JPH06135981A - Sialic acid derivative - Google Patents

Abstract

PURPOSE:To provide a new derivative useful as an intermediate for gangliosides such as GD3, GT1a. or GQ1b. CONSTITUTION:The objective derivative of formula I [R, R<x> and R<1> are each H or protecting group; n is 1 or 2; A is (2 3) or (2 6) glycoside bond; Sugar is (protecting group-bearing) galactose residue; where, R<x> may form lactone intramolecularly], e.g. 2-(trimethylsilyl)ethyl -0- (5-acetamido-3,5-dideoxy-D- glycero-alpha-D-galacto-2-nonuropyranosonicacid) - (2 8)-0-(5-acetamido-3,5-dideoxy-D- glycero-alpha-D-galacto-2-nonuropyranosonicacid) (2 3)-beta-galactopyranoside. The compound of the formula I can be obtained, for example, by lactonizing a compound of formula II followed by reaction with acetic anhydride and then reaction with thiophenol and a galactose derivative.

Description

Detailed Description of the Invention

[0001]

The present invention relates, for example ganglioside _{G D3, G T1a, G Q1b} , G Q1c, on the usefulness of the sialic acid derivative as a synthetic intermediate for gangliosides such as G _P1c.

In the present specification: * "Ac" indicates an acetyl group.

* "Ph" or "Phenyl" represents a phenyl group.

* "Bn" represents a benzyl group.

* "Bz" represents a benzoyl group.

* "Me" represents a methyl group.

* "SE" indicates a 2-trimethylsilylethyl group.

In the present specification, when the same compound (eg, general formula (i)) has many groups represented by the same symbol (eg, “R”), each “R” may be the same. Good or different.

[0009]

2. Description of the Related Art Gangliosides are present on the cell surface of animals, and their sugar chains are oriented extracellularly, recognizing external information and demonstrating the presence of self, hormones, viruses, bacterial toxins, and other receptors. Function, cell differentiation,
It is a generic name for sialosphingolipids that are deeply involved in basic life phenomena such as proliferation, canceration, and immunity. As a ganglioside, N-acetylneuraminic acid is α (2-8)
G _Q1b , G that is dimerized or _trimerized by binding
_Q1c or are such as G _P1c are known, in the ganglioside G _Q1b, are known to exhibit a nerve growth factor-like activity has been attracting attention. However, ganglioside exists only in a very small amount in the living body, and in addition to the diversity of oligosaccharide chain structures, the diversity of sialic acid and the ceramide molecule that is a lipid moiety is also obtained, and it is obtained as a pure pure compound from nature. Is extremely difficult. Therefore, it is essential to synthesize a single compound by synthetic means. However, the synthesis of the di- or trisialic acid derivative requires a multi-step synthetic process, side reactions are likely to occur, the separation of by-products is difficult, and the production yield problem is also considered. Gangliosides G _Q1b , G useful di- or trisialylated derivatives as synthetic intermediates such _Q1c or G _P1c has not been obtained.

[0010]

The object of the present invention is to solve the above is to provide a useful di- or trisialylated derivative ganglioside G _Q1b, as synthetic intermediates, such as G _Q1c or G _P1c.

[0011]

DISCLOSURE OF THE INVENTION As a result of intensive studies in view of the above-mentioned object, the present inventor found that the following novel di- or tri-N-acetylneuraminic acid derivative can be obtained from di- or tri-N-acetylneuraminic acid. It was found that

That is, the present invention has the following general formula (i):

[0013]

[Chemical 2]

[Wherein R, R ^x , R ', n, A and S
ugar is the same as above. ] The sialic acid derivative represented by this is provided.

In the present invention, examples of the protecting group represented by R include an acetyl group, a benzyl group and a benzoyl group.

In the present invention, the protective group represented by R ^x includes, in addition to the group represented by R, a group formed by forming a lactone with the carboxyl group of the adjacent N-acetylneuraminic acid residue. Be done.

In the present invention, examples of the protective group represented by R'include a lower alkyl group having 1 to 4 carbon atoms such as methyl group, ethyl group, propyl group and butyl group.

In the present invention, A represents a (2 → 3) or (2 → 6) glycosidic bond. Where (2 →
3) The bond means a bond between the hydroxyl group at the 2-position of N-acetylneuraminic acid and the hydroxyl group at the 3-position of galactose. The (2 → 6) bond means a bond between the 2-position hydroxyl group of N-acetylneuraminic acid and the 6-position hydroxyl group of galactose.

"Sugar" of the sialic acid derivative represented by the general formula (i) means a galactose residue which may have a protecting group represented by Gal, or a protecting group represented by Lac. It is a galactose residue that may be used. Here, the galactose residue which may have a protecting group represented by Gal means the following formula

[0020]

[Chemical 3]

[Wherein R ¹ is the same or different and is a hydrogen atom, acetyl group, benzyl group or benzoyl group, R ² is the same or different and is a hydrogen atom, acetyl group, benzyl group or benzoyl group, Alternatively, it shows a bond with the hydroxyl group at the 2-position of N-acetylneuraminic acid, and R
³ represents a hydroxyl group, a methoxy group, a benzyloxy group, an acetoxy group, a 2-trimethylsilylethoxy group, a methylthio group or a phenylthio group. ] It is a group represented by.

