Due to its abundance of physiologically active ingredients, one of the oldest medicinal herbs, elderberry (EB) L., is beneficial for both therapeutic and dietary purposes. This study determined the bioaccessibility of the phenolic compounds and the prebiotic potential of the polyphenols from freeze-dried EB powder (FDEBP), along with the antioxidant and antimicrobial activities of this extract. The most significant phenolic compounds in black EB are represented by anthocyanins (41.8%), predominating cyanidin-sambubiosides and cyanidin-glucosides (90.1% of the identified anthocyanins). The FRAP assay obtained the highest antioxidant activity value (185 ± 0.18 μmol Fe/g DW). The most sensitive to the antimicrobial activity of the extract was proven to be , and had the lowest minimum inhibitory concentration of 1.95 mg/mL. To determine the prebiotic potential of the polyphenols, the cell growth of five probiotic strains (, , , and ) was tested. The influence on cell growth was positive for all five probiotic strains used. Overall, the most significant increase ( < 0.05) was recorded at 1.5% FDEBP, on with a growth index (GI) of 152.44%, very closely followed by GI at 0.5% and 1% concentrations. The stability of the total phenolic compounds through simulated gastronitestinal digestion was increased (93%), and the bioaccessibility was also elevated (75%).