JP4599505B2 - Method for producing N-acetyl-β-hexosaminide derivative - Google Patents

Description

[0001]
[Technical field to which the invention belongs]
The present invention relates to a method for specifically producing an oligosaccharide having an N-acetyl-β-hexosaminide derivative, particularly 2-acetamido-2-deoxy-β-D-hexopyranoside. An N-acetylhexosamine oxazoline and a glycosyl acceptor are reacted to produce an N-acetyl-β-hexosaminide derivative, which is characterized by using an acid catalyst, particularly an ion exchange resin that can be recycled.
[0002]
[Prior art]
In general, the glycosidic bond forming reaction for synthesizing oligosaccharides is performed by activating the C-1 position of the glycosyl donor and simultaneously reacting with the hydroxyl group of the acceptor. However, many methods have the disadvantage that only low yields and poor stereoselectivity can be obtained while using expensive, highly toxic heavy metals or acid catalysts that are difficult to reuse as activators. Therefore, synthesis on an industrial scale was difficult (K. Toshima, Chem. Rev., 93, 1503 (1993)). In particular, in the synthesis of an N-acetyl-β-hexosaminide derivative, when an N-acetylhexosamine derivative having an active group at the 1-position is used as a sugar donor, an acetamide group at the C-2 position and a C- It is known that an oxazoline ring is formed between the 1-position, the yield is lowered, and it does not react with a hydroxyl group of a sugar having low reactivity. In the reaction to synthesize N-acetyl-β-hexosaminide derivatives to solve this problem, N-acetylhexosamine donors in which the acetamido group at the C-2 position is substituted with other substituents such as phthalimide group or azide group (J. Banoub, Chem. Rev., 92, 1167 (1992)). However, the synthesis of these derivatives requires a multi-step synthesis process. In order to perform an efficient synthesis on a large scale, it is important that the starting material is easily prepared.
[0003]
Attempts have been made to positively utilize the oxazoline derivative produced as a by-product during the glycosidic bond formation reaction in the glycosylation reaction. However, the method of Kiso et al. (Carbohydr. Res., 136, 309 (1985)) uses highly reactive metals and is not suitable for large-scale industrial production. In addition, the method of RL Thomas et al. (Carbohydr. Res., 183, 163 (1988)) uses an acid catalyst that is difficult to reuse, which is expensive in industrial production. In these reactions, when alcohols are used as receptors, glycosidic bonds are given, but when sugars are used as receptors, it is possible to obtain glycosidic bonds only with limited sugar receptors that are strictly protected. Known and lacks universality. Furthermore, these reactions are not suitable for mass synthesis because it is necessary to introduce a protecting group to a sugar hydroxyl group that is not involved in the opposite, and many steps are required.
[0004]
An example of a glycoside formation reaction using oxazoline of N-acetylhexosamine as an enzyme substrate has been reported (S. Kobayashi, J. Am. Chem. Soc., 118, 13113 (1996)). In this case, oxazoline and sugar receptor protecting groups are not necessary and are a practical method. However, the sugar receptor in this reaction is strictly regulated by the substrate specificity of the enzyme, and cannot be said to be a method that can be applied to all saccharides, alcohols and the like.
[0005]
[Problems to be solved by the invention]
Therefore, the object of the present invention is to provide an easy N-acetyl-β-type for many glycoside receptors such as sugars, alcohols, phenols, etc., with good yield and high β-selectivity and corresponding to large-scale preparation. It is to provide a process for producing hexosaminide derivatives, namely 2-acetamido-2-deoxy-β-D-glucopyranoside derivatives and 2-acetamido-2-deoxy-β-D-galactopyranoside derivatives.
[0006]
[Means for Solving the Problems]
