Cytotoxicity Assessment of Bacterial Cellulose Nanoparticles for Targeted Drug Delivery

Many U.S. Food & Drug Administration approved materials in nanomedicine technologies, like the copolymer poly(lactic-co-glycolic acid) polyethylene glycol (PLGA-PEG), involve chemically intensive syntheses and are difficult to manufacture at the kilogram scale. Although PLGA-PEG is an FDA approved platform, the synthetic polymeric material is challenging to produce sustainably, especially at commercial scale. Green engineering is one way to address the concerns of negative environmental impact and sustainability in formulation of nano-based delivery systems. We have recently developed bacterial cellulose nanoparticles (BCNPs) for sustainable drug delivery. The development of BCNPs was motivated by a synthesis method that has reduced environmental impact, an overall eco-friendly life cycle, and can be implemented following green engineering methodologies. A bacterial cellulose (BC) pellicle was first collected from a kombucha co-culture media (bacteria and yeast) from the air and water interface, washed with sodium hydroxide and water for isolation and purifications steps, and then dissolved in dimethylacetamide and lithium chloride. The dissolution of BC then underwent a nanoprecipitation with surfactant solution to formulate BCNPs. BCNPs are 100-200 nm in diameter, polydispersity index (<0.3, PDI) slightly negative zeta-potential, and have a primarily amorphous morphology, desired parameters for targeted drug delivery. The particle size and zeta-potential were measured by scanning transmission electron microscopy and NanoSizer, while the PDI was calculated. To further understand the use of BCNPs for targeted drug delivery, we evaluated the cytotoxicity of BCNPs in our established organotypic whole hemisphere (OWH) brain slice platform prepared from postnatal day 10 (P10) rat brains and compared to chitosan nanoparticles, field standards for polysaccharide-based nanoparticles. After plating 300 µm OWH slices on semi-permeable membranes in slice culture media, slices were cultured for 4 days in vitro (DIV). At 4 DIV, BCNPs were topically applied to slices at 3 doses: 5 µg, 25 µg, 75 µg. After 24 h of BCNPs exposure, slices were stained with propidium iodine (PI), a cell death stain, and TO-PRO-3, a positive nuclei stain. Slices were fixed in methanol and PI+ cells were imaged using confocal microscopy. Our results establish a sustainably derived platform in nanomedicine for targeted drug delivery applications, and can open opportunities for other therapeutics to be produced using green engineering.