Chondroitin sulphate purification method
The present invention relates to a downstream industrial method of purifying chondroitin sulphate obtained from animal cartilage, which produces a product fully compliant with the specifications required for the use of said compound in the pharmaceutical field.
1. A method for the separation of chondroitin sulphate (CS) and keratan sulphate (KS) from an aqueous solution containing them, comprising adding to the aqueous solution a water-miscible organic solvent under conditions of ionic strength, which induces formation of two liquid phases with an interface in-between: a dense phase in which CS concentrates, said dense phase primarily comprising said water-miscible organic solvent and a light phase in which KS concentrates, said light phase primarily comprising water, wherein said water-miscible organic solvents are selected from the group consisting of 2-propanol, 1-propanol, 2-propanol/2-butanol, ethanol and acetone.
2. Method according to claim 1 comprising:
a) preparing an aqueous solution of CS/KS and determining its conductivity;
b) optionally adding a salt able to dissociate in mono- or polyvalent ions to a pre-set conductivity value;
c) gradually adding an amount of one or more of said water-miscible organic solvents so as to induce a phase separation with concentration of CS in the dense phase and KS in the light phase.
3. Method according to claim 2 wherein the salt optionally added is sodium chloride.
4. Method according to claim 1 wherein the percentage by volume of each specific water-miscible organic solvent is determined by a graph correlating said percentage to the measured conductivity value, of the aqueous solution, or to the concentration of CS and KS, in the aqueous solution.
5. Method according to claim 1 wherein the dense phase containing CS is subjected to a further purification process.
6. Method according to claim 5 wherein the dense phase is diluted with water, thereafter adding said water-miscible organic solvent until the re-formation of two liquid phases: a dense phase in which CS concentrates and a light phase in which KS concentrates.
7. Method according to claim 1 , wherein CS and KS are respectively recovered from the two liquid phases by precipitating CS with two volumes of ethanol and drying the precipitate, and precipitating KS by adding sodium chloride and drying the precipitate.