The global textile dyeing industry releases approximately 200,000 tonnes of synthetic dyes into waterways annually, representing a major source of persistent organic pollutant contamination with documented endocrine-disrupting, carcinogenic, and mutagenic effects on aquatic ecosystems and human health. The transition from synthetic reactive dyeing to natural dyeing processes using plant-derived colorants offers environmental sustainability benefits but faces persistent technical challenges: natural dyes' inherent low substantivity to cotton fibres requires mordanting to achieve commercially acceptable colour strength and fastness properties, and the durability of natural dyed textiles has historically been inferior to synthetic dyed equivalents.This study investigates the enhancement of natural dyeing performance on pre-scoured mercerised cotton fabric using nano-mordants — nano-titanium dioxide (nano-TiO₂) and nano-zinc oxide (nano-ZnO) — relative to conventional mordants (alum, iron, tannin) across three natural dye sources (indigo from Indigofera tinctoria, marigold from Tagetes erecta, and pomegranate rind). Box-Behnken Response Surface Methodology (RSM) optimises four process variables — pH, temperature, time, and material:liquor ratio — to maximise colour strength (K/S value). Multifunctional performance evaluation includes wash, light, and rubbing fastness (ISO 105 series), and antimicrobial activity (AATCC 100) against Staphylococcus aureus and Escherichia coli. Life cycle assessment (LCA) following ISO 14040/44 quantifies and compares GWP, cumulative energy demand (CED), and freshwater eutrophication potential against standard reactive synthetic dyeing. The nano-TiO₂ + tannin combined mordanting strategy achieves K/S of 19.1 (pomegranate) — exceeding industry minimum of 12 — with wash fastness 4-5, light fastness 4, and >99.1% antimicrobial reduction, while reducing GWP by 67.5% relative to reactive synthetic dyeing.