Method for producing GlcNAc salts by enzymic method and purifying GlcNAc salts

Abstract

The invention discloses a method for producing GlcNAc salts by an enzymic method and purifying the GlcNAc salts, and belongs to the technical field of bioengineering. According to the method, N-acetyl-D-(+)-glucosamine is used as a raw material and is hydrolyzed by deacetylase to obtain GlcNAc and acetic acid; then, the GlcNAc salts are obtained through separation by eluting cation exchange columns by an acid solution; and meanwhile, a byproduct of sodium acetate is obtained through anion exchange recovery. The obtained GlcNAc salts are subjected to concentration, crystallization, decoloring and drying to obtain high-purity GlcNAc salt crystals. According to the method, a cyclic utilization process of enzymes, a cyclic recovery process of residue substrates and a recovery process of aceticacid are combined. The conversion rate of the N-acetyl-D-(+)-glucosamine and the total yield of the GlcNAc salt product are improved, and meanwhile, enzymes are cyclically utilized, so that the residue substrates circulate to be sufficiently converted, and the acetic acid is utilized as resources. Through operation conditions at the normal temperature, the resin loss rate is low; the generation quantity of hydrochloric acid waste liquid is very small; and the economic benefits of energy saving and consumption reduction and the effects of environment protection and safety are realized.

Description

technical field

[0001] The invention relates to enzymatic production of glucosamine salt and a purification method thereof, belonging to the technical field of bioengineering. Background technique

[0002] Glucosamine (GlcNAc) is an important hexosamine sugar, which is formed by replacing a hydroxyl group of glucose with an amino group, and is easily soluble in water and hydrophilic solvents. Widely present in nature, the chemical name is: 2-amino-2-deoxy-D-glucose, usually in the form of N-acetyl derivatives (such as chitin) or in the form of N-sulfate and N-acetyl-3-O-lactic acid The ether (muramic acid) form is present in microorganisms, polysaccharides of animal origin and conjugated polysaccharides. Glucosamine hydrochloride, molecular weight 215.5Da, white crystal, odorless, slightly sweet, soluble in water, slightly soluble in methanol, insoluble in organic solvents such as ethanol. The glucosamine molecule is not very stable and is prone to oxidation or degradation...

Examples

Embodiment 1

[0059] according to figure 1 The process route shown, the operation steps are as follows:

[0060] (1) Use acetylglucosamine solution with a concentration of 40-150g / L as raw material, add deacetylase solution or deacetylase preparation according to the ratio of 10-30U / g acetylglucosamine, and the pH range of the enzyme reaction is 4.0-8.0 , the reaction temperature is 25-45°C, and the reaction is stirred for 10-40min;

[0061] This step specifically removes the acetyl group in the N-acetylglucosamine molecule, and reacts to obtain an enzyme hydrolyzate whose main components are glucosamine and acetic acid. The pH of the enzyme reaction is preferably 7.0-8.0;

[0062] (2) The enzyme hydrolyzate after the step (1) reaction is transported in the ultrafiltration membrane equipment, the molecular weight cut-off of the ultrafiltration membrane is 5-200kDa, preferably 5-30kDa; Ultrafiltration membrane dialysate and deacetylase-containing membrane concentrate, the recovery rate of ...

Embodiment 2

[0078] according to figure 1 The process route shown, the operation steps are as follows:

[0079] (1) Collect 350m 3 The concentration is 102kg / m 3 The acetylglucosamine solution in the storage tank, pump the acetylglucosamine solution into the enzyme reaction tank, add the deacetylase solution according to the ratio of 10-25U / g acetylglucosamine, the pH of the enzyme reaction is 7.0-8.0, and the reaction temperature at 37°C, stirring for 30 minutes;

[0080] (2) Pump the enzyme hydrolyzate after the reaction in step (1) into the ultrafiltration membrane equipment, the molecular weight cut-off of the ultrafiltration membrane is 5000Da, add 60m 3 Pure water is used for the dialysis membrane concentrate, and the ultrafiltration membrane dialysis fluid and the membrane concentrate are collected separately; a total of 380m 3 The content is 56.3kg / m 3 of glucosamine dialysate; 30m of membrane concentrate 3 The enzyme liquid is returned to the deacetylase liquid storage tank to...

