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Method for purifying oxidized beta-nicotinamide adenine dinucleotide phosphate

A technology of nicotinamide adenine and dinucleotide, applied in chemical instruments and methods, organic chemistry, preparation of sugar derivatives, etc., can solve the problems of difficult separation of phosphate radicals, low product purity, low yield, etc., and achieve reduction Production cost, the effect of solving the difficult problem of phosphate residue

Active Publication Date: 2015-09-02
BONTAC BIO ENG SHENZHEN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In view of the shortcomings of the above-mentioned prior art, the object of the present invention is to provide a purification method for oxidized β-nicotinamide adenine dinucleotide phosphate, which aims to solve the problem that the phosphate group is difficult to separate in the prior art and the product purity low yield problem

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] 1. Concentration of crude product:

[0048] The pretreated coenzyme II solution is subjected to microfiltration and nanofiltration using membrane concentration equipment, microfiltration removes microorganisms, and nanofiltration uses a hollow fiber membrane with a molecular weight cut-off of 200 to concentrate the crude product to 40-60g / L.

[0049] 2. Purification:

[0050] Purification conditions:

[0051] Chromatographic column: 5cm*30cm;

[0052] Stationary phase: phenyl bonded silica gel;

[0053] Mobile phase: A phase: hydrochloric acid solution with pH 2; B phase: ethanol.

[0054] Flow rate: 50-80ml / min;

[0055] Detection wavelength: 260 nm;

[0056] Gradient: B%: 1% to 10% (40 min).

[0057] The injection volume is 10-15g.

[0058] Purification process: adjust the pH of the concentrated crude product solution to 2-4 with phosphoric acid solution or hydrochloric acid solution, rinse the chromatographic column with more than 30% ethanol solution, and the...

Embodiment 2

[0062] 1. Concentration of crude product:

[0063] The pretreated coenzyme II solution is subjected to microfiltration and nanofiltration using membrane concentration equipment, microfiltration removes microorganisms, and nanofiltration uses a hollow fiber membrane with a molecular weight cut-off of 200 to concentrate the crude product to 40-60g / L.

[0064] 2. Purification:

[0065] Purification conditions:

[0066] Chromatographic column: 15cm*30cm;

[0067] Stationary phase: phenyl bonded silica gel;

[0068] Mobile phase: A phase: hydrochloric acid solution with a pH of 3; B phase: ethanol;

[0069] Flow rate: 400-500ml / min;

[0070] Detection wavelength: 260 nm;

[0071] Gradient: B%: 1% ~ 10% (40 min);

[0072] The injection volume is 70-90g.

[0073] Purification process: adjust the pH of the concentrated crude product solution to 2-4 with phosphoric acid solution or hydrochloric acid solution, rinse the chromatographic column with more than 30% ethanol solution, ...

Embodiment 3

[0076] 1. Concentration of crude product:

[0077] The pretreated coenzyme II solution is subjected to microfiltration and nanofiltration using membrane concentration equipment, microfiltration removes microorganisms, and nanofiltration uses a hollow fiber membrane with a molecular weight cut-off of 200 to concentrate the crude product to 40-60g / L.

[0078] 2. Purification:

[0079] Purification conditions:

[0080] Chromatographic column: 30cm*30cm;

[0081] Stationary phase: phenyl bonded silica gel;

[0082] Mobile phase: A phase: hydrochloric acid solution with pH 4; B phase: ethanol;

[0083] Flow rate: 2500-3000ml / min;

[0084] Detection wavelength: 260 nm;

[0085] Gradient: B%: 1% ~ 10% (40 min);

[0086] The injection volume is 400-500g.

[0087] Purification process: adjust the pH of the concentrated crude product solution to 2-4 with phosphoric acid solution or hydrochloric acid solution, rinse the chromatographic column with more than 30% ethanol solution, a...

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Abstract

The invention discloses a method for purifying oxidized beta-nicotinamide adenine dinucleotide phosphate. The method for purifying oxidized beta-nicotinamide adenine dinucleotide phosphate concretely comprises the following steps: a, successively carrying out microfiltration and nanofiltration on a pretreated coenzyme II solution by adopting membrane concentration equipment, so that a concentrated crude product solution is obtained; b, regulating the pH value of the obtained crude product solution to 2-4, injecting a sample, loading the sample onto a reversed high performance liquid chromatography preparation column, carrying out gradient elution, and purifying, so that a purified sample solution is obtained; and c, carrying out nanofiltration on the purified sample solution by adopting the membrane concentration equipment, and freeze-drying by adopting a vacuum freeze-drying machine, so that purified coenzyme II is obtained. The coenzyme II prepared by adopting the method disclosed by the invention is high in purity, and the yield is high, so that the method disclosed by the invention has a good market prospect.

Description

technical field [0001] The invention relates to the field of nucleotide coenzymes, in particular to a method for purifying oxidized β-nicotinamide adenine dinucleotide phosphate. Background technique [0002] Oxidized β-nicotinamide adenine dinucleotide phosphate (referred to as: coenzyme Ⅱ, English: Nicotinamide adenine dinucleotide phosphate, NADP) is an extremely important nucleotide coenzyme, which is nicotinamide adenine dinucleotide (NAD) is a phosphorylated derivative of the 2'-position of the ribose ring linked to adenine, which participates in various anabolic reactions, such as the synthesis of lipids, fatty acids and nucleotides. It also serves as a medium for phosphate transfer and participates in various synthetic reactions. [0003] Coenzyme II has a wide range of uses. At present, the traditional purification process mostly uses ion exchange resin purification and other means. However, due to the influence of the synthesis process of Coenzyme II, the crude pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H19/207C07H1/06
CPCC07H21/02C07H1/06C07H19/207
Inventor 傅荣昭戴柱张琦
Owner BONTAC BIO ENG SHENZHEN
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