Simple method for the purification of lacto-N-neotetraose (LNnT) from carbohydrates obtained by microbial fermentation

A method for purification of lacto-N-neotetraose from other carbohydrates, wherein the lacto-N-neotetraose is produced by microbial fermentation and wherein the method comprises:

• a) separating biomass from a microbial fermentation broth comprising lacto-N-neotetraose to obtain a cell-free fermentation broth comprising lacto-N-neotetraose;
• b) subjecting the cell-free fermentation broth to ion-exchange resin treatment to remove charged material, wherein the cell-free fermentation broth is treated with a cation exchange resin and an anion exchange resin to obtain an aqueous solution comprising lacto-N-neotetraose;
• c) subjecting the aqueous solution comprising lacto-N-neotetraose obtained in step b) to:
• i) a first membrane filtration step to remove carbohydrates having a molecular weight smaller than that of lacto-N-neotetraose, wherein the first membrane is selected from a molecular weight cut-off of between about 300 and about 500 Dalton;
• ii) a second membrane filtration step to remove carbohydrates having a molecular weight larger than that of lacto-N-neotetraose, wherein the second membrane is selected from a molecular weight cut-off of between about 600 and about 800 Dalton, and
• d) a step for increasing the concentration of the lacto-N-neotetraose present in the filtrate of step c).