Dye wastewater containing non-fixed dyes discarded from different manufacturing industries is a major concern in environmental pollution. Amongst all other non-fixed dyes, anionic dyes hold a significant share in the dye wastewater (32–90%) stream, due to their extensive uses. In this study, cotton gin trash (CGT) is proposed for valorisation and utilisation as a bioadsorbent for the anionic dye. Gin trash was transformed into a film by a single-step process. Since −OH group rich CGT film tends to adsorb cationic dye, chitosan that has adsorption capability towards anionic dyes was used to modify CGT by introducing positive charges for the adsorption of anionic Acid Blue 25 (AB). The morphology, roughness, chemical structure and zeta potential of the raw CGT powder and chitosan-modified CGT (CHT–CGT) film were reported. The fabricated film showed roughness and pores in the surface favouring the dye adsorption. The adsorption process followed the physisorption phenomenon rather than the chemisorption process, where cationic CHT–CGT film attracted anionic AB. Kinetics and equilibrium adsorption of the system were described as favourable, fitting better with the Langmuir model compared to Freundlich, Dubinin–Radushkevich and Flory–Huggins isotherms. The maximum adsorption of the CHT–CGT film was 151.5 mg/g, compares favourably among other reported lignocellulosic waste. Besides, CHT–CGT film was found reusable after desorption, without significantly altering its removal efficiency. The results along with our previous report explore a sustainable pathway of adding value to CGT as a dye bioadsorbent from wastewater, where unmodified and chitosan-modified CGT films together have the potential to separate both cationic and anionic dyes concurrently.