Embodiment 3

[0095] according to figure 1 The process route shown is different in that, on the basis of Example 2, the nanofiltration membrane concentration equipment is replaced with reverse osmosis ceramic membrane equipment, that is, the column liquid under the negative column obtained in step (4) is collected to the reverse osmosis ceramic membrane equipment, the pore size of the reverse osmosis membrane is 1nm, and the operating pressure is 0.5-1.0MPa. The liquid concentrated by the reverse osmosis ceramic membrane equipment is returned to the enzyme reaction tank to participate in the next batch of enzyme reaction process.

[0096] In this production process, the production of 1 ton of glucosamine hydrochloride consumes 11 tons of pure water and produces 10.5 tons of waste water.

technical field

[0001] The invention relates to enzymatic production of glucosamine salt and a purification method thereof, belonging to the technical field of bioengineering. Background technique

[0002] Glucosamine (GlcNAc) is an important hexosamine sugar, which is formed by replacing a hydroxyl group of glucose with an amino group, and is easily soluble in water and hydrophilic solvents. Widely present in nature, the chemical name is: 2-amino-2-deoxy-D-glucose, usually in the form of N-acetyl derivatives (such as chitin) or in the form of N-sulfate and N-acetyl-3-O-lactic acid The ether (muramic acid) form is present in microorganisms, polysaccharides of animal origin and conjugated polysaccharides. Glucosamine hydrochloride, molecular weight 215.5Da, white crystal, odorless, slightly sweet, soluble in water, slightly soluble in methanol, insoluble in organic solvents such as ethanol. The glucosamine molecule is not very stable and is prone to oxidation or degradation...

Examples

Embodiment 1

[0059] according to figure 1 The process route shown, the operation steps are as follows:

[0060] (1) Use acetylglucosamine solution with a concentration of 40-150g / L as raw material, add deacetylase solution or deacetylase preparation according to the ratio of 10-30U / g acetylglucosamine, and the pH range of the enzyme reaction is 4.0-8.0 , the reaction temperature is 25-45°C, and the reaction is stirred for 10-40min;

[0061] This step specifically removes the acetyl group in the N-acetylglucosamine molecule, and reacts to obtain an enzyme hydrolyzate whose main components are glucosamine and acetic acid. The pH of the enzyme reaction is preferably 7.0-8.0;

[0062] (2) The enzyme hydrolyzate after the step (1) reaction is transported in the ultrafiltration membrane equipment, the molecular weight cut-off of the ultrafiltration membrane is 5-200kDa, preferably 5-30kDa; Ultrafiltration membrane dialysate and deacetylase-containing membrane concentrate, the recovery rate of ...

Embodiment 2

[0078] according to figure 1 The process route shown, the operation steps are as follows:

[0079] (1) Collect 350m 3 The concentration is 102kg / m 3 The acetylglucosamine solution in the storage tank, pump the acetylglucosamine solution into the enzyme reaction tank, add the deacetylase solution according to the ratio of 10-25U / g acetylglucosamine, the pH of the enzyme reaction is 7.0-8.0, and the reaction temperature at 37°C, stirring for 30 minutes;

[0080] (2) Pump the enzyme hydrolyzate after the reaction in step (1) into the ultrafiltration membrane equipment, the molecular weight cut-off of the ultrafiltration membrane is 5000Da, add 60m 3 Pure water is used for the dialysis membrane concentrate, and the ultrafiltration membrane dialysis fluid and the membrane concentrate are collected separately; a total of 380m 3 The content is 56.3kg / m 3 of glucosamine dialysate; 30m of membrane concentrate 3 The enzyme liquid is returned to the deacetylase liquid storage tank to...

Embodiment 3

[0095] according to figure 1 The process route shown is different in that, on the basis of Example 2, the nanofiltration membrane concentration equipment is replaced with reverse osmosis ceramic membrane equipment, that is, the column liquid under the negative column obtained in step (4) is collected to the reverse osmosis ceramic membrane equipment, the pore size of the reverse osmosis membrane is 1nm, and the operating pressure is 0.5-1.0MPa. The liquid concentrated by the reverse osmosis ceramic membrane equipment is returned to the enzyme reaction tank to participate in the next batch of enzyme reaction process.

[0096] In this production process, the production of 1 ton of glucosamine hydrochloride consumes 11 tons of pure water and produces 10.5 tons of waste water.

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