In the "Sugar" residue of the sialic acid derivative represented by the general formula (i), the lactose residue which may have a protecting group represented by Lac is represented by the following formula:

[0023]

[Chemical 4]

[Wherein R ^a is the same or different and is a hydrogen atom, acetyl group, benzyl group or benzoyl group, and R ^b is the same or different and is a hydrogen atom, acetyl group, benzyl group or benzoyl group, Alternatively, it shows a bond with the hydroxyl group at the 2-position of N-acetylneuraminic acid, and R
^c represents a hydroxyl group, a methoxy group, a benzyloxy group, an acetoxy group, a 2-trimethylsilylethoxy group, a methylthio group or a phenylthio group. ] It is a group represented by.

Among the sialic acid derivatives represented by the general formula (i) of the present invention, the compound in which the Sugar residue is a galactose derivative can be produced, for example, according to the following reaction process formula (I).

Reaction process formula (I)

[0027]

[Chemical 5]

[0028]

[Chemical 6]

[Wherein R, R ', R ¹ , R ² , R ³ , G
al, n and A are the same as above. ] The above reaction process formula (I)
In: * The compound of the formula (2) is obtained by subjecting di- or tri-N-acetylneuraminic acid (1) to methyl esterification and lactonization in anhydrous methanol under acidic conditions such as HCl and sulfuric acid, and then the lactone form 1 thereof. Acetic anhydride 10 to 2 per mol
It is obtained by using about 0 mol and reacting with pyridine or the like as a catalyst. The reaction temperature is about 20 to 40 ° C., and the reaction time is about 1 to 12 hours.

* The compound of the formula (3) can be obtained by reacting 1 mol of the acetyl protected compound of the formula (2) with about 1 to 4 mol of thiophenol. As the reaction solvent, dichloromethane, dichloroethane, toluene or the like can be used, the reaction temperature is about 20 to 30 ° C., and the reaction time is about 1 to 12 hours.

* The compound of the formula (ia) contains 2 mol of the galactose derivative (4) per 1 mol of the compound (3).
It can be obtained by using about 3 mol and reacting. As the reaction solvent, acetonitrile or the like can be used, the reaction temperature is about -40 to -20 ° C, and the reaction time is 24.
It is about 48 hours.

* The compound of the formula (i-b) is the compound (i
-A) can be obtained by deprotecting in the presence of a catalyst. As the catalyst, alkali metal alkoxides such as sodium methoxide, sodium ethoxide, potassium t-butoxide, and alkali metal hydroxides such as sodium hydroxide and potassium hydroxide are used. The reaction is carried out in water or an alcohol solvent such as methanol, ethanol or t-butanol using 0.01 to 0.02 mol of the above catalyst per 1 mol of the compound (ia). The reaction temperature is about 20 to 30 ° C, and the reaction time is 1
It is about 24 hours.

* Introducing a protecting group into the compound of formula (ib) by a conventional method can lead to another compound (i) of the present invention having a protecting group, for example, protecting a benzoyl group. When introduced as a group, about 3 to 4 mol of benzoic anhydride is used per 1 mol of the compound of formula (ib), pyridine or the like is used as a solvent, and a catalyst such as dimethylaminopyridine or the like is used if necessary. Then, the desired compound is obtained by reacting. The reaction temperature is about 0 to 30 ° C., and the reaction time is about 1 to 12 hours.

Among the compounds represented by the general formula (I) of the present invention, the compound in which Sugar is Lac can be produced according to the following reaction process formula (II).

Reaction process formula (II)

[0036]

[Chemical 7]

[Wherein R, R ', R ^a , R ^b , R ^c , n
And A are the same as above. ] In the above reaction process formula (II): * The compound of the formula (ic) contains 2 to 3 of any one of the above two kinds of Lac derivatives per 1 mol of the compound of the formula (3).
It can be obtained by reacting in a molar amount. As a solvent, acetonitrile or the like can be used, and the reaction temperature is -40.
The reaction time is about 24 to 48 hours.

* The compound of formula (id) has the formula (i-
It can be obtained by deprotecting the compound of c) in the presence of a catalyst. As the catalyst, alkali metal alkoxides such as sodium methoxide, sodium ethoxide and potassium t-butoxide, alkali metal hydroxides such as sodium hydroxide and potassium hydroxide, and palladium are used. The reaction is carried out by the compound (i
-C) It can be obtained by using about 0.01 to 0.02 mol of the catalyst for 1 mol of the reaction. As a reaction solvent, water or methanol, ethanol, t-
Butanol or the like can be used, the reaction temperature is about 20 to 40 ° C., and the reaction time is about 1 to 48 hours.

* Introducing a protecting group into the compound of formula (id) by a conventional method can lead to another compound (i) of the present invention having a protecting group, for example, protecting a benzoyl group. When introduced as a group, about 3 to 4 mol of benzoic anhydride is used per 1 mol of the compound of formula (id), pyridine or the like is used as a solvent, and a catalyst such as dimethylaminopyridine or the like is used if necessary. Then, the desired compound is obtained by reacting. The reaction temperature is about 0 to 30 ° C., and the reaction time is about 1 to 12 hours.

