Comprehensive Assessment of Functional Effects of Commonly Used Sugar Substitute Sweeteners on Ex Vivo Human Gut Microbiome

The composition and function of the human gut microbiome are often associated with health and disease status. Sugar substitute sweeteners are widely used food additives, although many studies using animal models have linked sweetener consumption to gut microbial changes and health issues. Whether sugar substitute sweeteners directly change the human gut microbiome functionality remains largely unknown. In this study, we systematically investigated the responses of five human gut microbiomes to 21 common sugar substitute sweeteners, using an approach combining high-throughput in vitro microbiome culturing and metaproteomic analyses to quantify functional changes in different taxa. Hierarchical clustering based on metaproteomic responses of individual microbiomes resulted in two clusters. The noncaloric artificial sweetener (NAS) cluster was composed of NASs and two sugar alcohols with shorter carbon backbones (4 or 5 carbon atoms), and the carbohydrate (CHO) cluster was composed of the remaining sugar alcohols. The metaproteomic functional responses of the CHO cluster were clustered with those of the prebiotics fructooligosaccharides and kestose. The sugar substitute sweeteners in the CHO cluster showed the ability to modulate the metabolism of Clostridia. This study provides a comprehensive evaluation of the direct effects of commonly used sugar substitute sweeteners on the functions of the human gut microbiome using a functional metaproteomic approach, improving our understanding of the roles of sugar substitute sweeteners on microbiome-associated human health and disease issues. IMPORTANCE The human gut microbiome is closely related to human health. Sugar substitute sweeteners as commonly used food additives are increasingly consumed and have potential impacts on microbiome functionality. Although many studies have evaluated the effects of a few sweeteners on gut microbiomes using animal models, the direct effect of sugar substitute sweeteners on the human gut microbiome remains largely unknown. Our results revealed that the sweetener-induced metaproteomic responses of individual microbiomes had two major patterns, which were associated with the chemical properties of the sweeteners. This study provided a comprehensive evaluation of the effects of commonly used sugar substitute sweeteners on the human gut microbiome.