Safety of the proposed amendment of the specifications for enzymatically produced steviol glycosides (E 960c): Rebaudioside D produced via enzymatic bioconversion of purified stevia leaf extract

Abstract

The EFSA Panel on Food Additives and Flavourings (FAF Panel) provides a scientific opinion on the safety of a proposed amendment of the specifications of enzymatically produced steviol glycosides (E 960c) with respect to the inclusion of rebaudioside D produced via enzyme‐catalysed bioconversion of purified stevia leaf extract. Rebaudioside D (95% on dry basis) is produced via enzymatic bioconversion of purified stevia leaf extract using uridine diphosphate (UDP)‐glucosyltransferase (UGT) and sucrose synthase enzymes produced by the genetically modified yeast K. phaffii UGT‐A, that facilitates the transfer of glucose to purified stevia leaf extract via glycosidic bonds. The same enzymes from K. phaffii UGT‐A may be used in the manufacturing process of the food additive, rebaudioside M produced via enzyme modification of steviol glycosides from stevia (E 960c(i)). The Panel considered that separate specifications would be needed for this food additive produced via the manufacturing process described in the current application, aligned with those already established for E 960c(i). The Panel concluded that there is no toxicological concern for Rebaudioside D produced via enzymatic bioconversion of purified stevia leaf extract using UDP‐glucosyltransferase and sucrose synthase produced by a genetically modified strain of the yeast K. phaffii. However, based on the available data, the Panel could not exclude the possibility that some residual amount of DNA coding for the kanamycin resistance gene could remain in the final product. Should this gene propagate in microbiota due to the presence of recombinant DNA in the final product, this would be of concern. Therefore, the Panel concluded that the safety of Rebaudioside D produced via this enzymatic bioconversion was not sufficiently demonstrated with the available data given that the absence of recombinant DNA was not shown.

Safety evaluation of the food additive steviol glycosides, predominantly Rebaudioside M, produced by fermentation using Yarrowia lipolytica VRM

“…The authors concluded that the metabolism of the test item indicated a rapid deglycosylation to a final steviol metabolite. These results are consistent with those previously considered in other scientific opinions of the Panel (EFSA FAF Panel, 2019, 2022.…”

“…The Panel considered that the metabolic fate of steviol glycosides, including steviol glycosides obtained via fermentation, leads to the aglycone which is absorbed. Given that all steviol glycosides follow the same metabolic pathways, the Panel considered that the current steviol glycosides would fall within the same group of substances (EFSA ANS Panel, 2010 ; EFSA FAF Panel, 2020 , 2022 ), and the group approach would be applicable. Therefore, the Panel considered that the already existing data on rebaudioside M and structural‐related steviol glycosides (EFSA ANS Panel, 2010 ; EFSA FAF Panel, 2019 , 2020 , 2021 , 2022 ), along with new supportive toxicity data on rebaudioside A produced fermentatively by Y. lipolytica (same manufacturing process of the proposed food additive), are sufficient.…”

“…For the other toxicity endpoints, no new studies were performed with the food additive, and no new studies relevant for the risk assessment were submitted by the applicant. Nonetheless, the Panel is of the opinion that a read‐across with regard to toxicity is applicable, considering the availability of toxicity studies on other previously evaluated steviol glycosides (EFSA ANS Panel, 2010 ; EFSA FAF Panel, 2020 , 2022 ). Therefore, no additional toxicity studies are required.…”

“…Data on ADME of some of the steviol glycosides currently listed in the EU specifications have been considered and summarised in previous EFSA opinions (EFSA ANS Panel, 2010 , 2015a ; EFSA FAF Panel, 2019 , 2020 , 2021 , 2022 ). According to these opinions, steviosides and steviol glycosides are not hydrolysed by digestive enzymes of the upper gastrointestinal tract due to the presence of ß‐glycosidic bonds.…”

“…In 2022, a new opinion on the safety of an additional proposed amendment to the specifications of the food additive steviol glycosides (E 960) was published and regarded rebaudioside D produced by enzymatic bioconversion of purified S. rebaudiana Bertoni leaf extract, using UDP glucosyltransferase (UGT) and sucrose synthase produced by a genetically modified strain of the yeast K. phaffii (EFSA FAF Panel, 2022 ).…”