Pomegranate, renowned for its delectable taste and remarkable nutritional profile, has witnessed a surge in both production and consumption. However, the by-products generated during industrial processes, such as peels and seeds, have the potential for adverse environmental impacts if not meticulously managed. Similarly, expired fruit juices or spillages that may occur during manufacturing and transportation contribute to agri-food waste. This study focused on the comprehensive assessment of pomegranate by-products and pomegranate juice using ascomycetes and zygomycetes filamentous fungi, namely Aspergillus oryzae, Rhizopus oligosporus, and Neurospora intermedia to obtain mycoprotein for sustainable vegan food production. The findings revealed that pomegranate juice, both fresh and expired commercial, contained essential nutrients for fungal biomass production (up to 0.024 g biomass/mL juice). Nonetheless, fresh juice emerges as a more potent medium in terms of protein production than commercial juice. Cultivating A. oryzae yielded a biomass of 0.39 (g biomass/g peel) from pomegranate peel, while concurrently raising the protein content of raw pomegranate peel from 30.89 g/kg to 85.41 g/kg. Furthermore, incorporating yeast extract into the peel medium not only resulted in an enhanced biomass yield of 0.49 (g biomass/g peel) but also significantly elevated the protein content to 198.63 g/kg. This study provides valuable insights into the potential of pomegranate peel and juice as promising substrate for fungal biomass production, offering opportunities for the development of innovative food and feed products.