Rheoliad 3(4)

ATODLEN 2Diwygio’r Atodiad i Reoliad (EU) Rhif 231/2012 ar gyfer ychwanegu manyleb ar gyfer E 960b glycosidau stefiol o eplesu (Yarrowia lipolytica)

1.  Yn Rheoliad y Comisiwn (EU) Rhif 231/2012, mae’r Atodiad (manylebau ar gyfer ychwanegion bwyd) wedi ei ddiwygio fel a ganlyn.

2.  Ar ôl y cofnod ar gyfer “E 960a” (glycosidau stefiol o Stevia), mewnosoder y cofnod a ganlyn—

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 AC 44H 70O 230.33
Rebaudioside BC 38H 60O 180.40
Rebaudioside DC 50H 80O 280.29
Rebaudioside MC 56H 90O 330.25
Molecular weight and CAS numberTrivial 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
Assay Not less than 95% of rebaudioside M, rebaudioside D, rebaudioside A, and rebaudioside B 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)
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 0.1 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.