How sweet it is: The next generation of natural, zero-calorie sweeteners

But it’s one thing to identify a superior rebaudioside. It’s quite another to turn it into a product — and a pure one at that.

Combining ancient processes with modern biotechnology

Conagen leverages fermentation — an ancient microbial process — to manufacture their suite of products, currently around 35. Through their integrated manufacturing chain, the team can move from design to high-throughput screening and fermentation rather quickly — a useful capability when it comes to producing Reb M.

“We do an enhanced bioconversion,” says Mattozzi, “wherein we take the extract from the leaf, and through enzymatic conversions we push what starts as a trace amount of Reb M to over 95% purity.”

Put that way, the process sounds simple. But it was only possible due to the team’s ability to test hundreds of different enzymatic processes in a high-throughput way. Why hundreds?

A critical characteristic of all rebaudiosides is the number of glycosyl groups found on the stevial core of the molecule, which affect the taste profile of the rebaudioside. Reb M has six glycosyl groups; several other rebaudiosides — Reb E, Reb V, Reb D, and Reb I, to name a few — have four or five glycosyl groups attached, making them potential Reb M precursors, and flavoring enhancers and sweeteners in their own right. How those groups are oriented and sequentially added to the steviol backbone yields hundreds of different combinations — all of which the Conagen team tested for the desired output: the best flavor. They then optimized growth conditions for each bio-conversion process.

“This has allowed us to access the mixture of different steviol glycosides that are already in the natural extract to converge them down on to Reb M,” explains Mattozzi. “Synthetic biology at work.”

The result? A commercial product called BESTEVIA® Reb M stevia leaf sweetener.

Third-generation natural sweeteners?

While the Reb M sweetener is a novel, improved solution, the Conagen team isn’t stopping there. They have identified a number of trace compounds in the stevia leaf extract that actually have more glycosyl groups — seven and even eight — than Reb M, which should make them sweeter and potentially give them an even more palatable taste profile than Reb M. Mattozzi sees these more complex glycosides as potential third-generation natural sweeteners within Conagen’s grasp based on their success making Reb M.

Regardless of what the future holds for the next generation of natural sweeteners, Conagen is in a good position to lead the way. As Mattozzi puts it, “Our knowledge on plant biochemistry, strain development, and fermentation allowed us to develop this process. However, it was our experience of scaling up processes that enabled us to develop a commercial product like BESTEVIA® Reb M.”

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