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Extraction of dyes from walnut using Soxhlet apparatus has been studied. The color components extracted and isolated from walnut shells were characterized by column chromatography, thin layer chromatography (TLC), nuclear magnetic resonance (NMR), mass spectroscopy (MS), and infrared (IR) techniques. Natural dye extract obtained from the walnut was used in dyeing polyamide fabrics with different mordants. The dyed fabrics were evaluated for antibacterial activity against pathogenic strains of Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria. As such, the relationship between antibacterial activity and dye concentration is investigated. Durability of antibacterial activity to laundering is also discussed. Results indicate that the polyamide dyed with walnut displayed excellent antibacterial activity in the presence of ferric sulfate, cupric sulfate, and potassium aluminum sulfate and exhibited good and durable fastness properties.