Methods for providing sialylated oligosaccharides

Patent metadata

Field	Value
Publication number	US-9539270-B2
Application number	US-201214363856-A
Country	US
Kind code	B2
Filing date	Dec 6, 2012
Priority date	Dec 7, 2011
Publication date	Jan 10, 2017
Grant date	Jan 10, 2017

Abstract

The invention relates to a method for providing analogs of human milk oligosaccharides (HMO), in particular oligosaccharides containing terminal sialic acid. The method comprises the steps of: a) providing a source of non-digestible galactooligosaccharides (GOS) containing at least two terminally bonded β-linked galactose residues; b) providing a sialic acid donor having (α2-3)-sialylated O-glycans; c) contacting said GOS with said sialic acid donor in the presence of an enzyme having trans-sialidase activity in an enzyme reaction mixture; and d) isolating from said enzyme reaction mixture a fraction comprising at least 20 percent by weight of disialylated galactooligosaccharides (di-Sia-GOS) based on the dry matter.

First claim

The invention claimed is:

1. A method for providing a composition comprising sialic acid containing oligosaccharides, comprising the steps of:
a) providing a source of non-digestible galactooligosaccharides (GOS) containing at least two terminally bonded β-linked galactose residues;
b) providing a sialic acid donor having (α2-3)-sialylated O-glycans;
c) contacting said GOS with said sialic acid donor in the presence of an enzyme having trans-sialidase activity in an enzyme reaction mixture; and
d) obtaining from said enzyme reaction mixture a fraction comprising at least 5 percent by weight of disialylated galactooligosaccharides (di-Sia-GOS) based on the dry matter.

2. Method according to claim 1 , wherein said source of galactooligosaccharide is the product obtained by enzymatic treatment of lactose or a mixture of lactose and trehalose with β-galactosidase (EC 3.2.1.23).

3. Method according to claim 1 wherein said di-Sia-GOS have a degree of polymerization (DP) within the range of DP4 to DP10.

4. Method according to claim 1 , wherein said sialic acid donor is a naturally occurring compound selected from sialic acids bonded to oligosaccharides, polysaccharides, polysialic acids, glycoproteins and glycolipids.

5. Method according to claim 4 , wherein said sialic acid donor is selected from the group consisting of glycosylated whey proteins and casein, and fragments of the same.

6. Method according to claim 4 , wherein said sialic acid donor is selected from the group consisting of glycosylated mucus proteins, and fragments of the same.

7. Method according to claim 6 , wherein said sialic acid donor is the product obtained from mucin, optionally followed by concentration of Sia-containing mucin polymers.

8. Method according to claim 1 , wherein said enzyme having trans-sialidase activity is encoded by a gene product from microorganisms of the Trypanosoma genus, preferably Trypanosoma cruzi or Trypanosoma congolense.

9. Method according to claim 1 , wherein said enzyme is recombinantly produced.

10. Method according to claim 1 , wherein the pH of the enzyme reaction mixture ranges between 4 and 6.

11. Method according to claim 1 , wherein step d) comprises a separation technique based on a difference in charge between components to be separated.

12. Method according to claim 1 , wherein step d) comprises a separation technique based on a difference in size between components to be separated.

13. A method for providing an infant formula, comprising
a) providing a source of non-digestible galactooligosaccharides (GOS) containing at least two terminally bonded β-linked galactose residues;
b) providing a sialic acid donor having (α2-3)-sialylated O-glycans;
c) contacting said GOS with said sialic acid donor in the presence of an enzyme having trans-sialidase activity in an enzyme reaction mixture; and
d) obtaining from said enzyme reaction mixture a fraction comprising at least 5 percent by weight of disialylated galactooligosaccharides (di-Sia-GOS) based on the dry matter, and
e) formulating said fraction into an infant formula together with a protein source, a fat source and a carbohydrate source.

14. Method according to claim 3 wherein said di-Sia-GOS have a degree of polymerization (DP) within the range of DP5 to DP8.

15. Method according to claim 5 , wherein said sialic acid donor is glycomacropeptide from κ-casein (GMP).

16. Method according to claim 6 , wherein said sialic acid donor is pig small intestinal glycoprotein (PSMG).

17. Method according to claim 10 , wherein the pH of the enzyme reaction mixture ranges between 4.8 and 5.8.

18. Method according to claim 11 , wherein step d) comprises anion exchange chromatography.

19. Method according to claim 12 , wherein step d) comprises size-exclusion chromatography.

Assignees

• Friesland Brands Bv
• Univ Groningen
• Darling Ingredients Nederland B V

Inventors

• Sallomons Erik
• Wilbrink Maarten Hotse
• Sanders Peter
• Kamerling Johannis Paulus
• Van Vuure Catherina Anna
• Hage Johannes Adrianus

Classifications

• A61P33/02

  Antiprotozoals, e.g. for leishmaniasis, trichomoniasis, toxoplasmosis · CPC title

• A61P31/04

  Antibacterial agents · CPC title

• A61P1/00

  Drugs for disorders of the alimentary tract or the digestive system · CPC title

• C12P19/26

  Preparation of nitrogen-containing carbohydrates · CPC title

• A61K31/702 Primary

  Oligosaccharides, i.e. having three to five saccharide radicals attached to each other by glycosidic linkages · CPC title