In the compound of the formula (ic) obtained by the above reaction, for example, the compound in which R ^c is a 2-trimethylsilylethyl group and A is (2 → 3) is prepared according to the following reaction process formula (III). Can lead to gangliosides.

Reaction process formula (III)

[0042]

[Chemical 8]

[0043]

[Chemical 9]

[0044]

[Chemical 10]

[Wherein R, R ', R ^a , R ^b , R ^c ,
n and A are the same as above. ] In the above reaction process formula (III): * The compound of formula (i-1) is obtained by deprotecting the SE group of the compound of formula (i-c) in the presence of trifluoroacetic acid. A halogen-based solvent such as dichloromethane is used as the reaction solvent, and 1 mol of the compound (ic) is used.
It is obtained by reacting 1.2 mol to an excess amount of trifluoroacetic acid. The reaction temperature is about 20 to 30 ° C., and the reaction time is about 1 to 3 hours.

* The compound of the formula (i-2) can be obtained by reacting the compound (i-1) with trichloroacetonitrile in a halogen solvent such as dichloromethane.
About 10 to 20 mol of trichloroacetonitrile is used with respect to 1 mol of the compound (i-1). The reaction temperature is about 0 ° C., and the reaction time is about 2 to 4 hours.

* A compound of formula (i-3) can be obtained by reacting a compound of formula (i-2) with a 2-azidosphingosine derivative in a solvent such as dichloromethane. The reaction is carried out with respect to 1 mol of the compound of the formula (i-2),
It is carried out using about 2-3 mol of the 2-azidosphingosine derivative. The reaction temperature is about -20 to 0 ° C, and the reaction time is 2
It is about 8 hours.

* The compound of formula (i-3) is reduced with H ₂ S and reacted with stearic acid in the presence of a dehydrating agent in a solvent such as dichloromethane to give a protected ganglioside (i-4) having ceramide bound thereto. Obtainable. The reaction is carried out using an excess amount of H ₂ S with respect to the compound of the formula (i-3) and using about 1.5 to 3 mol of stearic acid for 1 mol of the amine derivative produced. Reaction temperature is 20 ~
The reaction time is about 30 ° C. and about 10 to 20 hours.

* Then, the compound of formula (i-4) is deprotected to obtain the desired ganglioside. Deprotection is carried out according to a conventional method by using about 0.01 to 0.02 mol of a catalyst such as palladium-carbon, PtO ₂ , or Raney nickel with respect to 1 mol of the compound of the formula (i-4), water or methanol, ethanol, It can be carried out by hydrogenation in an alcohol solvent such as t-butanol.

The compound of the present invention can be purified by a conventional purification means such as solvent extraction, recrystallization, silica gel column chromatography and gel filtration chromatography.

[0051]

According to the present invention, it is possible to obtain a synthetic intermediate of which was difficult ganglioside conventional synthesis, thereby G _Q1b, is possible to easily obtain a natural ganglioside, such as G _Q1c or G _P1c I can do it now.

[0052]

EXAMPLES The present invention will be described in more detail below with reference to examples.

Example 1 Methyl 5-acetamido-2,4,7-tri-O-acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O- Acetyl-3,5
-Dideoxy-D-glycero-α-D-galacto-2-
Nonuropyranosilono-1 ', 9-lactone) -D-glycero-D-galacto-2-nonuropyranosylonate
Production of (2) (hereinafter abbreviated as compound (2)) O- (5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonic acid)-(2 → 8) -5-acetamide-3,5
-Dideoxy-D-glycero-α-D-galacto-2-
Nonuropyranosonic acid (hereinafter abbreviated as compound (1); 5.0 g, 8.3 mmol) and Amberlite IR-120 (H ⁺ ) (20 g) were suspended in methanol (300 ml), and the suspension was adjusted to 2 at 40 ° C. It was stirred for a day.
After completion of the reaction, TLC (dichloromethane: methanol =
After confirmation with 2: 1), the resin was filtered off and washed with methanol. The filtrate and washings were combined and concentrated under reduced pressure. The residue was dissolved in acetic anhydride (15 ml), cooled to 0 ° C. and pyridine (1
(5 ml) was added dropwise, and the mixture was stirred at 40 ° C for 1 day. The reaction solution was cooled to 0 ° C., methanol was added to decompose excess acetic anhydride, and the residue was azeotropically distilled with toluene, and the residue was extracted with dichloromethane. The organic layer was washed with 2M-hydrochloric acid and saturated sodium carbonate, and dried over sodium sulfate. This was separated by filtration and then concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (manufactured by Wako Pure Chemical Industries, Ltd., 200 mesh) to obtain an eluate (dichloromethane: methanol = 50 :).
Elution at 1) gave the title compound (2) (6.20 g, 84
%) Was obtained.