As a result of various studies to solve the above-mentioned problems, the present inventors have determined that N-acetylhexosamine oxazoline compound having no protecting group is a donor, a cation exchange resin is used as a catalyst, and N--to saccharides, alcohols and phenols. The present invention was completed by finding that an acetyl-β-hexosaminide derivative can be synthesized. The present invention will be described in detail below. That is, the present invention is [1] catalyzed by a cation exchange resin, an N-acetylhexosamine oxazoline compound represented by the general formula (1) is used as a sugar donor, and sugars, alcohols, or phenols are sugar-accepting. manufacturing method of a body N- acetyl -β1- Hekisosaminido derivatives, [2] N- acetyl -β1- Hekisosaminido derivative is 2-acetamido-2-deoxy-beta1-D-hexopyranoside derivative, to the [1] the method of manufacturing described, [3] N-acetyl -β1- Hekisosaminido derivative, 2-acetamido-2-deoxy-beta1-D-glucopyranoside derivative and 2-acetamido-2-deoxy-beta1-D-galactopyranoside derivative The production method according to [1] above, wherein the [4] N-acetyl-β1-hexosaminide derivative is GlcNAcβ1-6Man, GlcNAcβ1-3Man, GlcNAcβ1-6Galβ-OpNp, GlcNAcβ1-OMe and GalNAcβ1-OMe, Product described in [1] above And [5] the production method according to any one of [1] to [4], wherein the solvent is an aprotic polar solvent .
[0007]
DETAILED DESCRIPTION OF THE INVENTION
The synthesis of oligosaccharides having N-acetyl-β-hexosaminide derivatives is described. An oxazoline compound and a sugar acceptor are dissolved in an aprotic polar solvent. The aprotic solvent refers to an organic solvent such as dimethyl sulfoxide, dimethylformamide, hexamethylphosphoramide, triethyl phosphite. The amount ratio between the oxazoline compound and the sugar receptor is 1/100 to 100 equivalents, but increasing the concentration of the sugar receptor improves the reaction efficiency. Catalysts for activating the oxazoline ring to initiate the glycoside formation reaction include organic acids such as para-toluenesulfonic acid and camphorsulfonic acid, mineral acids such as hydrochloric acid and sulfuric acid, and cation exchange resins. Although there is no problem, an ion exchange resin that can be regenerated is effective from the viewpoint of operability and cost.
Examples of the ion exchange resin include strongly acidic and weakly acidic cation exchange resins such as Amberlist 15, Amberlist 15E, Amberlist 15WET, Amberlist 16WET, Amberlist 31, Amberlist 35WET, Amberlist A26, and Amberlist A21. Can be used.
[0008]
The synthesis of N-acetyl-β-hexosaminide derivatives, in which alcohol is β-glycosidically bonded to N-acetylhexosamine, proceeds almost quantitatively by dissolving in the alcohol that binds the oxazoline compound and then performing a glycoside formation reaction with an ion exchange resin. To do. When a high melting point alcohol is used as a substrate, it is dissolved in an aprotic polar solvent as in the oligosaccharide synthesis. The amount ratio of the oxazoline compound to the alcohol acceptor is 100 equivalents from the amount of catalyst, but the reaction efficiency is improved by increasing the concentration of the alcohol acceptor.
Examples of the present invention will be described below in more detail, but the present invention is not limited to these examples.
[0009]
【Example】
Synthesis example Synthesis of N-acetylcurcosamine oxazoline
By reacting a peracetylated derivative obtained by reacting N-acetylglucosamine with acetic anhydride and pyridine in the presence of a Lewis acid, an acetyl derivative (3) of oxazoline was obtained (Stork, W. et al. , J. Carbohydr. Chem. 2, 169-198 (1989)).
113 mg of the obtained oxazoline acetyl derivative (3) 113 mg was dissolved in methanol, 100 μL of 1N NaOMe was added, and the mixture was stirred at room temperature for 3 hours. The residue obtained after removing methanol under reduced pressure was purified by silica gel column chromatography (chloroform: methanol = 4: 1, 0.1% triethylamine) to obtain 69 mg of oxazoline (4) (quantitative). (Fig. 1)
