder containing sugar alcohols represented by the above formulas (1), (2), and (3). Obtained.

[Brief explanation of drawings]

The attached figure shows the growth promoting effect of the sugar alcohol of the present invention on bifidobacteria in comparison with lactose.

Claims (10)

[Claims] (1) General formula (I) Gal-(Gal)_nSor(I) (wherein, Gal represents a galactose residue, Sor represents a sorbitol residue, and n represents an integer from 1 to 4). sugar alcohol. (2) Formula O-β-D-galactopyranosyl-(1→6)-O-β
The sugar alcohol according to claim (1), which is represented by -D-galactopyranosyl-(1→4)-D-sorbitol. (3) Formula O-β-D-galactopyranosyl-(1→3)-O-β
The sugar alcohol according to claim (1), which is represented by -D-galactopyranosyl-(1→4)-D-sorbitol. (4) Formula O-β-D-galactopyranosyl-(1→4)-[O-
The sugar alcohol according to claim 1, which is represented by β-D-galactopyranosyl-(1→6)]-D-sorbitol. (5) Oligosaccharides are produced by allowing β-galactosidase or β-glucosidase to act on lactose or lactose-containing substances to perform a transfer reaction on galactose residues in lactose molecules, and then the glucose residues of the obtained oligosaccharides are A method for producing a sugar alcohol, characterized by reducing a group. (6) The method for producing a sugar alcohol according to claim (5), wherein the reduction of the glucose residue of the oligosaccharide is carried out by reacting with sodium borohydride in an aqueous solution or an ethanol solution. (7) The method for producing a sugar alcohol according to claim (5), wherein the reduction of the glucose residue of the oligosaccharide is carried out by reacting with hydrogen at high temperature and high pressure using a nickel catalyst. (8) A method for producing a sugar alcohol, which comprises allowing β-galactosidase or β-glucosidase to act on lactitol or a lactitol-containing substance to cause a transfer reaction to galactose residues in lactitol molecules. (9) General formula (I) Gal-(Gal)_n-Sor(I) (However, in the formula, Gal represents a galactose residue, Sor represents a sorbitol residue, and n represents an integer from 1 to 4) A composition for promoting the growth of Bifidobacterium, which contains a sugar alcohol represented by the following as an active ingredient. (10) Contains a sugar alcohol represented by the general formula (I) Gal-(Gal)_n-Sor(I) (in the formula, Gal represents a galactose residue and Sor represents a sorbitol residue, respectively) as an active ingredient. A constipation-improving composition.