Sustainable production and antibacterial efficacy of silver nanoparticles on cellulose nanofibers from mushroom waste

Underutilized agricultural wastes, such as spent mushroom substrate (SMS), present valuable opportunities for developing sustainable biomedical materials. In this study, cellulose nanofibers (CNFs) were successfully isolated from SMS through a chemo-mechanical process, while the water extract of SMS (WESMS) served as a green reducing agent for the simultaneous synthesis and <i>in situ</i> loading of silver nanoparticles (AgNPs) onto TEMPO-oxidized CNFs (AgNP/ToCNF). The chemical structure of the isolated cellulose was characterized using ATR-FTIR, while UV-vis spectroscopy confirmed the successful synthesis and AgNPs loading, showing a maximum absorbance at 424 nm. The resulting hybrid nanomaterial exhibited a nanofiber width diameter range of 273.5-318.5 nm, while the AgNPs had an average diameter of 34.04 nm. The antimicrobial efficacy of AgNP/ToCNF was evaluated against <i>S. aureus</i>, <i>P. aeruginosa</i>, and <i>E. coli</i> using agar well diffusion, broth microdilution, time-kill, and cell membrane leakage assays. AgNP/ToCNF exhibited MIC₉₀ values of 250 μg mL^-1 against <i>S. aureus</i> and 125 μg mL^-1 against <i>P. aeruginosa</i> and <i>E. coli</i>, whereas free-state AgNPs showed MIC₉₀ values of 62.5 μg mL^-1 against <i>S. aureus</i> and 31.25 μg mL^-1 against <i>P. aeruginosa</i> and <i>E. coli</i>. Both compounds demonstrated bactericidal activity against all three bacterial strains. Cytotoxicity was assessed using the LDH assay, revealing a concentration-dependent toxicity pattern. Notably, AgNP/ToCNF exhibited minimal toxicity to human dermal fibroblasts (HDFs) at concentrations ≤500 μg mL^-1 after 72 hours, while free-state AgNPs induced >67% cytotoxicity. Although CNFs derived from SMS lacked intrinsic antimicrobial activity, their incorporation with AgNPs significantly enhanced antibacterial efficacy while simultaneously reducing AgNPs-induced cytotoxicity in mammalian cells. These findings underscore the potential of SMS-derived CNFs as biocompatible nanocarriers for AgNPs and other antibacterial agents, offering a sustainable and eco-friendly approach to developing antimicrobial biomaterials. This study explores the feasibility of upcycling SMS into high-value biomedical products, creating opportunities for future applications in wound healing, antimicrobial coatings, and medical nanocomposites.