JPH07100710B2 - Method for epimerizing reducing oligosaccharides - Google Patents

Description

TECHNICAL FIELD The present invention relates to a method for epimerizing a reducing oligosaccharide.

[Conventional technology]

Oligosaccharides are saccharides formed by dehydration condensation of several monosaccharides found in a free state or bound to other biological substances in a living body through glycoside bonds.

In recent years, various sugars have been found that show activities such as antibacterial activity and carcinostatic activity, or sugars with other useful applications have been discovered. There is increasing interest as a substance or as a nutritional substance or low calorie sweetener.

Under these circumstances, as research on oligosaccharides and application development have been activated, and as these demands have been cultivated, the development of industrial acquisition methods for a wide variety of oligosaccharides, especially naturally rare oligosaccharides, has been developed. Has become necessary.

However, as a method for converting oligosaccharides that are relatively abundant in nature from the past into rare oligosaccharides, monosaccharides have long been the so-called epilation reaction via 1,2-enediol in weakly alkaline aqueous solutions. It is known, but when oligosaccharides were used as targets, there were almost no reports of the reaction, and Bilik's report [LISTY CUKROVARNICKE
94/1978 P88], the present situation is that only examples of epimerizing D-merbiose using molybdic acid are introduced. Therefore, it can be said that the oligosaccharide epimerization technology belongs to a new technical field to be developed.

In addition, the above-mentioned method of Bilik has a problem that the reaction temperature is as high as 90 ° C., the reaction time is too long as 6 hours, and the epilation rate is only 20%, which is not an industrial method. Met.

[Means for solving problems]

In view of such circumstances, the present inventor has earnestly studied an oligosaccharide epimerization method in which the reaction conditions are mild, and the epimerization rate and the sugar recovery rate are high. The inventors have found that the object can be achieved by using a catalyst, and have reached the present invention.

That is, the present invention is a salt of a metal selected from Ni, Co, Ca, Sr, La, Ce, Pr, Nd, Y, In and N, N, N'-trimethylethylenediamine or N, N, N ', N'. -A method for epimerizing a reducing oligosaccharide, which comprises adding a reducing oligosaccharide to a solution containing a complex formed from tetramethylethylenediamine and / or both and allowing the mixture to react with heating.

The epimerization of the present invention mainly means the epimerization of C-2 of the reducing terminal aldose residue of the oligosaccharide.

The present invention will be described in detail below.

The starting oligosaccharide used in the present invention is preferably a reducing oligosaccharide.

Here, the reducing oligosaccharide refers to one obtained by dehydration condensation between the hemiacetal hydroxyl group of one sugar and the alcoholic hydroxyl group of the other sugar.

As a specific example, D-melibiose [α-D-Galp
-(1 → 6) -D-Glcp], D-gentiobiose [β
-D-Glcp- (1 → 6) -D-Glcp], an isomaltooligosaccharide such as D-isomaltose [α-D-Glcp- (1
→ 6) -D-Glcp], Isomaltotriose [α-D-
Glcp- (1 → 6) -α-D-Glcp- (1 → 6) -α-D
-Glcp], isomalt hexaose [(α-D-Glcp-
(1- [→ 6) -α-D-Glcp- (1-] ₄ → 6) -α
-D-Glcp], and D-maltose [α-D-Glcp-
(1 → 4) -D-Glcp], D-laminaribiose [β-
D-Glcp- (1 → 3) -D-Glcp] and the like.

These oligosaccharides are C-2 of the aldose residue at the reducing end.
Is converted into an epimer and converted into each epimer.

Metal salts used in the present invention, Ni, Co, Ca, Sr, La, Ce, Pr, Nd, Y,
A salt of a metal selected from In, a halide thereof is preferable, and a chloride is particularly preferable. These metal salts form a complex with a specific diamine described later and serve as a catalyst for the epilation reaction.

Further, in the present invention, the metal salt and the diamine, which are the raw materials of the complex, may be added and the reducing oligosaccharide may be directly added thereto (the present inventor named the latter method one pot method).

In the present invention, the choice of diamine is particularly important,
N, N, N'-trimethylethylenediamine (hereinafter abbreviated as tmen) or N, N, N ', N'-tetramethylethylenediamine (hereinafter abbreviated as tetmen) is used.

It has been confirmed by the study of the present inventor that the length of the methylene chain of this diamine has an important influence on its action function.

In the one pot method, the amount ratio of the metal salt and the diamine can be appropriately selected according to other conditions, but is usually 1: 0.2 to 1:10, preferably 1: 0.5 to 1: 4. Further, the ratio of the raw sugar and the metal salt is 1: 0.2 to 1: 5, preferably 1: 0.5 to 1: 2. Next, when a complex is used, the amount ratio of the raw sugar and the complex is 1: 0.2.
˜1: 10, preferably 1: 0.5 to 1: 5.

The above-mentioned complex or metal salt and diamine dissolve them,
It is preferred that the reducing oligosaccharide is also dissolved in a solvent that can be used before use. As this solvent, lower alcohols such as methanol and ethanol, mixed solvents of water and lower alcohols, mixed solvents of DMF and lower alcohols, etc. are used, and methanol is particularly excellent.

