{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T05:04:13Z","timestamp":1787029453902,"version":"build-2736575974"},"reference-count":56,"publisher":"MDPI AG","issue":"16","license":[{"start":{"date-parts":[[2026,8,17]],"date-time":"2026-08-17T00:00:00Z","timestamp":1786924800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Polymers"],"abstract":"<jats:p>Magnetic poly(vinylphosphonic acid-co-ethylene glycol dimethacrylate)-TiO2 [m-poly(VPA-co-EGDMA)-TiO2] microspheres were synthesized by suspension polymerization and evaluated as supports for direct acetylcholinesterase (AChE) immobilization. The microspheres were characterized by SEM, FT-IR, BET, XRD, XPS, VSM, and zeta potential analyses and exhibited porous morphology, magnetic responsiveness, a specific surface area of 195 m2 g\u22121, and a swelling ratio of 40.19%. AChE was immobilized mainly through physical adsorption and multipoint non-covalent enzyme\u2013support interactions. The optimum pH was pH 10 for both free and immobilized AChE. Under the tested assay conditions, immobilized AChE showed apparent kinetic parameters of Km = 0.028 mM and Vmax = 0.132 EU.mL\u22121. Immobilized AChE retained 58.2% of its initial activity after 60 days at 4 \u00b0C and 24% after 20 reuse cycles. Overall, the microspheres offer a magnetically recoverable support for AChE immobilization, although further optimization is required to improve kinetic performance and long-term operational stability.<\/jats:p>","DOI":"10.3390\/polym18161999","type":"journal-article","created":{"date-parts":[[2026,8,17]],"date-time":"2026-08-17T10:04:36Z","timestamp":1786961076000},"page":"1999","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Magnetic Poly(VPA-co-EGDMA)-TiO2 Polymeric Microspheres as Supports for Acetylcholinesterase Immobilization: Kinetic Behavior, Stability, and Reusability"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7958-4533","authenticated-orcid":false,"given":"\u015eahin","family":"\u00d6zel","sequence":"first","affiliation":[{"name":"Department of Chemistry, Faculty of Science and Arts, Bursa Uluda\u011f University, Bursa 16059, Turkey"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,8,17]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1019","DOI":"10.1016\/j.procbio.2008.06.004","article-title":"Enzyme stability and stabilization\u2014Aqueous and non-aqueous environment","volume":"43","author":"Iyer","year":"2008","journal-title":"Process Biochem."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1451","DOI":"10.1016\/j.enzmictec.2007.01.018","article-title":"Improvement of enzyme activity, stability and selectivity via immobilization techniques","volume":"40","author":"Mateo","year":"2007","journal-title":"Enzym. Microb. Technol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1639","DOI":"10.1007\/s10529-009-0076-4","article-title":"Advances in enzyme immobilisation","volume":"31","author":"Brady","year":"2009","journal-title":"Biotechnol. Lett."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1016\/j.cbpa.2005.02.014","article-title":"Immobilised enzymes: Science or art?","volume":"9","author":"Cao","year":"2005","journal-title":"Curr. Opin. Chem. Biol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1038\/35051706","article-title":"Enzymes for chemical synthesis","volume":"409","author":"Koeller","year":"2001","journal-title":"Nature"},{"key":"ref_6","first-page":"1113","article-title":"Conducting polymer-based electrochemical biosensor for the detection of acetylthiocholine and pesticide via acetylcholinesterase","volume":"68","author":"Akdag","year":"2021","journal-title":"Biotechnol. Appl. Biochem."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1002\/elsc.201700130","article-title":"Covalent immobilization of acetylcholinesterase on a novel polyacrylic acid-based nanofiber membrane","volume":"18","author":"Ogan","year":"2018","journal-title":"Eng. Life Sci."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"16345","DOI":"10.1021\/acs.analchem.2c03290","article-title":"Colorimetric assay for acetylcholinesterase activity and inhibitor screening based on metal\u2013organic framework nanosheets","volume":"94","author":"Wang","year":"2022","journal-title":"Anal. Chem."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"5216","DOI":"10.1021\/cr900214c","article-title":"History and new developments of assays for cholinesterase activity and inhibition","volume":"110","author":"Miao","year":"2010","journal-title":"Chem. Rev."},{"key":"ref_10","first-page":"1118","article-title":"Acetylcholinesterase-based biosensor electrodes for organophosphate pesticide detection: I. Modification of carbon surface for immobilization of acetylcholinesterase","volume":"20","author":"Vakurov","year":"2004","journal-title":"Biosens. Bioelectron."