The human gut microbiota plays a vital role in host health by acting as a barrier against pathogens, boosting the immune system, and metabolizing complex carbon sources into beneficial compounds such as short-chain fatty acids (SCFAs). However, external factors like diet and xenobiotics, including food colorants, can impact this microbial community. Brilliant blue FCF (E-133), a common food dye that is not absorbed or metabolized by the body, thus, leading to significant exposure of the gut microbiota, and its effects on it are not well-documented. This study investigated the impact of brilliant blue on the simplified human gut microbiota model (SIHUMIx) over a seven-day exposure period, followed by a four-day recovery phase. Using continuous culture bioreactors along with metaproteomic and metabolomic analyses, the research uncovered substantial structural and functional changes in the microbiota. Our findings showed alterations in the species abundance such as B. thetaiotaomicron, B. longum, and C. butyricum, reductions in energy metabolism by-products, particularly lactate and butyrate, and disruptions in metabolic pathways involved in the metabolism of xenobiotics and amino acids. The results indicate that brilliant blue exposure compromises microbiota stability and functionality, highlighting the need for further research on its long-term effects and recovery mechanisms. This study underscores the importance of assessing the impacts of food additives on gut microbiota as part of comprehensive health risk evaluations.