A method for purifying nanoparticles using cationic modified monoliths and aqueous elution

Nanoparticles are widely used in the medical field for diagnosis and therapy. In particular, the use of nanoparticles containing vaccines has spread rapidly; hence, ensuring nanoparticle safety and minimizing their side effects have become important concerns worldwide. In this study, we used three types (NH2, poly-Lys, and trimethylaminopropyl) of cationic modified silica monoliths with cylindrical structures, diameters of 4.2 mm, and heights of 1.5 mm. Doxil, an anticancer nanomedicine, and exosomes, as typical nanoparticles, were separated from model leaked drugs (e.g., doxorubicin and oligonucleotides) and proteins (e.g., albumin) coexisting in nanoparticle sample solutions using these monoliths. Each nanoparticle solution (200 μL) was applied to each monolith followed by centrifugation at 9,100 g for 1 min. The ionic concentration of the elution solution was increased stepwise to determine the concentration required to elute the nanoparticles from each monolith by centrifugation. The NH2- and poly-Lys-modified monoliths separated and purified nanoparticles from leaked drugs or proteins coexisting in nanoparticle sample solutions. The nanoparticles were separated from other substances by changing the pH and concentration of the aqueous Tris buffer used as the eluent. Doxil was eluted with 500-1,000 mM Tris buffer (pH 8) when using the NH2-modified monolith, and with 200-1,000 mM Tris buffer (pH 6) when using the poly-Lys-modified monolith. Exosome was obtained using 1,000 mM Tris buffer (pH 8) and the NH2-modified monolith. The recovery efficiencies (ratio of nanoparticle content in the most abundant fraction to that in the sample solution before purification) of Doxil and exosome were 64% and 55%, respectively. Because this method can purify nanoparticles using only low-speed centrifugation for a few minutes, we expect it will be used to improve nanoparticle safety.