{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T00:13:06Z","timestamp":1788221586560,"version":"build-2803163510"},"reference-count":81,"publisher":"American Chemical Society (ACS)","issue":"33","license":[{"start":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T00:00:00Z","timestamp":1786665600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100031478","name":"NextGenerationEU","doi-asserted-by":"publisher","award":["NA"],"award-info":[{"award-number":["NA"]}],"id":[{"id":"10.13039\/100031478","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100007642","name":"Universit? degli Studi di Roma Tor Vergata","doi-asserted-by":"publisher","award":["HyNDiGoCat 2024"],"award-info":[{"award-number":["HyNDiGoCat 2024"]}],"id":[{"id":"10.13039\/501100007642","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100021856","name":"Ministero dell'Universit? e della Ricerca","doi-asserted-by":"publisher","award":["E53D22014620001"],"award-info":[{"award-number":["E53D22014620001"]}],"id":[{"id":"10.13039\/501100021856","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,8,25]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Biochar, a carbon-rich material produced through biomass pyrolysis, is gaining significant attention in electrochemical sensor development. This material offers dual advantages: it enhances electrode performance while providing functional groups for binding with recognition elements such as antibodies, enzymes, and aptamers. Despite growing interest, practical applications encounter several challenges. The biochar heterogeneous surface could lead to random immobilization of interaction sites and fouling, both of which limit sensor performance, particularly in complex matrices. In addition, surface modification strategies specifically designed to introduce molecular spacers on biochar for electrochemical biosensing remain comparatively underexplored. This work introduces a multistep chemical modification strategy designed to add reactive functional groups to the biochar surface and, in particular, is focused on the development of a molecular spacer on spent coffee ground-derived biochar (pristine biochar) by introducing a covalently bound molecular moiety through azide\u2013alkyne cycloaddition (clicked biochar). Each reaction intermediate was characterized through in-depth chemical\u2013physical characterization, confirming successful functionalization at each step. Crucially, the molecular spacer extends recognition elements further into solution while limiting large protein adsorption. To validate this approach, the performance of an enzymatic biosensor using alkaline phosphatase (AP) and a model substrate was evaluated. The functionalized biochar exhibits improved antifouling properties and better analytical performance compared to that of its unmodified counterpart. A kinetic study conducted using chronoamperometry (CA) shows an approximately 2.2-fold increase in the apparent maximum catalytic current for clicked biochar compared with pristine biochar, while the apparent Michaelis\u2013Menten constants remained comparable, confirming improved enzyme accessibility and enhanced catalytic efficiency of the covalently immobilized enzyme. Furthermore, electrochemical impedance spectroscopy (EIS) confirmed the antifouling ability of this material, resulting in a higher reduction in nonspecific adsorption compared to pristine biochar, as highlighted by a change in charge transfer resistance (Rct).<\/jats:p>","DOI":"10.1021\/acsomega.6c01575","type":"journal-article","created":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T17:02:14Z","timestamp":1786726934000},"page":"49163-49174","source":"Crossref","is-referenced-by-count":0,"title":["Click-Functionalized Biochar as a Spacer-Engineered\nInterface for Electrochemical Biosensing"],"prefix":"10.1021","volume":"11","author":[{"given":"Antonio","family":"Licheri","sequence":"first","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Veronica","family":"Mattarelli","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lorenzo","family":"Bartolucci","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via del Politecnico 