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"10777","DOI":"10.1002\/jcb.28369","article-title":"Acetylcholinesterase with mesoporous silica: Covalent immobilization, physiochemical characterization, and its application in food for pesticide detection","volume":"120","author":"Palanivelu","year":"2019","journal-title":"J. Cell. Biochem."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"210","DOI":"10.1002\/bit.21107","article-title":"Crosslinked enzyme aggregates in hierarchically-ordered mesoporous silica: A simple and effective method for enzyme stabilization","volume":"96","author":"Kim","year":"2007","journal-title":"Biotechnol. Bioeng."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Antony, C., Ghodke, P.K., and Thiyagarajan, S. (2022). Overview of the enzyme support system of immobilization for enhanced efficiency and reuse of enzymes. Enzymes in the Valorization of Waste, CRC Press.","DOI":"10.1201\/9781003187721-6"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Kaur, J. (2026). Advanced enzyme immobilization: Problems and solutions. Recent Trends in Enzyme Immobilization, Elsevier.","DOI":"10.1016\/B978-0-443-27475-6.00015-2"},{"key":"ref_15","unstructured":"Benaoura, A., Gasmi, M., Bouizzar, M.S., and Rira, M. (2026, August 10). Cefazolin Biodegradation by a Molecularly Identified Bacterium: Hybrid RSM-ANN-GA Optimization and Enzyme Immobilization on Fe3O4 Nanoparticles. Available online: https:\/\/ssrn.com\/abstract=5464694."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"205","DOI":"10.1080\/13102818.2015.1008192","article-title":"An overview of technologies for immobilization of enzymes and surface analysis techniques for immobilized enzymes","volume":"29","author":"Mohamad","year":"2015","journal-title":"Biotechnol. Biotechnol. Equip."},{"key":"ref_17","unstructured":"Hern\u00e1ndez Coronado, E.E. (2019). Efecto Sin\u00e9rgico de los Procesos de Fotocat\u00e1lisis y Foto-Fenton Heterog\u00e9neos en la Degradaci\u00f3n de Cefuroxima Bajo Radiaci\u00f3n Solar Simulada con el Sistema \u03b1-Fe2O3-TiO2\/H2O2. [Master\u2019s Thesis, Universidad Aut\u00f3noma de Nuevo Le\u00f3n]."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"141172","DOI":"10.1016\/j.ijbiomac.2025.141172","article-title":"Innovative immobilization of \u03b2-glucosidase on amidoximated acrylic fabric integrated with magnetite Fe3O4 nanoparticles","volume":"306","author":"Almulaiky","year":"2025","journal-title":"Int. J. Biol. Macromol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1802922","DOI":"10.1002\/adma.201802922","article-title":"Porous polymers as multifunctional material platforms toward task-specific applications","volume":"31","author":"Wu","year":"2019","journal-title":"Adv. Mater."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Li, J.-W., Cheng, C.-C., and Chiu, C.-W. (2024). Advances in multifunctional polymer-based nanocomposites. Polymers, 16.","DOI":"10.3390\/polym16233440"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Lee, J., and Patel, R. (2022). Wastewater treatment by polymeric microspheres: A review. Polymers, 14.","DOI":"10.3390\/polym14091890"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"e00157","DOI":"10.1002\/ppsc.202500157","article-title":"Precision-Engineered Poly (GMA-EGDMA) Microspheres for Multifunctional Applications","volume":"43","author":"Ali","year":"2026","journal-title":"Part. Part. Syst. Charact."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2436","DOI":"10.1039\/D4MH01618A","article-title":"Porous polymers: Structure, fabrication and application","volume":"12","author":"Liu","year":"2025","journal-title":"Mater. Horiz."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"\u00d6zel, \u015e., and Kara, A. (2026). Magnetically recoverable phosphonic acid-functionalized polymeric TiO2 microspheres for synergistic adsorption\u2013photocatalytic removal of Reactive Blue 221. Sep. Sci. Technol., 1\u201317.","DOI":"10.1080\/01496395.2026.2702974"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"88","DOI":"10.1016\/0006-2952(61)90145-9","article-title":"A new and rapid colorimetric determination of acetylcholinesterase activity","volume":"7","author":"Ellman","year":"1961","journal-title":"Biochem. Pharmacol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e202301824","DOI":"10.1002\/cbdv.202301824","article-title":"Design and synthesis of pyrazole carboxamide derivatives as selective cholinesterase and carbonic anhydrase inhibitors: Molecular docking and biological evaluation","volume":"21","author":"Durgun","year":"2024","journal-title":"Chem. Biodivers."