* Elemental analysis (molecular formula: C ₃₇ H ₅₀ O ₂₃ N ₂ ) C H N calculated value 49.89 5.66 3.14 measured value 49.75 5.83 2.96 * IR (film): ν (Cm ^-1 ) 3400, 3200, 3070, 2950, 1760,
1670

[0055]

[Chemical 11]

Example 2 Phenyl [methyl 5-acetamido-4,7-di-]
O-acetyl-3,5-dideoxy-2-thio-8-O
-(5-acetamido-4,7,8,9-tetra-O-
Acetyl-3,5-dideoxy-D-glycero-α-D
-Galact-2-nonuropyranosilono-1 ', 9-lactone) -D-glycero-D-galacto-2-nonulopyranosylonate (3) (hereinafter abbreviated as compound (3)) Compound ( 2) (1 g, 1.1 mmol) was dissolved in dichloromethane (10 ml) and thiophenol (0.5 m
1, 4.4 mmol) and boron trifluoride ether complex (2 ml, 8.8 mmol) were added, and the mixture was stirred at room temperature for 12 hours. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 15: 1), the reaction solution was extracted with dichloromethane. The organic layer was washed with saturated sodium carbonate and water in this order, and dried over sodium sulfate. This was separated by filtration and then concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (200, manufactured by Wako Pure Chemical Industries, Ltd.
Mesh) and eluted with an eluent (dichloromethane: methanol = 50: 1) to give the title compound (3) (940).
mg, 89%).

* Elemental analysis (molecular formula: C ₄₁ H ₅₂ O ₂₁ N ₂ S) C H N calculated value 52.34 5.57 2.98 measured value 52.31 5.27 2.90 * IR (film): ν (cm ⁻¹ ) 3400, 3200, 3070, 2900, 1750,
1660, 690 Example 3 2- (Trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosylono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonulopyranosylonate]-(2 → 3) -6
-O-benzoyl-β-D-galactopyranoside
(6) (hereinafter abbreviated as compound (6)) Compound (3) (600 mg, 650 μmol) and 2-
(Trimethyl) ethyl 6-O-benzoyl-β-D-galactopyranoside (4) (750 mg, 2.0 mmo
l) is dissolved in acetonitrile (25 ml), molecular sieves 3 Å (2.5 g) is added,
The mixture was stirred at room temperature for 5 hours. Then cool to -35 ° C and
-Iodosuccinimide (300 mg, 1.3 mmo
l), trifluormethane sulfonic acid (23 μl, 13
0 μmol) was added and the mixture was stirred at −35 ° C. for 2 days. After completion of the reaction, TLC (dichloromethane: methanol = 3)
After confirmation with 0: 1), the solid was filtered off and extracted with dichloromethane. The organic layer was washed successively with saturated sodium carbonate and saturated sodium thiosulfate, and dried over sodium sulfate. This was filtered off and concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (Wako Pure Chemical Industries, Ltd., 200 mesh) and eluted with an eluent (dichloromethane: methanol = 40: 1) to give the title compound. (6) (250 mg, 32%) and its β-isomer (75
mg, 10%) was obtained.

* Elemental analysis (molecular formula: C ₅₃ H ₇₄ O ₂₈ N ₂ Si) C H N calculated value 52.38 6.14 2.31 measured value 52.13 6.10 2.60 * IR (film): ν (cm ⁻¹ ) 3400, 3300, 3100-2900, 1750,
_{^{1670,860,840,710 * [α] 20 D:}} -21.8 ° (c = 1.1, CHCl
₃ )

[0059]

[Chemical 12]

Example 4 2- (Trimethylsilyl) ethyl O- (5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonuropyranosonic acid)-(2 →
8) -O- (5-acetamido-3,5-dideoxy-
Production of D-glycero-α-D-galacto-2-nonulopyranosonic acid)-(2 → 3) -β-galactopyranoside (7) (hereinafter abbreviated as compound (7)) Compound (6) (20 mg, 16.5 μmol) was dissolved in methanol (4 ml), sodium methoxide (catalyst) was added, and the mixture was stirred at 45 ° C. for 1 day and then 0.2 N
Add sodium hydroxide (catalyst) and add 4 more at 45 ° C.
Stir for hours. After confirming the completion of the reaction by TLC (butanol: ethanol: water = 2: 1: 1), IR-120 (H
⁺ ) Was used to neutralize the reaction solution. After the resin was filtered off and washed with methanol, the filtrate and washings were combined and concentrated under reduced pressure, and the resulting syrup was subjected to gel chromatography (Sephadex L).
H-20) and by eluting with an eluent (methanol), the title compound (7) (14 mg, quantitative) was obtained.

* Elemental analysis (molecular formula: C ₃₃ H ₅₇ O ₂₁ N ₂ Si) C H N calculated value 46.86 6.79 3.31 measured value 46.66 6.51 3.33 * [α] ²⁰ _D : + 60.0 ° (c = 0.03, Me
OH) Example 5 2- (trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosylono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonulopyranosylonate]-(2 → 6) -3
-O-benzoyl-β-D-galactopyranoside
(8) (hereinafter, abbreviated as compound (8)) Compound (3) (300 mg, 320 μmol) and 2-
(Trimethylsilyl) ethyl-3-O-benzoyl-β
-D-galactopyranoside (250 mg, 640 μmo
l) was treated by the same procedure as in Example 3 to obtain the title compound (8) (128 mg, 33%) and its β isomer. (35 mg, 9.0%) was obtained.

