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The Food Additives and Novel Foods (Authorisations and Miscellaneous Amendments) and Food Flavourings (Removal of Authorisations) (Scotland) Regulations 2024

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E 960C(ii) REBAUDIOSIDE M, AM AND D PRODUCED VIA ENZYMATIC CONVERSION OF HIGHLY PURIFIED STEVIOL GLYCOSIDES FROM STEVIA LEAF EXTRACTS

Synonyms
Definition

Steviol glycosides produced via enzymatic conversion of highly purified steviol glycosides (rebaudioside A or stevioside) from Stevia leaf extracts are composed predominantly of rebaudioside M, rebaudioside D, and rebaudioside AM.

Rebaudiosides D, M and AM are produced via enzymatic conversion of highly purified steviol glycoside (rebaudioside A or stevioside) extracts (95% steviol glycosides) obtained from Stevia rebaudiana Bertoni plant using UDP-glucosyltransferase and sucrose synthase enzymes produced by genetically modified strains of Escherichia coli (pPM294, pFAH170, and pSK041) that facilitate the transfer of glucose from sucrose and UDP-glucose to steviol glycosides via glycosidic bonds. After removal of the enzymes by solid-liquid separation and heat treatment, the purification involves concentration of the steviol glycosides by resin adsorption, followed by recrystallisation of the steviol glycosides resulting in a final product containing not less than 95 % of total steviol glycosides, including one or more of rebaudiosides D, M and AM.

Viable cells or DNA of Escherichia coli (pPM294, pFAH170, and pSK041) must not be detected in the food additive.

Chemical name

Rebaudioside M: 13-[(2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl)oxy]kaur-16-en-18-oic acid, 2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl ester

Rebaudioside D: 13-[(2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl)oxy]kaur-16-en-18-oic acid, 2-O-β-D-glucopyranosyl-β-D-glucopyranosyl ester

Rebaudioside AM: 13-[(2-O-β-D-glucopyranosyl-β-D-glucopyranosyl)oxy]kaur-16-en-18-oic acid, 2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl ester

Molecular formulaTrivial nameFormulaConversion factor
Rebaudioside MC56 H90 O330.25
Rebaudioside DC50 H80 O280.29
Rebaudioside AMC50 H80 O280.29
Molecular weight and CAS NoTrivial nameCAS NumberMolecular weight (g/mol)
Rebaudioside M1220616-44-31291.29
Rebaudioside D63279-13-01129.15
Rebaudioside AM2222580-26-71129.15
AssayNot less than 95 % of steviol glycosides on the dried basis, including one or more of rebaudiosides D, M and AM
DescriptionWhite to light yellow powder, approximately between 200 and 350 times sweeter than sucrose (at 5 % sucrose equivalency)
Identification
SolubilityFreely soluble to slightly soluble in water
pHBetween 4.5 and 7.0 (1 in 100 solution)
Purity
Total ashNot more than 1 %
Loss on dryingNot more than 6 % (105°C, 2h)
Residual solventNot more than 5000 mg/kg ethanol
ArsenicNot more than 0.015 mg/kg
LeadNot more than 0.2 mg/kg
CadmiumNot more than 0.015 mg/kg
MercuryNot more than 0.07 mg/kg
Residual proteinNot more than 5 mg/kg.

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