Melanoidins (MLDs), the final products of the Maillard reaction, are high molecular-weight polymers formed during the processing of black garlic. These compounds contribute to the distinctive color, flavor, and bioactive properties of black garlic. However, the impact of storage conditions on the structural integrity and bioactivities of black garlic melanoidins (BG-MLDs) remains largely unexplored. By using various in vitro simulations, this study investigated the effects of different storage conditions (at 4 and 35 ℃ for 0, 3, and 6 months) on the digestion, absorption, metabolism, and intestinal microbial profile of BG-MLDs during in vitro gastrointestinal digestion. Our results demonstrated that BG-MLDs resisted digestion regardless of storage conditions. BG-MLDs stored at 4 ℃ for 3 months significantly enriched the abundance of beneficial bacteria like Bifidobacterium longum and decreased harmful bacteria like Escherichia. Additionally, MLD treatment significantly increased the production of short-chain fatty acids (SCFAs), particularly propionate, butyrate, and isovalerate, with the 35 ℃ for 3 months of storage showing the most significant effect (increased by 3.74-fold). However, a longer storage at 35 ℃ (6 months) showed a decline in SCFA production. These findings highlight the critical role of a certain storage time (3 months) in preserving and potentially enhancing the prebiotic activity of BG-MLDs. By optimizing storage parameters, the bioactive potential of BG-MLDs can be used for their potential application as functional food ingredients or nutraceuticals, contributing to the prevention and management of chronic diseases associated with gut dysbiosis.