Extraction, separation and recovery of diterpene glycosides from Stevia rebaudiana plants

A process for recovering diterpene glycosides from Stevia rebaudiana plant material, said process comprising:

• sequentially extracting Stevia rebaudiana plant material with a first solvent comprising a liquid haloalkane of from one to four carbon atoms to remove impurities therefrom in a first extract substantially free of diterpene glycosides, and then with a second solvent comprising an alkanol of from one to three carbon atoms to give a second extract containing diterpene glycosides;
• chromatographing the resulting extracted diterpene glycosides on a liquid chromatography column having a packing comprising an ether oxygen-containing organic stationary phase covalently bonded through a silicon atom to a silica gel support;
• said extracted glycosides being introduced onto said column for chromatography thereon in solution in said second solvent or in another solvent having a high polarity not substantially greater than the polarity of the second solvent;
• said column being eluted with an alkanol of from two to four carbon atoms;
• and
• collecting individually eluate fractions rich in the respective diterpene glycosides.

A process for recovering diterpene glycosides from Stevia rebaudiana plant material, said process comprising:

• sequentially extracting Stevia rebaudiana leaves with chloroform to remove impurities therefrom in a first extract substantially free of diterpene glycosides, and then with methanol to give a second extract containing diterpene glycosides;
• chromatographing the resulting extracted diterpene glycosides on a liquid chromatography column having a packing comprising a glycerylpropyl moiety stationary phase covalently bonded through a silicon atom to a silica gel support having a mean particle size diameter of about 5-15 microns and a mean surface area of about 100-500 square meters per gram;
• said extracted glycosides being introduced onto said column for chromatography thereon in solution in a solvent comprised of 95-100% by volume of 1-propanol and the balance substantially water;
• said column being eluted with 1-propanol;
• and
• collecting individually eluate fractions rich in respective diterpene glycosides.

A process for separating a mixture of diterpene glycosides of the Stevia rebaudiana plant, said process comprising:

• introducing a solution of said mixture of diterpene glycosides in a solvent onto a liquid chromatography column, said column having a packing comprising an oxygen-containing organic stationary phase covalently bonded through a silicon atom to an inorganic support;
• said solvent comprising 95-100% by volume of an alkanol of from one to four carbon atoms and the balance substantially water;
• eluting said column with an alkanol of from two to four carbon atoms;
• and
• collecting individually eluate fractions rich in respective diterpene glycosides.