Process for the production of isomaltooligosaccharides
The present invention provides a method for the production of oligosaccharides by the fermentation of dextran-sucrase-producing microorganisms with sucrose and maltose. The disclosed process allows for the control of the final composition of the isomaltooligosaccharides by adjustments to pH and the initial ratio of sucrose to maltose.
1. A method for the preparation of oligosaccharides comprising:
(a) contacting a feedstock comprising sucrose and maltose having at least a 2.33:1 starting ratio of sucrose to maltose with dextransucrase producing bacterial cells in an aqueous culture medium, where the dextransucrase is of enzyme classification (EC) 2.4.1.5;
(b) fermenting the feedstock with the bacterial cells while controlling the culture medium at a pH between 4 and 6.0 by adding an acid or a base to the culture medium;
(c) removing the bacterial cells to produce a fermentation broth; and
(d) polishing the fermentation broth to produce a final composition;
wherein the final composition of the oligosaccharides comprises maltosyl-isomaltooligosaccharides with degrees of polymerization greater than 2 (DP2).
2. The method of claim 1 , wherein performing steps (a) to (c) is continuous.
3. The method of claim 1 , wherein the method is conducted as an immobilized cell process.
4. The method of claim 1 , wherein the method is conducted as a batch or a fed-batch operation.
5. The method of claim 1 , wherein the starting ratio of sucrose to maltose ranges up to 7:1.
6. The method of claim 1 , wherein the starting ratio is 2.33:1; 3.17:1; 3.2:1; or 2.75:1.
7. The method of claim 1 , wherein the ratio of sucrose to maltose is adjusted during the fermentation process by the addition of either more sucrose or more maltose.
8. The method of claim 1 , wherein the dextransucrase-producing bacterial cells are Leuconostoc mesenteroides, Leuconostoc citreum, Leuconostoc gasicomitatum , or Leuconostoc kimchii.
9. The method of claim 8 , wherein the dextransucrase-producing bacterial cells are Leuconostoc citreum ATCC 13146 or Leuconostoc citreum NRRL B-742 or Leuconostoc mesenteroides subsp. mesenteroides NRRL B-1299.
10. The method of claim 1 , wherein the dextransucrase-producing bacterial cells are Weisella confusa, Weissella cibaria, Lactococcus spp, Pediococcus spp, Pediococcus pentosaceus, Streptococcus mutans, Streptococcus oralis, Streptococcus sanguinis, Lactobacillis spp, or Lactobacillis reuteri.
11. The method of claim 1 , wherein the base comprises an alkali earth metal hydroxide or carbonate.
12. The method of claim 1 , wherein the bacterial cells are removed by centrifugation, filtration or clarification.
13. The method of claim 1 , wherein the polishing removes insoluble impurities or comprises decolorization or comprises de-ashing or comprises removing protein or comprises removing organic acids or comprises combinations of these steps.
14. The method of claim 13 , wherein the decolorization utilizes activated charcoal or activated carbon or comprises using a weak base anion resin or comprises a combination of these steps.
15. The method of claim 13 , wherein the de-ashing comprises using a strong acid cation resin to remove metal ions.
16. The method of claim 13 , wherein the de-ashing comprises a two-step process using a strong acid cation resin followed by a weak base anion resin.
17. The method of claim 13 , wherein the removing protein comprises heating, evaporation, centrifugation, filtration, using activated carbon, using a weak base anion resin, or a combination thereof.
18. The method of claim 13 , wherein the removing the organic acids utilizes a chromatographic grade gel-type strong acid cation exchange resin in calcium form (SAC-Ca++).
19. The method of claim 13 , wherein the removing organic acids comprises utilizing a weak base anion resin.
20. The method of claim 1 , wherein the final composition of the oligosaccharides produced comprises isomaltooligosaccharides with an α-(1>4) linkage at the reducing end and α-( 1 →6) linkages with a degree of polymerization between 3 and 10 or between 3 and 9.
21. The method of claim 20 , wherein the isomaltooligosaccharides further comprise α-(1→4), α-(1→3) and/or α-(1→2) branching.
22. The method of claim 1 , further comprising providing the oligosaccharides as a concentrated solution.
23. The method of claim 1 , further comprising providing the oligosaccharides as a powder produced by drying or by spray drying or by freeze drying.
24. The method of claim 1 , wherein the aqueous culture medium comprises a filter sterilized sugar and salt stock solution.