High-Purity Steviol Glycosides

a technology of steviol glycosides and steviol, which is applied in the direction of transferases, tobacco, tobacco treatment, etc., can solve the problems of unsuitable commercial use methods

Inactive Publication Date: 2016-07-14
PURECIRCLE SDN BHD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although methods are known for preparing steviol glycosides from Stevia rebaudiana, many of these methods are unsuitable for use commercially.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

In-Vivo Production of UGT76G1

[0285]NcoI and NdeI restriction sides were added to the original nucleic sequence as described in Genbank accession no. AAR06912.1. After codon optimization the following nucleic sequence was obtained:

(SEQ ID NO: 1)CCATGGCCCATATGGAAAACAAAACCGAAACCACCGTTCGTCGTCGTCGCCGTATTATTCTGTTTCCGGTTCCGTTTCAGGGTCATATTAATCCGATTCTGCAGCTGGCAAATGTGCTGTATAGCAAAGGTTTTAGCATTACCATTTTTCATACCAATTTTAACAAACCGAAAACCAGCAATTATCCGCATTTTACCTTTCGCTTTATTCTGGATAATGATCCGCAGGATGAACGCATTAGCAATCTGCCGACACATGGTCCGCTGGCAGGTATGCGTATTCCGATTATTAACGAACATGGTGCAGATGAACTGCGTCGTGAACTGGAACTGCTGATGCTGGCAAGCGAAGAAGATGAAGAAGTTAGCTGTCTGATTACCGATGCACTGTGGTATTTTGCACAGAGCGTTGCAGATAGCCTGAATCTGCGTCGTCTGGTTCTGATGACCAGCAGCCTGTTTAACTTTCATGCACATGTTAGCCTGCCGCAGTTTGATGAACTGGGTTATCTGGATCCGGATGATAAAACCCGTCTGGAAGAACAGGCAAGCGGTTTTCCGATGCTGAAAGTGAAAGATATCAAAAGCGCCTATAGCAATTGGCAGATTCTGAAAGAAATTCTGGGCAAAATGATTAAACAGACCAAAGCAAGCAGCGGTGTTATTTGGAATAGCTTTAAAGAACTGGAAGAAAGCGAACTGGAAACCGTGATTCGTGAAATTCCGGCACCGAGCTTTCTGATTCCGCTGCCGA...

example 2

In-Vitro Production of UGT76G1

[0290]The S30 T7 High Yield Protein expression system kit from Promega was used. 4 μg of UGT76G1_pET30a+ plasmid from E. coli EC100 was mixed with 80 μL of S30 premix plus and 72 μL of S30 T7 extract was added. Nuclease-free water was added in order to obtain a total volume of 200 μL and the resulting solution was incubated for 2 h at 30° C. 180 μL was used in the catalytic test reaction.

example 3

In-Vitro Production of UGT91D2

[0291]NcoI and NdeI restriction sides were added to the original nucleic sequence as described in Genbank accession no. ACE87855.1. After codon optimization the following nucleic sequence was obtained:

(SEQ ID NO: 2)CCATGGCACATATGGCAACCAGCGATAGCATTGTTGATGATCGTAAACAGCTGCATGTTGCAACCTTTCCGTGGCTGGCATTTGGTCATATTCTGCCGTATCTGCAGCTGAGCAAACTGATTGCAGAAAAAGGTCATAAAGTGAGCTTTCTGAGCACCACCCGTAATATTCAGCGTCTGAGCAGCCATATTAGTCCGCTGATTAATGTTGTTCAGCTGACCCTGCCTCGTGTTCAAGAACTGCCGGAAGATGCCGAAGCAACCACCGATGTTCATCCGGAAGATATTCCGTATCTGAAAAAAGCAAGTGATGGTCTGCAGCCGGAAGTTACCCGTTTTCTGGAACAGCATAGTCCGGATTGGATCATCTATGATTATACCCATTATTGGCTGCCGAGCATTGCAGCAAGCCTGGGTATTAGCCGTGCACATTTTAGCGTTACCACCCCGTGGGCAATTGCATATATGGGTCCGAGCGCAGATGCAATGATTAATGGTAGTGATGGTCGTACCACCGTTGAAGATCTGACCACCCCTCCGAAATGGTTTCCGTTTCCGACCAAAGTTTGTTGGCGTAAACATGATCTGGCACGTCTGGTTCCGTATAAAGCACCGGGTATTAGTGATGGTTATCGTATGGGTCTGGTTCTGAAAGGTAGCGATTGTCTGCTGAGCAAATGCTATCATGAATTTGGCACCCAGTGGCTGCCGCTGCTGGAAACCCTGCATCAGGTTCCGGTTGTTCCGGTGGGT...

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Abstract

Methods of preparing highly purified steviol glycosides, particularly rebaudiosides A, D and M are described. The methods include utilizing recombinant microorganisms for converting various staring compositions to target steviol glycosides. In addition, novel steviol glycosides reb D2 and reb M2 are disclosed, as are methods of preparing the same. 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.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application claims priority to U.S. Provisional Patent Application No. 61 / 827,922, filed May 28, 2013; U.S. Provisional Patent Application No. 61 / 843,544, filed Jul. 8, 2013; U.S. Provisional Patent Application No. 61 / 861,528, filed Aug. 2, 2013; U.S. Provisional Patent Application No. 61 / 881,166, filed Sep. 23, 2013; U.S. Provisional Patent Application No. 61 / 885,084, filed Oct. 1, 2013; U.S. Provisional Patent Application No. 61 / 904,751, filed Nov. 15, 2013; U.S. Provisional Patent Application No. 61 / 913,482, filed Dec. 9, 2013; U.S. Provisional Patent Application No. 61 / 921,635, filed Dec. 30, 2013; U.S. Provisional Patent Application No. 61 / 925,329, filed Jan. 9, 2014 and U.S. Provisional Patent Application No. 61 / 939,855, filed Feb. 14, 2014. The above-referenced priority documents are incorporated fully herein in their entirety.TECHNICAL FIELD[0002]The present invention relates to a biocatalytic process for preparing compositio...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A23L1/236A23L2/60C07H15/256C12P19/56A23L27/30
CPCA23L1/2363C07H15/256A23V2002/00A23L2/60C12P19/56C07H1/06C12N9/10A24B15/302A23L27/33Y02P20/582A23V2250/258A24B15/10
Inventor PRAKASH, INDRABUNDERS, CYNTHIASONI, PANKAJMARKOSYAN, AVETIKCYRILLE, JARRINBADIE, AURELIENHALLE, ROBER TER
Owner PURECIRCLE SDN BHD
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