* Elemental analysis (molecular formula: C ₅₃ H ₇₄ O ₂₈ N ₂ Si) C H N calculated value 52.38 6.14 2.31 measured value 52.47 6.01 2.30 * IR (film): ν (cm ⁻¹ ) 3400, 3300, 3100-2900, 1750,
1670, 860, 840, 710 * [α] ²⁰ _D : -18.6 ° (c = 0.4, CHCl
₃ )

[0063]

[Chemical 13]

Example 6 2- (Trimethylsilyl) ethyl O- (5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonic acid)-(2 →
8) -O- (5-acetamido-3,5-dideoxy-
Production of D-glycero-α-D-galacto-2-nonulopyranosonic acid)-(2 → 6) -β-D-galactopyranoside (9) (hereinafter abbreviated as compound (9)) Compound ( 8) (20 mg, 17 μmol) was treated in the same manner as in Example 4 to give the title compound (9) (14 m
g, quantitative).

* Elemental analysis (molecular formula: C ₃₃ H ₅₇ O ₂₁ N ₂ Si) C H N calculated value 46.86 6.79 3.31 measured value 46.87 6.90 3.20 * [α] ²⁰ _D : -30.0 ° (c = 0.03, Me
OH) Example 7 2- (trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosilono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonuropyranosylonate]-(2 → 3) -O
-(2,6-di-O-benzyl-β-D-galactopyranosyl)-(1 → 4) -2,3,6-tri-O-benzyl-β-D-glucopyranoside (11) (hereinafter , Abbreviated as compound (11)) Compound (3) (300 mg, 320 μmol) and 2-
(Trimethylsilyl) ethyl O- (2,6-di-O-
Benzyl-β-D-galactopyranosyl)-(1 → 4)
-2,3,6-Tri-O-benzyl-β-D-glucopyranoside (10) (570 mg, 640 μmol) was treated in the same manner as in Example 3 to give the title compound (11).
(170 mg, 31%) was obtained.

* Elemental analysis (molecular formula: C ₈₇ H ₁₁₀ O ₃₂ N ₂ Si) C H N calculated value 60.62 6.43 1.63 measured value 60.52 6.59 1.64 * IR (film): ν (cm ⁻¹ ) 3400, 3300, 3150-2870, 1750,
1670, 860, 840, 700 * [α] ²⁰ _D : -12.3 ° (c = 0.7, CH ₂
Cl ₂ ) Example 8 Methyl 5-acetamido-2,4,7-tri-O-acetyl-3,5-dideoxy-8-O- [5-acetamido-4,7-di-O-acetyl-3, 5-Dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-D-galacto-2-nonuropyranosylono-1 ′ ', 9'
-Lactone) -D-glycero-α-D-galacto-2-
Nonuropyranosilono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonuropyranosylonate (13) (hereinafter abbreviated as compound (13)) O- (5- Acetamide-3,5-dideoxy-D-glycero-α-D-galacto-2-nonuropyranosonic acid)-(2 → 8) -O- (5-acetamido-
3,5-Dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonic acid)-(2 → 8)-
(5-acetamido-3,5-dideoxy-D-glycero-D-galacto-2-nonulopyranosonic acid) (12) (100 mg, 112 μmol) and IR
Suspend -120 (H ⁺ ) in methanol and add 2 at 40 ° C.
It was stirred for a day. Completing the reaction by TLC (dichloromethane:
After confirmation with methanol = 1: 1), the resin was filtered off and washed with methanol. The filtrate and washings were combined and concentrated under reduced pressure, and the resulting syrup was dissolved in acetic anhydride (0.3 ml),
The mixture was cooled to ° C, pyridine (0.3 ml) was added dropwise, and the mixture was further stirred at 40 ° C for 1 day. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 15: 1), the reaction solution was extracted with dichloromethane. The organic layer is 2M-hydrochloric acid,
It was washed with saturated sodium carbonate and dried over sodium sulfate. This was filtered off and concentrated under reduced pressure. The obtained syrup was subjected to silica gel column chromatography (Wako Pure Chemical Industries, Ltd., 200 mesh) and eluted with an eluent (dichloromethane: methanol = 25: 1) to give the title. The compound (13) (67 mg, 48%) was obtained.