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3278-0414","authenticated-orcid":true,"given":"Rocco","family":"Cancelliere","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8369-5789","authenticated-orcid":true,"given":"Claudia","family":"Mazzuca","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pietro","family":"Mele","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via del Politecnico 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Silvia","family":"Orlanducci","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6970-0318","authenticated-orcid":true,"given":"Laura","family":"Micheli","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0398-8408","authenticated-orcid":true,"given":"Riccardo","family":"Salvio","sequence":"additional","affiliation":[{"name":"Tor Vergata University of Rome , , Via Della Ricerca Scientifica 1 , ,","place":["Roma, Italy, 00133"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,8,14]]},"reference":[{"key":"2026083119150170700_cit1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jece.2022.107260","article-title":"Emerging trends\nin the application of carbon-based materials: A review","volume":"10","author":"Egbedina","year":"2022","journal-title":"J. Environ. Chem. Eng."},{"key":"2026083119150170700_cit2","doi-asserted-by":"publisher","first-page":"301","DOI":"10.1016\/j.carbon.2022.07.023","article-title":"The Role of Carbon-Based Materials for Fuel Cells Performance","volume":"198","author":"Jaleh","year":"2022","journal-title":"Carbon"},{"key":"2026083119150170700_cit3","doi-asserted-by":"publisher","first-page":"721","DOI":"10.1016\/j.carbon.2021.07.053","article-title":"Carbon-based materials for fast charging\nlithium-ion batteries","volume":"183","author":"Li","year":"2021","journal-title":"Carbon"},{"key":"2026083119150170700_cit4","doi-asserted-by":"publisher","first-page":"417","DOI":"10.1016\/j.mset.2019.04.006","article-title":"Recent development\nof carbon based materials for energy storage devices","volume":"2","author":"Iqbal","year":"2019","journal-title":"Mater. Sci. Energy Technol."},{"key":"2026083119150170700_cit5","doi-asserted-by":"publisher","first-page":"23710","DOI":"10.1039\/c2jm34066f","article-title":"KOH activation of carbon-based materials for energy\nstorage","volume":"22","author":"Wang","year":"2012","journal-title":"J. Mater. Chem."},{"key":"2026083119150170700_cit6","doi-asserted-by":"publisher","DOI":"10.1002\/adhm.202101834","article-title":"Carbon-Based Materials\nfor Articular Tissue Engineering: From Innovative Scaffolding Materials\ntoward Engineered Living Carbon","volume":"11","author":"Islam","year":"2022","journal-title":"Adv. Healthcare\nMater."},{"key":"2026083119150170700_cit7","doi-asserted-by":"publisher","DOI":"10.1016\/j.cobme.2021.100274","article-title":"Biosensing applications of carbon-based\nmaterials","volume":"18","author":"Joshi","year":"2021","journal-title":"Curr. Opin. Biomed. 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Sonochem."},{"key":"2026083119150170700_cit11","doi-asserted-by":"publisher","DOI":"10.1002\/cctc.202301477","article-title":"Enzyme Mimics\nBased on Guanidinocalix[4]arene\/Nanodiamond Hybrid Systems with Phosphodiesterase\nActivity","volume":"16","author":"Vezzoni","year":"2024","journal-title":"ChemCatChem"},{"key":"2026083119150170700_cit12","doi-asserted-by":"publisher","DOI":"10.1016\/j.ccr.2025.216479","article-title":"An equal terms comparison of the proficiency of artificial\nphosphodiesterases by using simple models of RNA or DNA as benchmarks\u2013the\ntakeaway to design next generation supramolecular catalysts","volume":"531","author":"Casnati","year":"2025","journal-title":"Coord. Chem. Rev."},{"key":"2026083119150170700_cit13","doi-asserted-by":"publisher","first-page":"557","DOI":"10.1007\/s10311-020-01084-9","article-title":"Environmental applications of carbon-based\nmaterials: a review","volume":"19","author":"Gopinath","year":"2021","journal-title":"Environ. Chem. Lett."},{"key":"2026083119150170700_cit14","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/j.jiec.2016.06.002","article-title":"Production and utilization of biochar:\nA review","volume":"40","author":"Cha","year":"2016","journal-title":"J. Ind. Eng. Chem."