High-purity steviol glycosides
Methods of preparing highly purified steviol glycosides, particularly rebaudiosides M, D, E and I are described. The methods include utilizing enzyme preparations and recombinant microorganisms for converting various staring compositions to target steviol glycosides. The highly purified rebaudiosides are useful as non-caloric sweetener in edible and chewable compositions such as any beverages, confectioneries, bakery products, cookies, and chewing gums.
1 . A method for producing rebaudioside M comprising the steps of:
a. providing a starting composition comprising rebaudioside A;
b. providing an enzyme preparation or microorganism containing the enzymes: UGTSr of SEQ ID NO: 3 or a mutant variant thereof having >98% amino-acid sequence identity, and SuSy_At of SEQ ID NO: 1 and UGTS1 of SEQ ID NO: 2;
c. contacting the enzyme preparation or microorganism with a medium containing the starting composition to produce a medium comprising at least rebaudioside M.
2 . The method of claim 1 further comprising the step of:
d. separating the rebaudioside M from the medium to provide a highly purified rebaudioside M.
3 . The method of claim 2 , wherein the rebaudioside M content is greater than 95% by weight on a dry basis.
4 . The method of claim 1 , wherein the microorganism is selected from the group consisting of E. coli, Saccharomyces sp., Aspergillus sp., Pichia sp., Bacillus sp., and Yarrowia sp.