{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,12]],"date-time":"2026-03-12T04:05:03Z","timestamp":1773288303972,"version":"3.50.1"},"reference-count":36,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2022,3,16]],"date-time":"2022-03-16T00:00:00Z","timestamp":1647388800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["JCM"],"abstract":"<jats:p>The aim was to evaluate the antibacterial efficacy and penetration depth into dentinal tubules of a solution of chitosan nanodroplets (NDs) loaded with Benzalkonium Chloride (BAK). Seventy-two human single-root teeth with fully formed apex were used. Cylindrical root dentin blocks were longitudinally sectioned and enlarged to a size of a Gates Glidden drill #4. After sterilization, root canals were infected with Enterococcus faecalis ATCC 29212 and further incubated for three weeks. Specimens were assigned to three experimental groups (n = 20), plus positive (n = 6) and negative (n = 6) controls. In the first group, irrigation was achieved with 2 mL of NDs solution loaded with BAK (NDs-BAK), in the second with 2 mL of 5% sodium hypochlorite (NaOCl) and in the last with 2 mL of 2% chlorhexidine (CHX). Specimens were rinsed and vertically fractured. Confocal laser scanning microscopy (CLSM) and viability staining were used to analyze the proportions of dead and live bacteria quantitatively. The volume ratio of red fluorescence (dead) was calculated in 3D reconstructions. Data were analyzed by one-way ANOVA and post hoc Bonferroni tests (p &lt; 0.05). The ratio of red fluorescence over the whole green\/red fluorescence resulted in a significant comparison of NDs-BAK with NaOCl (p &lt; 0.01) and NaOCl with CHX (p &lt; 0.01). No differences were found between NDs-BAK and CHX (p &gt; 0.05). The mean depth of efficacy was, respectively: NDs-BAK 325.25 \u03bcm, NaOCl 273.36 \u03bcm and CHX 246.78 \u03bcm with no statistical differences between groups. The NaOCl solution showed the highest antimicrobial efficacy, but nanodroplets with BAK seemed to have the same effect as CHX with a high depth of efficacy.<\/jats:p>","DOI":"10.3390\/jcm11061650","type":"journal-article","created":{"date-parts":[[2022,3,16]],"date-time":"2022-03-16T22:09:58Z","timestamp":1647468598000},"page":"1650","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Confocal Laser Scanner Evaluation of Bactericidal Effect of Chitosan Nanodroplets Loaded with Benzalkonium Chloride"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2523-8149","authenticated-orcid":false,"given":"Mario","family":"Alovisi","sequence":"first","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0365-0626","authenticated-orcid":false,"given":"Damiano","family":"Pasqualini","sequence":"additional","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2949-4556","authenticated-orcid":false,"given":"Narcisa","family":"Mandras","sequence":"additional","affiliation":[{"name":"Department of Public Health and Pediatrics, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2542-622X","authenticated-orcid":false,"given":"Janira","family":"Roana","sequence":"additional","affiliation":[{"name":"Department of Public Health and Pediatrics, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Pietro","family":"Costamagna","sequence":"additional","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4057-1428","authenticated-orcid":false,"given":"Allegra","family":"Comba","sequence":"additional","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Roberta","family":"Cavalli","sequence":"additional","affiliation":[{"name":"Department of Drug Science and Technology, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4256-5774","authenticated-orcid":false,"given":"Anna","family":"Luganini","sequence":"additional","affiliation":[{"name":"Department of Life Sciences and Systems Biology, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alfredo","family":"Iandolo","sequence":"additional","affiliation":[{"name":"Department of Medicine, Surgery and Dentistry, Salerno Medical School, University of Salerno, 84084 Salerno, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8701-8486","authenticated-orcid":false,"given":"Lorenza","family":"Cavallo","sequence":"additional","affiliation":[{"name":"Department of Public Health and Pediatrics, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7380-1675","authenticated-orcid":false,"given":"Nicola","family":"Scotti","sequence":"additional","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elio","family":"Berutti","sequence":"additional","affiliation":[{"name":"Department of Surgical Sciences, Dental School, University of Turin, 10126 Turin, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,3,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1034\/j.1601-1546.2002.20101.x","article-title":"Endodontic epidemiology and treatment outcome: General considerations","volume":"2","author":"Eriksen","year":"2002","journal-title":"Endod. Top."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"712","DOI":"10.1111\/iej.13467","article-title":"The global prevalence of apical periodontitis: A systematic review and meta-analysis","volume":"54","author":"Michelon","year":"2021","journal-title":"Int. Endod. J."