CN1199977C - Simple process for synthesizing 3,6-branched mannopentaose - Google Patents

Abstract

The present invention relates to mannose core pentasaccharide α-D-mannose 1→2-α-D-mannose 1→6-(α-D-mannose 1→2- A simple chemical synthesis of α-D-mannose 1→3)-D-mannose.

Description

Facile Synthesis of 3,6-Branched Mannopentaose

The invention belongs to the field of preparation of biologically active oligosaccharides, in particular to a synthesis method of oligosaccharides which can be used for drug screening.

The oligosaccharide chain linked to asparagine is referred to as N-sugar chain, which usually has a manno-oligosaccharide containing 7-9 mannosyl groups, and this manno-oligosaccharide always contains a 3, 6-branched manno-core pentasaccharide , as shown in the following structural formula

The structure of N-glycan chains is characterized by at least one oligosaccharide diantennium branched on mannose with 3, 6. In addition, high mannose N-glycan chains with 5-9 mannose are complex type N -The predecessor of sugar chain synthesis, so the appearance or increase of high mannose sugar chains in tumor glycoproteins indicates that the processing of sugar chains is incomplete and immature due to the need for malignant cell proliferation and increased glycoprotein synthesis speed. Synthesis of mannose core pentasaccharide is the basis for the preparation of related antigens or diagnostic reagents, and is an essential raw material for glycobiology research. In addition, the combination of mannocorepentasaccharide and CoA can be applied to the separation and purification of this protein, which has potential industrial application prospects.

The purpose of the present invention is to complete the synthesis of the above-mentioned mannose core pentasaccharide in a one-step method, using 1,2-ethylidene-β-D-mannoside as the acceptor, and mannobiose trichloride linked by α1→2 Acetimide ester is used as sugar group donor, and mannopentaose can be easily synthesized by using Lewis acid as catalyst. Compared with the reported method, it is greatly simplified and suitable for batch preparation.

The synthetic method of the present invention is:

Using mannoside 1 as the glycosyl acceptor and α1→2-linked mannobiose trichloroacetimidate 2 as the glycosyl donor, one-step condensation into mannopentaose 3 linked at the 3 and 6 positions, removing the iso The propylidene and acetyl groups give mannopentaose 4. The following reaction formula is shown:

Bz = benzoyl, Ac = acetyl

The glycosyl donor 2 of the α1→2-linked mannanbiose trichloroacetimide ester was condensed from the mannose orthoester 7 to obtain the α1→2-linked mannanbiose 8, and the allyl group was removed and then trichloro Acetonitrile activates the 1 position to obtain 2, and the orthoester 7 of the sugar is prepared from acetylbromomannose 5, as shown in the following reaction formula:

Bz = benzoyl, Ac = acetyl All = allyl

The condensation reaction needs to use an organic solvent halogenated alkanes, ethers or amides as a solvent, and a Lewis acid as a catalyst.

The organic solvent used is preferably dichloromethane, and the Lewis acid used is preferably trimethylsilyl triflate or boron trifluoride.

The present invention will be described in detail below in conjunction with the examples.

(1) Synthesis of protected manna core pentasaccharide 3

1,2 Ethylidene mannose 1 (1.03 g, 5 mmol), glycosyl donor 2 (11.5 g, 10 mmol) of α1→2-linked mannobiose trichloroacetimide ester was dissolved in 100 In milliliter of dichloromethane, cool to -10°C, add 100 microliters of trimethylsilyl trifluoromethanesulfonate under stirring, then let the reactant naturally warm to room temperature, add 200 microliters of triethylamine after five hours to finish After reaction, the solvent was distilled off under reduced pressure, and intermediate 3 (16.5 g, 77%) was obtained after separation by column chromatography. m.p141-144°C; [α] _D -11.5° (c1.1, CHCl ₃ ); ¹ H NMR δ7.98-7.25 (m, 60H, Bz-H), 5.67 (dd, 1H, J 3.0Hz, J 1.7Hz), 5.64(dd, 1H, J 3.1Hz, J 1.7Hz), 5.53(s, 1H), 5.26(q, 1H, CH ₃ -CH), 5.20(s, 1H), 5.12(d, 1H, J 1.6Hz), 5.08(d, 1H, J 1.7Hz), 5.05(d, 1H, J 1.7Hz), 2.01(s, 3H, _CH3CO ), 2.00(s, 3H, _CH3CO ) , 1.50 (d, 3H, CH ₃ -CH).

(2) Synthesis of manna core pentasaccharide 4

The protected manna core pentasaccharide 3 (520 mg, 0.26 mmol) was dissolved in 10 ml of 90% trifluoroacetic acid, stirred at room temperature, monitored by TLC (ethyl acetate/petroleum ether 1/1), the reaction After completion, the solvent was distilled off under reduced pressure, and the intermediate was obtained after separation by column chromatography. The intermediate was dissolved in 50 ml of methanol saturated with ammonia. After one week, the reaction was completed, and the solvent was distilled off under reduced pressure to obtain the powder After washing with ethyl acetate, 489 mg of mannose pentasaccharide was obtained with a yield of 69%.

