Nowadays, cotton fabric treatment with nanoparticles has been increasingly high because of its unique properties. In this study, using the bioactive compounds in Psidium guava leave extracts, zinc precursors were reduced to obtain ZnO nanoparticles using two extracts, namely, water extract and ethanol extract. A comparative evaluation was carried out to analyze and develop a wider understanding of the effect of processing parameters on the ZnO nanoparticles. The ZnO nanoparticles have been characterized using a particle size analyzer, Transmission Electronic Microscopy (TEM), X-ray diffraction, and Thermal Gravimetrical Analysis (TGA) to determine their structure. The cotton fabric was treated with ZnO nanoparticles in the presence and absence of different natural biopolymers, namely; alginate, chitosan, and carboxymethyl cellulose (CMC). The treated cotton samples were characterized by atomic absorption and scanning electron microscopy (SEM), UPF value, antimicrobial activity, and self-cleaning properties. It was found that the cotton fabric treated with ZnO-chitosan nanocomposites gives the higher UPF values and antimicrobial activity against Staphylococcus aureus, Escherichia coli, and Candida albicans of all other treated fabrics.