Preparation of Cationic Lignin Nanoparticles

Never-dried softwood kraft pulp fibers (Metsä Board Husum mill, Sweden) were used to prepare TCNF following the method of (Saito et al. 2007 (link)). Sulfur-free birch lignin produced by BLN process (named after the company that initially developed the technology) was provided by CH Bioforce Oy (Finland). Briefly, the process involves hot water (150 °C) extraction of hemicelluloses from biomass followed by alkali cooking to extract lignin, all in oxygen-starved conditions (von Schoultz 2015 ). The LNPs were prepared by anti-solvent nanoprecipitation in acetone and water (Figueiredo et al. 2021 (link)). These LNPS were cationized by coating with glycidyltrimethylammonium chloride (GTAC)-treated lignin to produce cationic LNPs (cLNPs) (Agustin et al. 2022 (link); Sipponen et al. 2017 (link)). The detailed preparation of TCNF, LNPs, and cLNPs and their basic characteristics are available in the Supporting Information (Fig. S2). Hexadecane, GTAC, cyclohexanone, and acetone were purchased from Fisher Scientific (Finland). The pharmaceutical compounds, which are certified reference materials, dialysis membrane tubing (Spectra/Por 7, 6–8 kDa molecular weight cut-off (MWCO), Spectra/Por 1, 1 kDa MWCO), glass microfibre filters with a 0.7 µm pore size (Whatman GF/F), chromasolv-grade acetonitrile and methanol were obtained from Sigma-Aldrich (Finland).