A new nanozyme with peroxidase-like activity for simultaneous phosphoprotein isolation and detection based on metal oxide affinity chromatography: Monodisperse-porous cerium oxide microspheres


YILDIRIM D., Gokcal B., BÜBER E., Kip C., DEMİR M., Tuncel A.

CHEMICAL ENGINEERING JOURNAL, vol.403, 2021 (Peer-Reviewed Journal) identifier identifier

  • Publication Type: Article / Article
  • Volume: 403
  • Publication Date: 2021
  • Doi Number: 10.1016/j.cej.2020.126357
  • Journal Name: CHEMICAL ENGINEERING JOURNAL
  • Journal Indexes: Science Citation Index Expanded, Scopus, Academic Search Premier, PASCAL, Aerospace Database, Aqualine, BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, Chimica, Compendex, Food Science & Technology Abstracts, INSPEC, Pollution Abstracts, Veterinary Science Database, Civil Engineering Abstracts
  • Keywords: Microfluidic system, Silica, Nanoparticles, Immobilized metal affinity chromatography, Casein, Phosphopeptides, HIGHLY SELECTIVE ENRICHMENT, COLORIMETRIC DETECTION, SILICA MICROSPHERES, NANOPARTICLES, PURIFICATION, NANORODS, CAPTURE, GLUCOSE, PHOSPHOPEPTIDES, NANOCOMPOSITES

Abstract

Monodisperse-porous cerium oxide (CeO2) microspheres obtained by a new method were used for simultaneous phosphoprotein isolation and detection for the first time. The synthesis method, "staged sol-gel templating protocol" provided monodisperse-porous CeO2 microspheres 5.0 mu m in size, with the specific surface areas and the pore volumes up to 80 m(2)/g and 0.41 cm(3)/g, respectively. CeO2 microspheres were directly used as a metal oxide affinity chromatography (MOAC) sorbent without any post-functionalization protocol. The isolation/enrichment of phosphoproteins from complex biological samples such as milk and human serum was performed using CeO2 microspheres as the sorbent, with the purities up to 99%. In batch MOAC runs, the equilibrium adsorption capacities of 105.3 and 82.4 mg phosphoprotein/g sorbent were obtained for alpha-casein and beta-casein, respectively. CeO2 microspheres also exhibited peroxidase-like activity and were proposed as a new nanozyme for colorimetric determination of phosphoprotein concentration in complex samples. Maximum substrate consumption rate and Km were determined as 384.6 mu M/min and 2885.8 mu M, respectively, according to Michealis-Menten model. The peroxidase-like activity of CeO2 microspheres linearly decreased with the increasing phosphoprotein concentration while no appreciable change in this magnitude was observed with the increasing non-phosphoprotein concentration. This behavior was explained by the increase in the pseudospecific phosphoprotein adsorption onto CeO2 microspheres with the increasing phosphoprotein concentration. Similar linear tendencies between peroxidase-like activity and phosphoprotein concentration observed in the complex samples such as human serum and milk allowed the determination of phosphoprotein concentration up to 300 and 400 mu g/mL, respectively, using CeO2 microspheres as the nanozyme. This study provided a new, unique material having a dual function termed as isolation and detection of large phosphorylated molecules without applying any post-functionalization protocol.