Methods for providing sialylated oligosaccharides

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.