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Moehlenbrock, M.J., and Minteer, S.D. (2016). Introduction to the field of enzyme immobilization and stabilization. Enzyme Stabilization and Immobilization: Methods and Protocols, Springer.","DOI":"10.1007\/978-1-4939-6499-4_1"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"324","DOI":"10.1016\/0003-2697(78)90350-0","article-title":"Modified Ellman procedure for assay of cholinesterases in crude enzymatic preparations","volume":"86","author":"Gorun","year":"1978","journal-title":"Anal. Biochem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"6290","DOI":"10.1039\/C2CS35231A","article-title":"Modifying enzyme activity and selectivity by immobilization","volume":"42","author":"Rodrigues","year":"2013","journal-title":"Chem. Soc. Rev."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1016\/j.procbio.2019.11.001","article-title":"High throughput covalent immobilization process for improvement of shelf-life, operational cycles, relative activity in organic media and enzymatic kinetics of urease and its application for urea removal from water samples","volume":"90","author":"Jangi","year":"2020","journal-title":"Process Biochem."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3550","DOI":"10.1002\/elps.200406096","article-title":"Immobilized microfluidic enzymatic reactors","volume":"25","author":"Foret","year":"2004","journal-title":"Electrophoresis"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1016\/j.memsci.2003.06.004","article-title":"Progress in enzymatic membrane reactors\u2014A review","volume":"242","author":"Rios","year":"2004","journal-title":"J. Memb. Sci."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"95","DOI":"10.1016\/j.procbio.2017.03.025","article-title":"Immobilization and characterization of human carbonic anhydrase I on amine functionalized magnetic nanoparticles","volume":"57","author":"Sahin","year":"2017","journal-title":"Process Biochem."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Bi\u00e9, J., Sepodes, B., Fernandes, P.C.B., and Ribeiro, M.H.L. (2022). Enzyme immobilization and co-immobilization: Main framework, advances and some applications. Processes, 10.","DOI":"10.3390\/pr10030494"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"116031","DOI":"10.1016\/j.fuel.2019.116031","article-title":"Adsorption pore structure and its fractal characteristics of coals by N2 adsorption\/desorption and FESEM image analyses","volume":"257","author":"Li","year":"2019","journal-title":"Fuel"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"128615","DOI":"10.1016\/j.cej.2021.128615","article-title":"N-doped magnetic three-dimensional carbon microspheres@TiO2 with a porous architecture for enhanced degradation of tetracycline and methyl orange via adsorption\/photocatalysis synergy","volume":"411","author":"Zhou","year":"2021","journal-title":"Chem. Eng. J."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"2885","DOI":"10.1002\/adsc.201100163","article-title":"Potential of different enzyme immobilization strategies to improve enzyme performance","volume":"353","author":"Rodrigues","year":"2011","journal-title":"Adv. Synth. Catal."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"14139","DOI":"10.3390\/molecules190914139","article-title":"Inorganic materials as supports for covalent enzyme immobilization: Methods and mechanisms","volume":"19","author":"Zucca","year":"2014","journal-title":"Molecules"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"4402","DOI":"10.1021\/acscatal.8b04921","article-title":"Multienzymatic cascade reactions via enzyme complex by immobilization","volume":"9","author":"Hwang","year":"2019","journal-title":"ACS Catal."},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Baruwa, A., Gbadeyan, O.J., and Permaul, K. (2026). Application of Biotechnology in the Synthesis of Nanoparticles\u2014A Review. Molecules, 31.","DOI":"10.3390\/molecules31091415"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"385101","DOI":"10.1088\/1361-6528\/acda35","article-title":"Nanotechnology and enzyme immobilization: A review","volume":"34","author":"Oke","year":"2023","journal-title":"Nanotechnology"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1002\/elsc.200900001","article-title":"Immobilization of acetylcholinesterase in nanofibrous PVA\/BSA membranes by electrospinning","volume":"10","author":"Moradzadegan","year":"2010","journal-title":"Eng. Life Sci."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"e00311","DOI":"10.1042\/BSR20150154","article-title":"Acetylcholinesterase immobilization and characterization, and comparison of the activity of the porous silicon-immobilized enzyme with its free counterpart","volume":"36","author":"Saleem","year":"2016","journal-title":"Biosci. Rep."