* Elemental analysis (molecular formula: C ₅₂ H ₆₉ O ₃₂ N ₃ ) C H N calculated value 60.53 6.39 1.59 measured value 60.43 6.55 1.60 * IR (film): ν (Cm ⁻¹ ) 3400,3100-2900,1750,1670 Example 9 Phenyl [methyl 5-acetamido-4,7-di-
O-acetyl-3,5-dideoxy-2-thio-8-O
-[5-acetamido-4,7-di-O-acetyl-
3,5-dideoxy-8-O- (5-acetamido-
4,7,8,9-Tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosilono-1 ″, 9′-lactone) -D-glycero -Α-D-galacto-2-nonulopyranosilono-1 ',
9-lactone] -D-glycero-D-galacto-2-nonuropyranosylonate] (14) (hereinafter, referred to as compound (1
The compound (13) (120 mg, 96 μmol) was treated in the same manner as in Example (2) to give the title compound (14) (100 mg, 80%). * Elemental analysis (molecular formula: C ₅₆ H ₇₁ O ₃₀ N ₃ S) C H N calculated value 51.81 5.51 3.24 measured value 51.67 5.68 3.06 * IR (film): ν (cm ^-1 ) 3400, 3100-2900, 1750, 1670,
690 Example 10 2- (Trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7-di-
O-acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O-acetyl-
3,5-dideoxy-D-glycero-α-D-galacto-2-nonuropyranosilono-1 ″, 9′-lactone)-
D-glycero-α-D-galacto-2-nonuropyranosilono-1 ′, 9-lactone) -D-glycero-α-D-
Galact-2-Nonuropyranosylonate]-(2 → 3)
-O- (2,6-di-O-benzyl-β-D-galactopyranosyl)-(1 → 4) -2,3,6-tri-O-benzyl-β-D-glucopyranoside (15) (Less than,
Preparation of compound (15) Abbreviated compound (14) (100 mg, 77 μmol) and compound (10) (140 mg, 150 μmol) in acetonitrile (5 ml), and added molecular sieve 3 angstrom (500 mg) at room temperature. Stir for 5 hours. Then, it cooled at -30 degreeC, N-iodosuccinimide (52 mg, 230 micromol) and trifluoromethanesulfonic acid (2 microliter, 23 micromol) were added, and -3
The mixture was stirred at 0 ° C for 5 days. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 25: 1), the solid matter was separated by filtration and extracted with dichloromethane. The organic layer was washed successively with saturated sodium carbonate and saturated sodium thiosulfate, and dried over sodium sulfate. This was filtered off and concentrated under reduced pressure. The obtained syrup was subjected to silica gel column chromatography (Wako Pure Chemical Industries, Ltd., 200 mesh) and eluted with an eluent (dichloromethane: methanol = 25: 1) to give the title. Compound (15) (78 mg, 49
%) Was obtained.

* Elemental analysis (molecular formula: C ₁₀₂ H ₁₂₉ O ₄₁ N ₃ Si) C H N calculated value 58.87 6.25 2.02 measured value 58.73 6.42 1.84 * IR (film): ν (cm ⁻¹ ) 3400, 3300, 3100-2900, 1750,
1670, 860, 840, 700 * [α] ²⁰ _D : -15.3 ° (c = 1.6, CHCl
₃ ) Example 11 2- (trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosylono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonulopyranosylonate]-(2 → 6) -3
-O-benzyl-β-D-galactopyranoside (1
7) (hereinafter abbreviated as compound (17)) Compound (3) (300 mg, 320 μmol) and 2-
(Trimethylsilyl) ethyl 3-benzyl-β-D-
Galactopyranoside (16) (310 mg, 840 μm)
was treated by the same method as in Example 5 to obtain the title compound (17) (183 mg, 48%) and its β-isomer (61 mg, 18%). Compound (1
The physical property values of 7) are shown below.

* Elemental analysis (molecular formula: C ₅₃ H ₇₆ O ₂₇ N ₂ Si) C H N calculated value 52.99 6.38 2.33 measured value 52.79 6.10 2.35 * IR (film): ν (cm ⁻¹ ) 3400, 3300, 3100-2900, 1750,
1670, 860, 840, 700 * [α] ²⁰ _D : -23.5 ° (c = 1.2, CHCl
₃ ) Example 12 2- (trimethylsilyl) ethyl O- [methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosilono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonulopyranosylonate]-(2 → 3)-
2,4,6-Tri-O-benzoyl-β-D-galactopyranoside (18) (hereinafter abbreviated as compound (18))
Production of Compound (6) (200 mg, 165 μmol) was dissolved in pyridine (1 ml), and benzoic anhydride (150 mg, 6
60 μmol) and 4-dimethylaminopyridine (2
3 mg, 165 μmol) was added, and the mixture was stirred at room temperature for 10 hours. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 50: 1), the mixture was cooled to 0 ° C. and methanol was added to decompose excess reagent. Then concentrate under reduced pressure,
The obtained syrup was extracted with dichloromethane, and the organic layer was washed with 2M-hydrochloric acid and water in this order and dried over sodium sulfate. This is filtered off, washed with dichloromethane, and the filtrate and washings are combined and concentrated under reduced pressure. The obtained syrup is subjected to silica gel column chromatography (manufactured by Wako Pure Chemical Industries, Ltd., 200 mesh) to obtain an eluate (dichloromethane). : Methanol = 30: 1) to give the title compound (18) (180 mg, 77%).

* Elemental analysis (molecular formula: C ₆₇ H ₈₂ O ₃₀ N ₂ Si) C H N calculated value 56.53 5.81 1.97 measured value 56.50 5.51 1.89 * IR (film): ν (cm ⁻¹ ) 3400, 3100-2900, 1750, 1670,
860, 840, 710 * [α] ²⁰ _D : -5.6 ° (c = 0.7, CHC
l ₃ )

[0071]

[Chemical 14]

Example 13 O- [methyl 5-acetamido-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O -Acetyl-3,5
-Dideoxy-D-glycero-α-D-galacto-2-
Nonuropyranosilono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonuropyranosylonate]-(2 → 3) -1-O-acetyl-2,4,6- Tri-O-benzoyl-β-D-galactopyranose
(19) (hereinafter abbreviated as compound (19)) Compound (18) (180 mg, 125 μmol) was dissolved in toluene (2 ml), and acetic anhydride (0.2 ml) and boron trifluoride ether complex (60 μl, 25
(0 μmol) was added, and the mixture was stirred at room temperature for 1 day. Completion of the reaction by TLC (dichloromethane: methanol = 25:
After confirmation in 1), it was extracted with dichloromethane. The organic layer was washed with saturated sodium hydrogen carbonate and water in this order, and dried over sodium sulfate. This was filtered off and concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (Wako Pure Chemical Industries, Ltd., 200 mesh) and eluted with an eluent (dichloromethane: methanol = 40: 1) to give the title compound. (19) (142 mg, 82%) was obtained.

