Development of boronic acid-functionalized mesoporous silica-coated core/shell magnetic microspheres with large pores for endotoxin removal

Abstract

Endotoxins are found almost everywhere and possess high toxicity in vivo and in vitro. Here we design a novel boronate affinity material, called boronic acid-functionalized mesoporous silica-coated core/shell magnetic microspheres (Fe3O4@nSiO2@mSiO2-BA) with large pores (pore size > 20 nm) based on the chemical structure and physical properties of endotoxins, for facile and highly efficient removal of endotoxins. Dual modes for endotoxin removal were proposed and confirmed in this work: the endotoxin aggregates with size < 20 nm were bound with boronic acid ligands chemically modified on the inner and outer surface of the large pores of Fe3O4@nSiO2@mSiO2-BA microspheres; while the larger endotoxin micelles (size >20 nm) were absorbed on the outer surface of the prepared material based on boronate affinity. Transmission electron microscopy (TEM), X-ray diffraction (XRD), nitrogen adsorption/desorption isotherms and Fourier transform infrared (FT-IR) spectroscopy confirm that Fe3O4@nSiO2@mSiO2-BA microspheres possess core/shell structure, uniform diameter (520 nm), high surface area (205.57 m2/g), large mesopores (21.8 nm) and boronic acid ligands. The purification procedures of Fe3O4@nSiO2@mSiO2-BA microspheres for endotoxin were optimized, and 50 mM NH4HCO3 (pH 8.0) and 0.05 M fructose were selected as loading/washing, elution buffers, respectively. The binding capacity of Fe3O4@nSiO2@mSiO2-BA microspheres for endotoxin was calculated to be 60.84 EU/g under the optimized conditions. Finally, the established analytical method was applied to remove endotoxins from plasmid DNA. After endotoxin removal, the endotoxin content in plasmid DNA was reduced from 0.0026 to 0.0006 EU/mL for two-fold concentration, and from 0.0088 to 0.0022 EU/mL for five-fold concentration after binding, respectively. Additional advantages of the prepared boronate affinity material include excellent stability, reusability/repeatability, and low cost. Boronate affinity materials with large pores could thus prove to be powerful adsorbents for endotoxin removal and the potential applications in the aspects of biological research, pharmaceutical industry, and life health.