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CN-119255975-B - Preparation method of sodium hyaluronate oligosaccharide composition, composition prepared by preparation method and application of composition

CN119255975BCN 119255975 BCN119255975 BCN 119255975BCN-119255975-B

Abstract

The invention belongs to the field of biochemistry, and particularly relates to a preparation method of a sodium hyaluronate oligosaccharide composition, and further discloses the prepared composition and application. According to the preparation method of the sodium hyaluronate oligosaccharide, macromolecular sodium hyaluronate is used as a raw material, an acidic aqueous solution is prepared, ethanol is added for precipitation reaction, the supernatant is separated, impurities are removed through an ultrafiltration membrane, the second ethanol precipitation reaction is carried out, and the separated precipitate is purified through a reverse-phase ODS column, so that the sodium hyaluronate oligosaccharide with the required performance can be obtained. The sodium hyaluronate oligosaccharide composition prepared by the method comprises disaccharide, tetrasaccharide, hexasaccharide, octasaccharide, decasaccharide and dodecasaccharide, and has the advantages of better product stability, excellent water solubility, good transdermal property, moisture retention, good oral absorbability and the like.

Inventors

  • GUO WENJUAN
  • CAO JIE
  • LI WENCHAO
  • FU QU
  • QU SHOUWEI
  • FU JUNCHANG

Assignees

  • 北京瑞济善健康科技有限公司
  • 付俊昌

Dates

Publication Date
20260512
Application Date
20220819

Claims (7)

  1. 1. A method for preparing a sodium hyaluronate oligosaccharide composition, comprising the steps of: (1) Preparing macromolecular sodium hyaluronate serving as a raw material to form an acidic aqueous solution and reacting; The molecular weight of the macromolecular sodium hyaluronate is 5 ten thousand-200 ten thousand Da; the acid adopted for preparing the acid aqueous solution is sulfuric acid or hydrochloric acid, and the concentration of the acid is 0.5-4.0M; the mass ratio of the macromolecular sodium hyaluronate to the acid is 1:5-15; The temperature of the reaction step is 50-150 ℃ and the reaction time is 3-15 hours; (2) Regulating pH of the reaction solution to neutrality, adding ethanol to reach ethanol concentration of 50-70%, and collecting first precipitate and supernatant; (3) Removing impurities from the supernatant with ultrafiltration membrane, adding ethanol to ethanol concentration of 70-90%, and collecting second precipitate; the molecular weight cut-off of the ultrafiltration membrane is 1500-3000da; (4) Dissolving the second precipitate in water, and purifying with reversed-phase ODS column.
  2. 2. The method for preparing a sodium hyaluronate oligosaccharide composition according to claim 1, wherein in the step (4), the purification step comprises a step of eluting with 0.5 to 3v/v% aqueous ethanol solution and a step of eluting with 40 to 60v/v% aqueous ethanol solution, collecting an eluted fraction of 40 to 60v/v% ethanol, recovering ethanol, and drying.
  3. 3. The method for preparing a sodium hyaluronate oligosaccharide composition according to claim 1 or 2, further comprising the step of dissolving the first precipitate in water, purifying with an activated carbon column, collecting the purified product and repeating steps (1) to (2) using the purified product as a raw material; The step of purifying by using the activated carbon column comprises the step of eluting by using water and the step of eluting by using 20-40v/v% ethanol water solution, collecting the elution fraction of the 20-40v/v% ethanol water solution, recovering ethanol and drying to obtain the required purified product.
  4. 4. The method for preparing the sodium hyaluronate oligosaccharide composition according to claim 1 or 2, wherein: controlling the second precipitate to be dissolved by adding water until the concentration is 10-25mg/mL; controlling the first precipitate to be dissolved by adding water until the concentration is 20-35mg/mL.
  5. 5. A sodium hyaluronate oligosaccharide composition prepared by the method of any one of claims 1-4, wherein the oligosaccharide composition comprises sodium hyaluronate disaccharide, sodium hyaluronate tetrasaccharide, sodium hyaluronate hexasaccharide, sodium hyaluronate octasaccharide, sodium hyaluronate decasaccharide, and sodium hyaluronate dodecasaccharide; wherein, based on 100% of the total amount of the oligosaccharide composition, The sum of the mass contents of the sodium hyaluronate hexasaccharide, the sodium hyaluronate octasaccharide and the sodium hyaluronate decasaccharide is 40-60wt%; The content of the sodium hyaluronate disaccharide is 10-24wt%, the content of the sodium hyaluronate tetrasaccharide is 20-33wt%, and the content of the sodium hyaluronate dodecasaccharide is 3-8wt%; The mass ratio of the sodium hyaluronate hexasaccharide, the sodium hyaluronate octasaccharide and the sodium hyaluronate decasaccharide is 20-26:13-19:7-17.
  6. 6. The sodium hyaluronate oligosaccharide composition according to claim 5, wherein the sodium hyaluronate oligosaccharide composition has an average molecular weight of 1100-160 da and a 1wt% aqueous solution pH of 6-7.
  7. 7. Use of the sodium hyaluronate oligosaccharide composition of any one of claims 5 or 6 or the sodium hyaluronate oligosaccharide composition prepared by the method of any one of claims 1-5 for the preparation of a food product, a medical product and/or a daily chemical product.