},{"key":"2026083119150170700_cit15","doi-asserted-by":"publisher","first-page":"240","DOI":"10.1016\/j.fuel.2017.12.054","article-title":"Properties of biochar","volume":"217","author":"Weber","year":"2018","journal-title":"Fuel"},{"key":"2026083119150170700_cit16","doi-asserted-by":"publisher","DOI":"10.1016\/j.jaap.2021.105405","article-title":"A critical\nreview on production, modification and utilization of biochar","volume":"161","author":"Xie","year":"2022","journal-title":"J. Anal. Appl. 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Eng."},{"key":"2026083119150170700_cit29","doi-asserted-by":"publisher","first-page":"8951","DOI":"10.1007\/s10853-020-05499-1","article-title":"Fabrication of\nscreen-printed electrodes: opportunities and challenges","volume":"56","author":"Suresh","year":"2021","journal-title":"J. Mater. Sci."},{"key":"2026083119150170700_cit30","doi-asserted-by":"publisher","first-page":"865","DOI":"10.1007\/s00604-014-1181-1","article-title":"Screen-printed electrodes for biosensing:\na review (2008\u20132013)","volume":"181","author":"Taleat","year":"2014","journal-title":"Microchim. 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Rev."},{"key":"2026083119150170700_cit34","doi-asserted-by":"publisher","DOI":"10.1093\/rb\/rbaf007","article-title":"Analytical methods in studying cell force sensing: principles, current\ntechnologies and perspectives","volume":"12","author":"Liu","year":"2025","journal-title":"Regener. Biomater."},{"key":"2026083119150170700_cit35","doi-asserted-by":"publisher","DOI":"10.1016\/j.snb.2024.137224","article-title":"Construction of an\nelectrochemical aptasensor based on a covalent organic framework for\nrapid and ultrasensitive detection of Staphylococcus aureus","volume":"428","author":"Zhang","year":"2025","journal-title":"Sens. Actuators, B"},{"key":"2026083119150170700_cit36","doi-asserted-by":"publisher","DOI":"10.3389\/fenrg.2020.00138","article-title":"Impact of Biomass Source and Pyrolysis\nParameters on Physicochemical Properties of Biochar Manufactured for\nInnovative Applications","volume":"8","author":"Uroic\u0301 S\u030ctefanko","year":"2020","journal-title":"Front. 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Bioelectron."},{"key":"2026083119150170700_cit40","doi-asserted-by":"publisher","DOI":"10.1016\/j.scitotenv.2022.159797","article-title":"Biochar: An environmentally friendly\nplatform for construction of a SARS-CoV-2 electrochemical immunosensor","volume":"858","author":"Valenga","year":"2023","journal-title":"Sci. Total Environ."},{"key":"2026083119150170700_cit41","doi-asserted-by":"publisher","DOI":"10.3389\/fchem.2023.1251360","article-title":"Label-free electrochemical\nimmunosensor as a reliable point-of-care device for the detection\nof Interleukin-6 in serum samples from patients with psoriasis","volume":"11","author":"Cancelliere","year":"2023","journal-title":"Front. Chem."},{"key":"2026083119150170700_cit42","doi-asserted-by":"publisher","first-page":"163","DOI":"10.1016\/j.talanta.2019.05.101","article-title":"Quick electrochemical\nimmunoassay for hantavirus detection based on biochar platform","volume":"204","author":"Martins","year":"2019","journal-title":"Talanta"},{"key":"2026083119150170700_cit43","doi-asserted-by":"publisher","DOI":"10.1016\/j.talanta.2023.125162","article-title":"Fouling-resistant electrode for electrochemical sensing based on\ncovalent-organic frameworks TpPA-1 dispersed cabon nanotubes","volume":"267","author":"Li","year":"2024","journal-title":"Talanta"},{"key":"2026083119150170700_cit44","doi-asserted-by":"publisher","DOI":"10.1016\/j.hazadv.2022.100226","article-title":"Adsorption of Pollutants from Wastewater by Biochar:\nA Review","volume":"9","author":"Jagadeesh","year":"2023","journal-title":"J. Hazard. Mater. Adv."},{"key":"2026083119150170700_cit45","doi-asserted-by":"publisher","DOI":"10.1016\/j.envpol.2022.119609","article-title":"Biochar-microorganism\ninteractions for organic pollutant remediation: Challenges and perspectives","volume":"308","author":"Mukherjee","year":"2022","journal-title":"Environ. Pollut."},{"key":"2026083119150170700_cit46","doi-asserted-by":"publisher","DOI":"10.1016\/j.cej.2023.148329","article-title":"Effect of\nbiochar chemical modification (acid, base and hydrogen peroxide) on\ncontaminants content depending on feedstock and pyrolysis conditions","volume":"481","author":"Hawryluk-Sidoruk","year":"2024","journal-title":"Chem. Eng. J."