¹ H NMR (D ₂ O) δ5.32, 5.20, 5.15, 5.13, 4.85.

(3) 3,4,6-tri-O-benzoyl-β-D-mannose 1,2-allyl orthoester (7)

2,3,4,6-Tetra-O-acetyl-α-D-mannose bromide (8.22 g, 20 mmol) was added to allyl alcohol (40 mL), and 2,4-di Pyridine (2.3mL, 20mmol), the reaction was carried out at room temperature, and detected by TLC (1:1 petroleum ether/ethyl acetate). After the reaction was completed, it was concentrated under reduced pressure, and the residue was suspended in MeOH in anhydrous methanol ( 20 mL), was added methanol solution of sodium methoxide (2M, 0.4 mL), the mixture was stirred overnight, and the reaction was completed. The reaction solution was concentrated under reduced pressure, and the residue was separated by column chromatography to obtain powder 6 (4.72 g, 90%). 6 was dissolved in pyridine (30mL), and benzoyl chloride (8.2mL, 70mmol) was added dropwise under stirring. After three hours, the reaction was complete, and the reaction solution was post-treated by conventional methods, and column chromatography (3/1 petroleum ether/ethyl acetate) to obtain crystalline compound 7 (10.33 g, ~100%).

(4) 2-O-acetyl-3,4,6-tri-O-benzoyl-α-D-mannose-(1→2)-3,4,6-tri-O-benzoyl -α-D-Mannose allyl glycoside (8)

Compound 7 was dried under high vacuum for two hours, then dissolved in dichloromethane (80mL). TMSOTf (100 microliters) was added dropwise under nitrogen protection at -42°C. After 2 hours, TLC (3/1 petroleum ether/acetic acid ethyl ester) indicated that the reaction was complete. Neutralized with triethylamine, concentrated under reduced pressure, and purified by column chromatography to obtain 8 as a solid (6.72 g, 66%). mp 148-150°C; [α] _D +3.0°(c 1.3 _C ^_ ₅₉ H ₅₂ O ₁₈ : C, 67.56; H, 4.96. Found: C, 67.60; H, 4.92.

(5) 2-O-acetyl-3,4,6-tri-O-benzoyl-α-D-mannose-(1→2)-3,4,6-tri-O-benzoyl -α-D-mannose trichloroacetimidate (2) Compound 8 (524 mg, 0.5 mmol) was added to 90% acetic acid (10 ml) containing sodium acetate (293 mg, 3 mmol), and then PdCl ₂ (89 mg, 0.5 mmol), the mixture was stirred at room temperature for 12 hours, TLC (2/1 petroleum ether/ethyl acetate) indicated that the reaction was complete, diluted with dichloromethane (30 mL), diluted with water, saturated sodium bicarbonate The solution was washed, the organic phase was concentrated under reduced pressure, and purified by column chromatography to obtain 2-O-acetyl-3,4,6-tri-O-benzoyl-α-D-mannose-(1→2)- 3,4,6-Tri-O-benzoyl-α-D-mannose (456 mg), dissolved in dichloromethane (20 mL), added _CCl3CN (0.1 mL, 2 mmol) and DBU (14 microliters, 0.18 mmol), the reaction was completed after 6 hours, the reaction solution was concentrated under reduced pressure, and purified by column chromatography to obtain crystallized compound 2 (473 mg, two-step yield 90%), mp 139-142 ° C; [α] _D -1.5° (c 1.1, CHCl ₃ ), ¹ H NMR δ 8.64, 8.02-7.26, 6.59, 6.09, 5.95-5.87, 5.70, 5.20, 4.68-4.48, 2.03. Calculated for C ₅₈ H ₄₈ O ₁₈ Cl ₃ N: C, 60.3 1; H, 4.16. Found: C, 60.34; H, 4.15.

Claims (4)

1. With 1,2-ethylidene-β-D-mannoside as acceptor, mannobiose trichloroacetimide ester connected with α1→2 as glycosyl donor, and Lewis acid as catalyst The simple and easy synthetic method of mannose pentasaccharide is characterized in that: Using mannoside 1 as the glycosyl acceptor and α1→2-linked mannobiose trichloroacetimidate 2 as the glycosyl donor, one-step condensation into mannopentaose 3 linked at the 3 and 6 positions, removing the iso The propylidene group and the acetyl group promptly obtain mannopentaose 4, as shown in the following reaction formula:

Bz = benzoyl, Ac = acetyl 2. according to the method for claim 1, the glycosyl donor 2 of the mannobiose trichloroacetimide ester of α1→2 connection is condensed by mannose orthoester 7, obtains the mannanbiose 8 of α1→2 connection, desorbs After the allyl group is removed, the 1-position is activated with trichloroacetonitrile to obtain 2, and the sugar orthoester 7 is prepared from acetylbromomannose 5, as shown in the following reaction formula:

Bz = benzoyl, Ac = acetyl All = allyl 3. The condensation reaction described in claim 1 and 2 needs to use organic solvent halogenated alkanes, ethers or amides as solvent, and needs to use Lewis acid as catalyst. 4. The organic solvent used in claim 3 is dichloromethane, and the Lewis acid used is trimethylsilyl triflate or boron trifluoride.