The reaction temperature is set in the range of room temperature to 150 ° C, but especially 4
0 to 100 ° C is preferable. If the temperature is too high, side reactions occur and the yield decreases. On the other hand, if it is too low, the reaction rate is too slow, which is not preferable. The reaction pressure is not particularly limited and may be normal pressure.

The reaction time is one of the features of the method of the present invention, but it may be an extremely short time within 1 hour as compared with the conventional method. Usually 5-30 minutes is sufficient.

In addition, although better results can be obtained by stirring during the reaction, this is not essential.

As described above, in the method of the present invention, the above metal salt and diamine or a complex thereof is added to a solvent such as methanol, a reducing oligosaccharide is added, and the mixture is heated for 5 to 30 minutes under stirring to form an epilation. Allow the reaction to take place.

In order to take out the epimer after the epilation reaction, it is treated with a weakly acidic aqueous solution and then separated.

Water may be added to the reaction solution as needed, and an acid such as dilute hydrochloric acid or dilute sulfuric acid may be added to the reaction solution under weak acidity (usually around pH 6.5). Preferably, the reaction is carried out at room temperature or higher for several minutes or longer with stirring.

Although various known methods can be used for the separation operation, using a cation exchange resin and an anion exchange resin,
A method of removing the metal ion, complex ion, halogen ion and the like and then taking out the desired epimer by concentration or the like is simple and preferable.

〔Example〕

 The present invention will be described below with reference to examples.

In addition, high-performance liquid chromatography (hereinafter referred to as HP
LC), but specifically, a post-column detection method using 2-cyanoacetamide as a color reagent (for details, see
S.Honda et al. Anal. Chem. 52.1079 (1982)) using a fluorescence detector.

Example 1 NiCl ₂ .6H ₂ O 132 mg and tetmen 129 mg were added to methanol 30 ml.
Dissolved in. Next, 200 mg of D-melibiose was dissolved in this, and then reacted at 60 ° C. under normal pressure for 10 minutes.

After the reaction, the mixture was immediately cooled, 30 ml of water was added, 2N HCl was added, and the mixture was stirred at room temperature for 30 minutes.

The reaction solution was deionized using a cation exchange resin Dowex 50W-X2 and an anion exchange resin DoWex MSA-1.

As a result of analyzing the sugar in the obtained liquid by HPLC, 48% of the initially added D-melibiose was converted to an epimer. The D-melibiose: epimer ratio was 44:56. Further, sugars other than D-melibiose and its epimer were not detected at all. The sugar recovery rate (yield) relative to the raw sugar was as high as 85%.

From these results, it can be seen that the present invention is an excellent method with high epilation rate, high selectivity and high recovery rate.

Next, Table 1 shows the results of the same procedure as above except that the reaction temperature was changed to 40 ° C, 80 ° C and 100 ° C.

Examples 2 to 19 The catalyst system was a combination of various metal salts and diamines as shown in Table 2 (the amount ratio in this case was D-melibiose / metal salt / diamine = 1/1/2 (molar ratio)). Or the complex (in this case, the amount ratio was D-melibiose / complex = 1/1), and the same procedure as in Example 1 was carried out. The results are shown in Table-2.

Using Examples 20-22 _{[Ni (H 2 O) 2 (} tmen) 2 ] Cl ₂ as a catalyst, the table and the solvent -
Example 1 was carried out in the same manner as in Example 1 except that it was changed as shown in FIG.

The results are shown in Table-3.

Examples 23 to 32 Solvent methanol, reaction temperature of 60 ° C., reaction time of 10 minutes, etc. were the same as in Example 1 except that the reaction conditions (starting sugar and catalyst) shown in Table 4 were the same as those of Examples 23 to 32. Carried out. The results are shown in Table-4.

[Effects of the Invention] The present invention allows the reaction to proceed under mild conditions that have never been achieved before.
Moreover, it is a method of epilating a reducing oligosaccharide having a high epilation rate and excellent selectivity.

Thus, according to the present invention, it has become possible to obtain valuable epimers of oligosaccharides, which are considered to be more important in the future, in large quantities with high quality.

Claims (5)

[Claims] 1. A salt of a metal selected from Ni, Co, Ca, Sr, La, Ce, Pr, Nd, Y, In and N, N, N'-trimethylethylenediamine or N, N, N ', N. A method for epimerizing a reducing oligosaccharide, which comprises adding a reducing oligosaccharide to a solution containing a complex formed of'-tetramethylethylenediamine and / or both, and allowing the mixture to react with heating. 2. The method for epimerizing a reducing oligosaccharide according to claim 1, wherein the metal salt is a halide. 3. The method for epimerizing a reducing oligosaccharide according to claim 1, wherein the solution is a lower alcohol or a mixed solution of lower alcohol and water or DMF. 4. The method for epimerizing a reducing oligosaccharide according to claim 1, wherein the heating reaction is carried out at 40 to 100 ° C. 5. A raw material reducing oligosaccharide is D-melibiose,
The method for epimerizing a reducing oligosaccharide according to claim 1, which is selected from D-gentiobiose, isomaltooligosaccharide, D-maltose, and D-laminaribiose.