<p>Stevia is a sweetener found in the leaves of the plant <i>Stevia rebaudiana</i> Bertoni. It is 50 to 350 times sweeter than sugar and has almost no calories. α-1,6 glucosylated steviol glycosides (GSG) is a novel stevia with a unique chemical structure and improved sensory properties. Considering their potential application in various foods and beverages, we conducted in vitro metabolism study to assess the digestion safety of GSG. In the simulated gastric fluid and simulated intestinal fluid assays, there weren’t significant changes in GSG and steviol glycosides (SG). The anaerobic metabolism study using human fecal homogenates showed complete decomposition of GSG into steviol, similar to common SG such as rebaudioside A, rebaudioside M, and stevia extract, a SG mixture extracted from <i>Stevia rebaudiana</i> Bertoni. Overall, the results of the in vitro digestion study show that ingested GSGs can be digested similar to other SG that have established safety for consumption.</p> Graphical Abstract <p></p>