* Elemental analysis (molecular formula: C ₆₄ H ₇₂ O ₃₁ N ₂ Si) C H N calculated value 56.30 5.31 2.05 measured value 56.16 5.48 1.87 * [α] ²⁰ _D : + 7.7 ° (c = 0.9, CHC
l ₃ ) * IR (film): ν (cm ⁻¹ ) 3400, 3100-2950, 1750, 1670,
710

[0074]

[Chemical 15]

Example 14 Methyl O- [methyl 5-acetamido-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-
Acetamide-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosylono-1 ′, 9-lactone)-
D-glycero-α-D-galacto-2-nonuropyranosylonate]-(2 → 3) -2,4,6-tri-O-benzoyl-1-thio-β-D-galactopyranoside ( Two
0) (hereinafter abbreviated as compound (20)) Compound (19) (140 mg, 100 μmol) was dissolved in dichloromethane (1 ml) and cooled to 0 ° C. Methylthiotrimethylsilane (35 μl, 250 μmol)
And boron trifluoride ether complex (25μ
1, 100 μmol) and then stirred at room temperature for 4 hours. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 25: 1), the reaction solution was extracted with dichloromethane. The organic layer was washed with saturated sodium carbonate and water in this order, and dried over sodium sulfate. This was separated by filtration and then concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (manufactured by Wako Pure Chemical Industries, Ltd., 200 mesh) to obtain an eluate (dichloromethane: methanol = 40: 1).
And eluted with the title compound (20) (106 mg, 76
%) Was obtained.

* Elemental analysis (molecular formula: C ₆₃ H ₇₂ O ₂₉ N ₂ S) C H N calculated value 55.91 5.36 2.07 measured value 56.13 5.10 2.33 * [α] ²⁰ _D : + 10.7 ° (c = 1.4, CHCl
₃ ) * IR (film): ν (cm ^-1 ) 3400, 3100-2900, 1750, 1670,
710

[0077]

[Chemical 16]

Example 15 2- (Trimethylsilyl) ethyl O- (5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonuropyranosilonic acid)-(2
→ 8) -O- (5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonuropyranosylonic acid)-(2 → 3) -O- (β-D- Preparation of galactopyranosyl)-(1 → 4) -β-D-glucoside (21) (hereinafter abbreviated as compound (21)) Compound (11) (100 mg, 58 μmol) was added to methanol (5 ml) and acetic acid (1 ml). ) Was dissolved in a mixed solvent. Next, palladium-carbon (100 mg) was added, and the mixture was stirred at 45 ° C for 2 days while blowing hydrogen gas. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 5: 1), the solid matter was separated by filtration and washed with methanol. The filtrate and washings were combined and concentrated under reduced pressure, the obtained syrup was dissolved in methanol (10 ml), sodium methoxide (catalytic amount) was added, and the mixture was stirred at room temperature for 1 day. Then, 0.2N-potassium hydroxide aqueous solution (catalyst amount) was added and stirred for 4 hours. After confirming the completion of the reaction by TLC (butanol: ethanol: water = 2: 2: 1),
The reaction solution was neutralized with IR-120 (H ⁺ ). The resin was filtered off, washed with methanol, the filtrate and washings were combined and concentrated under reduced pressure, the obtained syrup was subjected to column chromatography (Sephadex LH-20), and eluted with an eluent (methanol), The title compound (21) (50 mg,
85%).

* Elemental analysis (molecular formula: C ₃₉ H ₆₈ O ₂₇ N ₂ Si) C H N calculated value 45.70 6.68 2.37 measured value 45.56 6.86 2.55 * [α] ²⁰ _D : -13.3 ° (c = 0.9, MeO
H) Example 16 2- (Trimethylsilyl) ethyl O- (methyl 5-
Acetamide-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamide-4,7,8,
9-Tetra-O-acetyl-3,5-dideoxy-D-
Glycero-α-D-galacto-2-nonulopyranosylono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonuropyranosylonate]-(2 → 3) -O
-(2,4,6-tri-O-acetyl-β-D-galactopyranosyl)-(1 → 4) -2,3,6-tri-O-
Acetyl-β-D-glucopyranoside (22) (hereinafter,
Production of compound (22) Abbreviated compound (11) (100 mg, 58 μmol) was dissolved in a mixed solvent of methanol (10 ml) and acetic acid (10 ml). Next, palladium-carbon (100 mg) was added, and the mixture was stirred at 45 ° C for 2 days while blowing hydrogen gas. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 5: 1), the solid matter was separated by filtration and washed with methanol. The filtrate and washings were combined and concentrated under reduced pressure, the obtained syrup was dissolved in pyridine (2 ml), acetic anhydride (1 ml) was added, and the mixture was stirred at room temperature for 1 day. Completion of the reaction by TLC (dichloromethane: methanol = 20: 1)
After confirmation with, the mixture was cooled to 0 ° C. and methanol was added to decompose excess reagent. Then, the mixture was concentrated under reduced pressure, and the obtained syrup was extracted with dichloromethane. The organic layer was washed with 2M-hydrochloric acid and saturated sodium carbonate, and dried over sodium sulfate. This was separated by filtration, washed with dichloromethane, and the filtrate and washings were combined and concentrated under reduced pressure, and the syrup thus obtained was subjected to silica gel column chromatography (manufactured by Wako Pure Chemical Industries, Ltd., 2
00 mesh) and eluted with the eluent (dichloromethane: methanol = 30: 1) to give the title compound (22)
(80 mg, 90%) was obtained.

