Robust non-toxic macroscale beads with antibacterial and contaminant scavenging properties for aquaculture

Non-toxic macroscale (few millimeters in diameter) cationic antibacterial beads were made for suppression of bacteria in fish culture. The beads were formed by the diffusion-driven layer by layer (dd-LBL) process using graphene oxide substrate and branched polyethyleneimine and were stabilized by post-assembly crosslinking process in which carboxyl groups on the graphene oxide were crosslinked with amine groups on the branched polyethyleneimine using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) chemistry. The crosslinked beads have negligible in vitro toxicity to 3T3 cells and zebrafish embryos. Further, the crosslinked beads exhibit excellent antibacterial activity, which can reach over 99.9% inactivation against Escherichia coli (E. coli) in 30 min even with ultrahigh (~ 108 CFU/mL) E. coli concentration. The antibacterial activity of the crosslinked beads can be regenerated with a simple procedure and remains high in the 10th cycle of regeneration. The crosslinked beads have much improved mechanical stability compared to the raw (uncrosslinked) beads. The highly porous structure and abundant functional groups of the crosslinked beads give them high adsorption capacity towards various forms of organic matter such as fish waste. Addition of zeolite nanoparticles during the bead formation process and subsequent chemical crosslinking with hyaluronic acid makes these beads suitable for removal of total ammonia nitrogen (TAN) while preserving the antibacterial activity and biocompatibility.