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Chaari, M., Ben Amara, F., Smaoui, S., Ali, D.S., Ben Hlima, H., and Mousavi Khaneghah, A. (2026). Biosensing Technologies for Ensuring the Safety of Meat Products: A Critical Review. Crit. Rev. Anal. Chem., 1\u201317.","DOI":"10.1080\/10408347.2025.2610326"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"223","DOI":"10.2174\/1568026614666141229112734","article-title":"Sol-gel encapsulation of biomolecules and cells for medicinal applications","volume":"15","author":"Wang","year":"2015","journal-title":"Curr. Top. Med. Chem."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"6735","DOI":"10.1021\/acs.jafc.1c01091","article-title":"Multiple roles of mesoporous silica in safe pesticide application by nanotechnology: A review","volume":"69","author":"Kong","year":"2021","journal-title":"J. Agric. Food Chem."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"72","DOI":"10.1016\/j.bioelechem.2012.02.005","article-title":"The stabilization of Au NP\u2013AChE nanocomposites by biosilica encapsulation for the development of a thiocholine biosensor","volume":"86","author":"Buiculescu","year":"2012","journal-title":"Bioelectrochemistry"},{"key":"ref_48","doi-asserted-by":"crossref","unstructured":"Salem, K.S., Rashid, T.U., Islam, M.M., Khan, M.N., Sharmeen, S., Rahman, M.M., and Haque, P. (2016). Recent Updates on Immobilization of Microbial Cellulase. New and Future Developments in Microbial Biotechnology and Bioengineering, Elsevier.","DOI":"10.1016\/B978-0-444-63507-5.00011-3"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"e70038","DOI":"10.1111\/1750-3841.70038","article-title":"The current research status of immobilized lipase performance and its potential for application in food are developing toward green and healthy direction: A review","volume":"90","author":"Wang","year":"2025","journal-title":"J. Food Sci."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1016\/S0003-2670(02)00518-4","article-title":"Immobilization of acetylcholinesterase on screen-printed electrodes: Comparative study between three immobilization methods and applications to the detection of organophosphorus insecticides","volume":"464","author":"Andreescu","year":"2002","journal-title":"Anal. Chim. Acta"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"2530","DOI":"10.1016\/j.ijbiomac.2018.09.025","article-title":"Magnetic nanoparticles as versatile carriers for enzymes immobilization: A review","volume":"120","author":"Bilal","year":"2018","journal-title":"Int. J. Biol. Macromol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1080\/07328303.2017.1347668","article-title":"Carrageenan for encapsulation and immobilization of flavor, fragrance, probiotics, and enzymes: A review","volume":"36","author":"Chakraborty","year":"2017","journal-title":"J. Carbohydr. Chem."},{"key":"ref_53","unstructured":"Gong, F., Zhang, G., and Su, Z. (2013). Microspheres for Separation of Bioactive Small Molecules. Microspheres and Microcapsules in Biotechnology: Design, Preparation and Applications, Jenny Stanford Publishing."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"324","DOI":"10.1109\/JSTQE.2014.2381491","article-title":"Photocurable polymer nanocomposites for magnetic, optical, and biological applications","volume":"21","author":"Song","year":"2014","journal-title":"IEEE J. Sel. Top. Quantum Electron."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"639","DOI":"10.1080\/24701556.2023.2187418","article-title":"A review on magnetic polymeric nanocomposite materials: Emerging applications in biomedical field","volume":"53","author":"Vijayasri","year":"2023","journal-title":"Inorg. Nano-Metal. Chem."},{"key":"ref_56","doi-asserted-by":"crossref","unstructured":"Das, R., Dwevedi, A., and Kayastha, A.M. (2021). Current and future trends on polymer-based enzyme immobilization. Polymeric Supports for Enzyme Immobilization: Opportunities and Applications, Elsevier.","DOI":"10.1016\/B978-0-12-819206-1.00004-1"}],"container-title":["Polymers"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4360\/18\/16\/1999\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,17]],"date-time":"2026-08-17T10:20:37Z","timestamp":1786962037000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4360\/18\/16\/1999"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,8,17]]},"references-count":56,"journal-issue":{"issue":"16","published-online":{"date-parts":[[2026,8]]}},"alternative-id":["polym18161999"],"URL":"https:\/\/doi.org\/10.3390\/polym18161999","relation":{},"ISSN":["2073-4360"],"issn-type":[{"value":"2073-4360","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,17]]}}}