* Elemental analysis (molecular formula: C ₆₄ H ₉₂ O ₃₈ N ₂ Si) C H N calculated value 50.39 6.08 1.84 measured value 50.36 5.78 1.76 * [α] ²⁰ _D : -4.9 ° (c = 1.8, CHC
l ₃ ) * IR (film): ν (cm ⁻¹ ) 3400, 3100-2900, 1750, 1670,
860,840

[0081]

[Chemical 17]

Example 17 O- [methyl 5-acetamido-4,7-di-O-acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O) -Acetyl-3,5
-Dideoxy-D-glycero-α-D-galacto-2-
Nonuropyranosilono-1 ′, 9-lactone) -D-glycero-α-D-galacto-2-nonulopyranosylonate]-(2 → 3) -O- (2,4,6-tri-O -Acetyl-β-D-galactopyranosyl)-(1 → 4)-
2,3,6-tri-O-acetyl-D-glucopyrano-
Preparation of compound (23) (hereinafter abbreviated as compound (23)) Compound (22) (75 mg, 50 μmol) was added to dichloromethane (0.6 ml) and trifluoroacetic acid (1.3).
(ml) mixed solvent and stirred at room temperature for 1.5 hours. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 15: 1), ethyl acetate was added. Then, the mixture was concentrated under reduced pressure, and the obtained syrup was subjected to silica gel column chromatography (200 mesh manufactured by Wako Pure Chemical Industries, Ltd.) to obtain an eluate (dichloromethane: methanol = 3).
Elution with 0: 1) gave the title compound (23) (63 mg,
90%).

* Elemental analysis (molecular formula: C ₅₈ H ₈₀ O ₃₈ N ₂ ) C H N calculated value 49.72 5.66 1.97 measured value 49.52 5.38 1.99 * IR (film): ν (Cm ⁻¹ ) 3400, 3300, 3100-2900, 1750,
1670 Example 18 O- [methyl 5-O-acetamide-4,7-di-O
-Acetyl-3,5-dideoxy-8-O- (5-acetamido-4,7,8,9-tetra-O-acetyl-
3,5-Dideoxy-D-glycero-α-D-galacto-2-nonuropyranosilono-1 ′, 9-lactone) -D
-Glycero-α-D-galacto-2-nonuropyranosylonate]-(2 → 3) -O- (2,4,6-tri-O-
Acetyl-β-D-galactopyranosyl)-(1 → 4)
Production of -O- (2,3,6-tri-O-acetyl-α-D-glucopyranosyl)-(1 → 1) -trichloroacetimidate (24) (hereinafter abbreviated as compound (24)) Compound ( 23) (60 mg, 42 μmol) was dissolved in dichloromethane (1 ml) and cooled to 0 ° C. And
Trichloroacetonitrile (0.2 ml, 160 μmo
l) and 1,8-diazabicyclo [5,4,0] -7-undecene (10 μl, 50 μmol) were added, and the mixture was stirred at 0 ° C. for 2 hours. After confirming the completion of the reaction by TLC (dichloromethane: methanol = 15: 1), the reaction solution was subjected to silica gel column chromatography (manufactured by Wako Pure Chemical Industries, Ltd., 200 mesh), and the eluent (dichloromethane: methanol = 25: 1). ), And the title compound (24) (57 mg, 86%) was obtained.

* Elemental analysis (molecular formula: C ₆₁ H ₈₃ O ₃₈ N ₃ Cl ₃ ) C H N calculated value 46.59 5.32 2.67 actual value 46.45 5.49 2.49 * IR (film) : Ν (cm ^-1 ) 3400, 3100-2900, 1750, 1670 * [α] ²⁰ _D : + 27.4 ° (c = 1.1, CHCl)
₃ ) From the above compound (24), the above reaction process formula (III)
It has been confirmed that the desired ganglioside having ceramide bound thereto can be obtained according to the above.

[0085]

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Yoji Tsukada, Inventor Yoji Tsukada 23 Fukami Dewayashiki-cho, Fushimi-ku, Kyoto-shi, Kyoto Prefecture B904 Fushimi B904 (72) Inventor Yasuhiro Ota 1 of 59, Gokasho Ichiban, 59, Uji, Kyoto Ichi Bankan 501

Claims (1)

[Claims] 1. The following general formula (i): [In the formula, R, R ^x and R ′ are the same or different and each represents a hydrogen atom or a protective group, n is 1 or 2, and A is
Represents a (2 → 3) or (2 → 6) glycoside bond, and Sugar represents a galactose residue which may have a protecting group or a lactose residue which may have a protecting group. However, R ^x may form an intramolecular lactone. ] The sialic acid derivative represented by.