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Preparation method of alpha-2,3-sialyllactulose

A technology of sialylation and lactulose, which is applied to the preparation of sugar derivatives, chemical instruments and methods, sugar derivatives, etc., can solve problems such as the difficulty of lactulose sialic acid glycosylation

Active Publication Date: 2016-09-21
HENAN INST OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to overcome the very difficult problem of carrying out sialic acid glycosylation of lactulose by chemical methods

Method used

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  • Preparation method of alpha-2,3-sialyllactulose
  • Preparation method of alpha-2,3-sialyllactulose
  • Preparation method of alpha-2,3-sialyllactulose

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0011] Raw material formula lactulose 40mg, cytidine sodium triphosphate (CTP) 95mg, sialic acid (Neu5Ac) 53mg, dissolved in 3000mg ultrapure water, adjusted pH 8.5 with 3 mol / L NaOH, added 1 mol / L pH 8 .5 Tris-HCl to make the concentration in the reaction system 100 mmol / L; 2 mol / L MgCl 2 , Make the concentration in the reaction system 20 mmol / L; 1mg CMP-sialyltransferase (NmCSS); 2mg sialyltransferase, namely PmST 1 . The mixture was then placed in a 37°C gas bath shaker and reacted for 20 hours. The degree of reaction was detected by silica gel thin layer chromatography (TLC), and the developing solvent of TLC was n-propanol:methanol:water=5:2:1. After the lactulose reaction is complete, add an equal volume of 95% ethanol to the reaction solution, mix well, put it in a refrigerator at 4°C for 30 minutes, then centrifuge at 7000 rpm for 30 minutes, and concentrate the supernatant by rotary evaporation. The concentrate was purified by silica gel chromatography. The ratio of ...

Embodiment 2

[0013] Lactulose 40mg, cytidine triphosphate (CTP) 110mg, sialic acid (Neu5Ac) 60mg, dissolved in 3000mg ultrapure water, adjusted pH 8.5 with 3 mol / L NaOH, and added 1 mol / L pH 8.5 Tris-HCl to make the concentration in the reaction system 100mM; 2mol / L MgCl 2 , The concentration in the reaction system is 20 mmol / L; 1.5mg CMP-sialyltransferase (NmCSS); 2.5mg sialyltransferase (PmST 1 ). The mixture was then placed in a 37°C gas bath shaker and reacted for 24 hours. The degree of reaction was detected by silica gel thin layer chromatography (TLC), and the developing solvent of TLC was n-propanol:methanol:water=5:2:1. After the lactulose reaction is complete, add an equal volume of 95% ethanol to the reaction solution, mix well, put it in a refrigerator at 4°C for 30 minutes, then centrifuge at 7000 rpm for 30 minutes, and concentrate the supernatant by rotary evaporation. The concentrate was purified by silica gel chromatography. The ratio of the purification solvent is ethyl ...

Embodiment 3

[0015] 40mg lactulose, 90mg cytidine triphosphate (CTP) 90mg, sialic acid (Neu5Ac) 50mg, dissolved in 3000mg ultrapure water, adjusted pH8.5 with 3mol / L NaOH, added 1mol / L pH8.5 Tris-HCl to make the concentration in the reaction system 100 mmol / L; 2 mol / L MgCl 2 , Make the concentration in the reaction system 20 millimoles / liter; 1.2mg of CMP-sialyltransferase, namely NmCSS; 2.3mg of sialyltransferase, namely PmST 1 . The mixture was then placed in a 37°C gas bath shaker and reacted for 26 hours. The degree of reaction was detected by silica gel thin layer chromatography (TLC), and the developing solvent of TLC was n-propanol:methanol:water=5:2:1. After the lactulose reaction is complete, add an equal volume of 95% ethanol to the reaction solution, mix well, put it in a refrigerator at 4°C for 30 minutes, then centrifuge at 7000 rpm for 30 minutes, and concentrate the supernatant by rotary evaporation. The concentrate was purified by silica gel chromatography. The ratio of th...

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Abstract

The invention discloses a preparation method of alpha-2,3-sialyllactulose. The method comprises the following steps: dissolving lactulose, sodium cytidine triphosphate and sialic acid in ultrapure water according to certain proportions, adjusting the pH value to 8.5 by using 3mol / L of NaOH, and adding 1mol / L of Tris-HCl with the pH value of 8.5; putting the above obtained mixture in a 37DEG C air bath oscillator, and reacting the mixture for 18-30h; detecting the reaction degree through adopting silica gel thin-layer chromatography (TLC), wherein a TLC developing solvent comprises n-propanol, methanol and water according to a ratio of 5:2:1; adding isopyknic 95% ethanol to a reaction solution obtained after completing the reaction of lactulose, uniformly mixing the reaction solution with ethanol, allowing the obtained mixed solution to stand in a 4DEG C refrigerator for 30min, centrifuging the obtained solution at 7000r / min for 30min, and carrying out rotary evaporative concentration on the obtained supernatant; and purifying the obtained concentrate by using a silica gel chromatographic column, and freeze-drying the purified concentrate to obtain a pure product.

Description

Technical field [0001] The invention belongs to the technical field of preparation of functional oligosaccharides, and specifically relates to a preparation method of alpha-2,3-sialylated lactulose. Background technique [0002] In the medical field, the potential of sialic acid oligosaccharides as antibacterial agents is very large. In addition, the expression of sialic acid on the cell membrane is also closely related to tumor generation and metastasis. The sialic acid analogs Zanamivir and Tamiflu have been approved by the US FDA as new drugs for the treatment of influenza. [0003] Existing research results show that sialic acid plays an extremely important role in many life phenomena of organisms, especially the research on functional oligosaccharides related to sialic acid is in the ascendant. Therefore, the synthesis of sialylated oligosaccharides and the sialylation modification with natural products as the parent, and the related biological activity research have formed ...

Claims

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Application Information

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IPC IPC(8): C07H1/00C07H7/033
CPCC07H1/00C07H7/033
Inventor 曾洁高海燕贾甜胡雅婕张瑞瑶孙俊良李光磊张军合
Owner HENAN INST OF SCI & TECH
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