},{"key":"2026083119150170700_cit47","doi-asserted-by":"publisher","DOI":"10.1016\/j.chemosphere.2022.137238","article-title":"Chemical modification of biochars\nas a method to improve its surface properties and efficiency in removing\nxenobiotics from aqueous media","volume":"312","author":"Tomczyk","year":"2023","journal-title":"Chemosphere"},{"key":"2026083119150170700_cit48","doi-asserted-by":"publisher","DOI":"10.1016\/j.cej.2022.140310","article-title":"Interfacial chemistry\nof mercury on thiol-modified biochar and its implication for adsorbent\nengineering","volume":"454","author":"Huang","year":"2023","journal-title":"Chem. Eng. J."},{"key":"2026083119150170700_cit49","doi-asserted-by":"publisher","first-page":"338","DOI":"10.1016\/j.wasman.2021.09.020","article-title":"Study on the performance\nof HNO(3)-modified biochar for enhanced medium temperature anaerobic\ndigestion of food waste","volume":"135","author":"Gao","year":"2021","journal-title":"Waste Manage."},{"key":"2026083119150170700_cit50","doi-asserted-by":"publisher","first-page":"25842","DOI":"10.1021\/acsomega.5c02113","article-title":"Surface Modification\nof Biochar to Prepare Environmentally Friendly Electrochemical Biosensors\nfor Detection of Cardiac Troponin T","volume":"10","author":"Faria","year":"2025","journal-title":"ACS Omega"},{"key":"2026083119150170700_cit51","doi-asserted-by":"publisher","first-page":"519","DOI":"10.1007\/s40726-022-00238-3","article-title":"Biochar Modification Methods for\nAugmenting Sorption of Contaminants","volume":"8","author":"Kumar","year":"2022","journal-title":"Curr. Pollut.\nRep."},{"key":"2026083119150170700_cit52","doi-asserted-by":"publisher","DOI":"10.3389\/fsens.2025.1552969","article-title":"Electrochemical\nand biosensing applications of biochar and biochar-based composites","volume":"6","author":"Tshoko","year":"2025","journal-title":"Front. Sens."},{"key":"2026083119150170700_cit53","doi-asserted-by":"publisher","first-page":"26886","DOI":"10.1039\/D5RA02682B","article-title":"Laccase-immobilized\nbiochar as a unique host matrix for electrochemical detection of gallic\nacid: a sustainable engineering approach","volume":"15","author":"Thomas","year":"2025","journal-title":"RSC\nAdv."},{"key":"2026083119150170700_cit54","doi-asserted-by":"publisher","first-page":"33595","DOI":"10.1021\/acsomega.5c04276","article-title":"Biochar for Electrochemical Sulfanilamide Detection:\nComparative Evaluation of Carbon Paste and Screen-Printed Electrodes","volume":"10","author":"Cabral","year":"2025","journal-title":"ACS Omega"},{"key":"2026083119150170700_cit55","doi-asserted-by":"publisher","first-page":"607","DOI":"10.1177\/0263617419867519","article-title":"Effect of nitric\nacid oxidation on the surface of hydrochars to sorb methylene blue:\nAn adsorption mechanism comparison","volume":"37","author":"Nguyen","year":"2019","journal-title":"Adsorpt.\nSci. Technol."},{"key":"2026083119150170700_cit56","doi-asserted-by":"publisher","DOI":"10.1016\/j.chemosphere.2022.134422","article-title":"The molecular markers provide complementary information for biochar\ncharacterization before and after HNO(3)\/H(2)SO(4) oxidation","volume":"301","author":"Chang","year":"2022","journal-title":"Chemosphere"},{"key":"2026083119150170700_cit57","doi-asserted-by":"publisher","first-page":"350","DOI":"10.1016\/j.jaap.2017.07.015","article-title":"Evolution of the chemical composition, functional group,\npore structure and crystallographic structure of bio-char from palm\nkernel shell pyrolysis under different temperatures","volume":"127","author":"Ma","year":"2017","journal-title":"J. Anal. Appl. Pyrolysis"},{"key":"2026083119150170700_cit58","doi-asserted-by":"publisher","DOI":"10.3390\/su16177497","article-title":"Selective\nPhenolics Recovery from Aqueous Residues of Pyrolysis Oil through\nComputationally Designed Green Solvent","volume":"16","author":"Qaisar","year":"2024","journal-title":"Sustainability"},{"key":"2026083119150170700_cit59","doi-asserted-by":"publisher","DOI":"10.1016\/j.jece.2025.115477","article-title":"Mutual interaction\nof pyrolysis operating conditions and surface morphology for the electrochemical\nperformance of biochar-modified screen-printed electrodes","volume":"13","author":"Cancelliere","year":"2025","journal-title":"J. Environ. Chem. 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