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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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1.  In the appropriate place, insert the following entry—

“E 960b STEVIOL GLYCOSIDES FROM FERMENTATION (YARROWIA LIPOLYTICA)

Synonyms
Definition

Steviol glycosides from Yarrowia lipolytica consist of a mixture predominantly composed of rebaudioside M, with some rebaudioside D, and smaller amounts of rebaudioside A and rebaudioside B. The manufacturing process comprises two main phases.

The first phase involves fermentation of a non-toxigenic non-pathogenic strain of Yarrowia lipolytica VRM that has been genetically modified with heterologous genes to overexpress steviol glycosides. Removal of biomass by solid-liquid separation and heat treatment is followed by concentration of the steviol glycosides.

The second phase involves purification by employing ion-exchange chromatography, followed by recrystallisation of the steviol glycosides resulting in a final product containing not less than 95% of rebaudiosides M, D, A, and B.

Viable cells or the DNA of Yarrowia lipolytica VRM must not be detected in the food additive.

Chemical name

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

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

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 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

Molecular formulaTrivial nameFormulaConversion factor
Rebaudioside AC44 H70 O230.33
Rebaudioside BC38 H60 O180.40
Rebaudioside DC50 H80 O280.29
Rebaudioside MC56 H90 O330.25
Molecular weight and CAS NoTrivial nameCAS NumberMolecular weight (g/mol)
Rebaudioside A58543-16-1967.01
Rebaudioside B58543-17-2804.88
Rebaudioside D63279-13-01129.15
Rebaudioside M1220616-44-31291.29
AssayNot less than 95 % of rebaudioside A, rebaudioside B, rebaudioside D and rebaudioside M on the dried basis
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)
hPurity
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 0.1 mg/kg
LeadNot more than 0.1 mg/kg
CadmiumNot more than 0.01 mg/kg
MercuryNot more than 0.05 mg/kg
Residual proteinNot more than 20 mg/kg
Microbiological criteria
Total (aerobic) plate countNot more than 1000 CFU/g
YeastNot more than 100 CFU/g
MouldsNot more than 100 CFU/g
Escherichia coliNegative in 1g
Salmonella spp.Negative in 25g”.

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