Example 1: Synthesis of GlcNAcβ1-6Man and GlcNAcβ1-3Man Mannose 337 mg (1.87 mmol) and oxazoline (4) 38 mg (0.18 mmol) synthesized in the synthesis example were dissolved in 0.6 mL of anhydrous dimethyl sulfoxide, and 300 mg of Amberlyst 15E (Organo) was added and stirred at room temperature for 12 hours. After the ion exchange resin was filtered off, the filtrate was diluted with 1 L of water, applied to an activated carbon column (2 cm × 20 cm), and eluted with a 10% aqueous ethanol solution to obtain 20 mg of the desired disaccharide mixture. GlcNAcβ1-6Man 9 mg (0.023 mmol) and GlcNAcβ1-3Man 4 mg (0.010 mmol) were obtained by purification with HPLC (TSK, Amide-80 column, 21.5φx30 cm, 75% acetonitrile, 80 ° C., 7 ml / min). (Figure 2)
[0010]
Example 2: Synthesis of GlcNAcβ1-6Galβ-OpNp Paranitrophenyl-β-D-galactopyranoside (Galβ-O-pNp) 124 mg (0.41 mmol) and oxazoline (4) 42 mg (0.21 mmol) in 1 mL anhydrous It was dissolved in dimethylformamide, 100 mg of Amberlyst 15E was added thereto, and the mixture was stirred at room temperature for 1 hour. After the ion exchange resin was filtered off, the filtrate was removed under reduced pressure. The obtained residue was purified using HPLC (ODS column 10% acetonitrile) to obtain 2.5 mg (0.024 mmol) of the desired GlcNAcβ1-6 Galβ-OpNp. (Figure 3)
[0011]
Example 3 Synthesis of GlcNAcβ1-OMe Oxazoline (4) 782 mg (3.85 mmol) was dissolved in 8 mL of methyl alcohol, 300 mg of Amberlyst 15E was added thereto, and the mixture was stirred at room temperature for 1 hour. After the ion exchange resin was filtered off, the filtrate was removed under reduced pressure. The obtained residue was recrystallized from isopropyl alcohol / isopropyl ether to obtain 320 mg (1.36 mmol) of GlcNAcβ-OMe. (Fig. 4)
[0012]
Example 4: Synthesis of GalNAcβ1-OMe
Oxazoline (5) 500 mg (3.85 mmol) prepared using N-acetylgalactosamine as a raw material in the same manner as in the synthesis example is dissolved in 30 mL of anhydrous methyl alcohol, and sodium hydroxide solution (1 M, 200 μl) is added at room temperature. Deacetylation was carried out by stirring for 10 minutes. Subsequently, Amberlyst 15E was added until neutral and stirred at room temperature for 1 hour. After the ion exchange resin was filtered off, the filtrate was concentrated under reduced pressure to obtain the product (298 mg). ¹ H NMR of the obtained compound was consistent with that of GalNAcβ1-OMe synthesized separately. (Fig. 5)
[0013]
【The invention's effect】
The compound obtained by the method of the present invention is an N-acetyl-β-hexosaminide derivative such as an oligosaccharide constituting a glycoprotein sugar chain or paranitrophenyl-2-acetamido-2-deoxy-β-D-glycosaminide, It is useful as a probe such as an antibody, as a carrier for DDS, as a substrate for measuring enzyme activity, as a pharmaceutical, a synthetic raw material, a research reagent, or the like.
[Brief description of the drawings]
1 is a synthetic reaction formula of an oxazoline derivative. FIG. 2 is a synthetic reaction formula of a reaction between an oxazoline derivative and mannose (Example 1).
FIG. 3 shows a synthetic reaction formula of a reaction between an oxazoline derivative and paranitraphenyl galactopyranoside (Example 2).
FIG. 4 is a synthetic reaction formula for the reaction of a glucooxazoline derivative and methanol (Example 3).
FIG. 5 shows a synthetic reaction formula for the reaction of a galactooxazoline derivative and methanol (Example 4).

Claims (5)

N-acetyl- β1- catalyzed by a cation exchange resin using N-acetylhexosamine oxazoline compound represented by the general formula (1) as a sugar donor and sugars, alcohols, or phenols as a sugar acceptor. A method for producing a hexosaminide derivative.

(In the formula, R1, R2, R3 and R4 represent the substituents shown in Table 1.)

The production method according to claim 1, wherein the N-acetyl-β1-hexosaminide derivative is a 2-acetamido-2-deoxy-β1-D-hexopyranoside derivative. The N-acetyl-β1-hexosaminide derivative is a 2-acetamido-2-deoxy-β1-D-glucopyranoside derivative and a 2-acetamido-2-deoxy-β1-D-galactopyranoside derivative. Manufacturing method. The production method according to claim 1, wherein the N-acetyl-β1-hexosaminide derivatives are GlcNAcβ1-6Man, GlcNAcβ1-3Man, GlcNAcβ1-6Galβ-OpNp, GlcNAcβ1-OMe and GalNAcβ1-OMe. The manufacturing method of any one of Claims 1-4 whose solvent is an aprotic polar solvent.

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