Description

Preparation method of sodium hyaluronate oligosaccharide composition, composition prepared by preparation method and application of composition Technical Field The invention belongs to the field of biochemistry, and particularly relates to a preparation method of a sodium hyaluronate oligosaccharide composition, and further discloses the prepared composition and application. Background Sodium hyaluronate is sodium glucuronate, has a chemical formula of (C14H 20N011 Na) N, is an inherent component in human body, and has no species specificity, and widely exists in tissues such as placenta, amniotic fluid, crystalline lens, articular cartilage, dermis layer of skin and the like, and cytoplasm and intercellular substance of organs, and has lubricating and nourishing effects on cells and organelles contained in the sodium hyaluronate. Moreover, sodium hyaluronate is a major component of extracellular matrix, and also an important component of skin, vitreous, joint synovial fluid and cartilage tissue, has unique physicochemical properties and biological functions, and plays an important role in maintaining tissue integrity and in tissue formation and remodeling during infection, injury, and embryonic development. Sodium hyaluronate is classified according to its molecular weight, and generally includes macromolecular sodium hyaluronate (molecular weight range: 160-160 Da), medium molecular sodium hyaluronate (molecular weight range: 20-160 Da), small molecular sodium hyaluronate (molecular weight range: 1-20 Da), and low molecular sodium hyaluronate (molecular weight range: 3000-1 Da). Although sodium hyaluronate has been widely used in the fields of medical science, daily chemical products, nutritional foods, pet foods and the like, sodium hyaluronate currently used has a large molecular weight and is too viscous, so that the sodium hyaluronate has poor transdermal absorption, poor oral absorption, poor bioavailability and the like, and has a plurality of limitations in application fields. Thus, the development of low molecular weight sodium hyaluronate and even sodium hyaluronate oligosaccharides has been a concern for those skilled in the art. In the prior art, the preparation method of the low-molecular sodium hyaluronate product is based on the degradation of macromolecular sodium hyaluronate and mainly comprises a physical method, a chemical method, a biological enzymolysis method and the like, but the problems of larger molecular weight of degradation products, wide molecular weight range of the products, unstable degradation effect, higher cost and the like generally exist. The average molecular weight of the sodium hyaluronate prepared by the method can be controlled to be 8000-20000Da, and the minimum molecular weight is about 5000Da, and the molecular weight range of the sodium hyaluronate prepared by the method is wider and can not meet the market demand. The method for producing the hyaluronic acid with small molecular weight by using the oxidative degradation method of hydrogen peroxide and ascorbic acid disclosed in Chinese patent CN101293934A has the problems of organic solvent residue, complex post-treatment process, environmental damage and the like. For another example, the methods disclosed in chinese patent CN103484513a and CN101507733a for preparing small molecule oligomeric hyaluronic acid by enzymatic method, but the methods apply genetic recombination technology, and the methods and operations are very complex and have high cost. In summary, sodium hyaluronate products in the prior art generally have the problems of unstable quality and large product difference, such as excessive molecular weight distribution, uneven color, low solubility and long dissolution time, and the preparation method of sodium hyaluronate generally has the problems of low efficiency (about 50%, more than 90% is difficult to reach), high cost, residual organic solvent and the like. Summary of The Invention Technical problem Therefore, the technical problem to be solved by the invention is to provide a preparation method of the sodium hyaluronate oligosaccharide composition, which has the advantages of simple process operation, low production cost and higher production efficiency; The second technical problem to be solved by the invention is to provide a sodium hyaluronate oligosaccharide composition product which is prepared by the preparation method and has stable quality, high solubility and quick dissolution, wherein the sodium hyaluronate oligosaccharide composition product not only can effectively improve the transdermal absorption effect, but also can effectively improve the skin moisture content and has good moisture retention; The third technical problem to be solved by the present invention is to provide the use of the above-mentioned sodium hyaluronate oligosaccharide composition. Solution to the problem Technical solution In order to solve the technical problems, the method for preparing the sodium hyal