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1170","DOI":"10.1016\/j.joen.2012.05.023","article-title":"Prevalence of periapical radiolucency and root canal treatment: A systematic review of cross-sectional studies","volume":"38","author":"Pak","year":"2012","journal-title":"J. Endod."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1097\/01.DON.0000129039.59003.9D","article-title":"Current challenges and concepts in the preparation of root canal systems: A review","volume":"30","author":"Peters","year":"2004","journal-title":"J. Endod."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"54","DOI":"10.1007\/s10266-021-00631-2","article-title":"Micro-CT evaluation of rotary and reciprocating glide path and shaping systems outcomes in maxillary molar curved canals","volume":"110","author":"Alovisi","year":"2021","journal-title":"Odontology"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"30","DOI":"10.1111\/j.1601-1546.2005.00152.x","article-title":"Mechanical preparation of root canals: Shaping goals, techniques and means","volume":"10","author":"Hulsmann","year":"2005","journal-title":"Endod. Top."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1860","DOI":"10.1016\/j.joen.2014.06.023","article-title":"Root Canal Preparation of Mandibular Molars with 3 Nickel-Titanium Rotary Instruments: A Micro-Computed Tomographic Study","volume":"40","author":"Zhao","year":"2014","journal-title":"J. Endod."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"387","DOI":"10.1111\/iej.12628","article-title":"Micro-CT evaluation of several glide path techniques and ProTaper Next shaping outcomes in maxillary first molar curved canals","volume":"50","author":"Alovisi","year":"2017","journal-title":"Int. Endod. J."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1016\/j.joen.2005.09.014","article-title":"Root canal irrigants","volume":"32","author":"Zehnder","year":"2006","journal-title":"J. Endod."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1016\/j.cden.2009.12.001","article-title":"Irrigation in endodontics","volume":"54","author":"Haapasalo","year":"2010","journal-title":"Dent. Clin. N. Am."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"791","DOI":"10.1016\/j.joen.2009.03.010","article-title":"Review of contemporary irrigant agitation techniques and devices","volume":"35","author":"Gu","year":"2009","journal-title":"J. Endod."},{"key":"ref_12","first-page":"9","article-title":"Irrigants for non-surgical root canal treatment in mature permanent teeth","volume":"12","author":"Fedorowicz","year":"2012","journal-title":"Cochrane Database Syst. Rev."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1131","DOI":"10.1111\/iej.13307","article-title":"The influence of the addition of surfactants to sodium hypochlorite on the removal of hard tissue debris","volume":"53","author":"Guerreiro","year":"2020","journal-title":"Int. Endod. J."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"288","DOI":"10.1111\/j.1365-2591.2008.01540.x","article-title":"The properties and applications of chlorhexidine in endodontics","volume":"42","author":"Mohammadi","year":"2009","journal-title":"Int. Endod. J."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1525","DOI":"10.1016\/j.joen.2012.07.008","article-title":"Wetting properties and critical micellar concentration of benzalkonium chloride mixed in sodium hypochlorite","volume":"38","author":"Bukiet","year":"2012","journal-title":"J. Endod."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"821","DOI":"10.1016\/j.joen.2012.03.012","article-title":"Decreased bacterial adherence and biofilm growth on surfaces coated with a solution of benzalkonium chloride","volume":"38","author":"Jaramillo","year":"2012","journal-title":"J. Endod."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"846","DOI":"10.1016\/j.jdent.2015.05.003","article-title":"The effect of benzalkonium chloride additions to AH Plus sealer. Antimicrobial, physical and chemical properties","volume":"43","author":"Cassar","year":"2015","journal-title":"J. Dent."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1417","DOI":"10.1016\/j.joen.2016.05.021","article-title":"Antibacterial nanoparticles in endodontics: A review","volume":"42","author":"Shrestha","year":"2016","journal-title":"J. Endod."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1007\/s11095-018-2371-z","article-title":"\u2018In Vitro\u2019, \u2018In Vivo\u2019 and \u2018In Silico\u2019 investigation of the anticancer effectiveness of oxygen-loaded chitosan-shelled nanodroplets as potential drug vector","volume":"35","author":"Khadjavi","year":"2018","journal-title":"Pharm. Res."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1275","DOI":"10.1016\/j.joen.2012.05.006","article-title":"The effect of tissue inhibitors on the antibacterial activity of chitosan nanoparticles and photodynamic therapy","volume":"38","author":"Shrestha","year":"2012","journal-title":"J. Endod."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"929","DOI":"10.2217\/fmb.15.27","article-title":"Antimicrobial chitosan nanodroplets: New insights for ultrasound-mediated adjuvant treatment of skin infection","volume":"10","author":"Banche","year":"2015","journal-title":"Future Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Argenziano, M., Bressan, B., Luganini, A., Finesso, N., Genova, T., Troia, A., Giribaldi, G., Banche, G., Mandras, N., and Cuffini, A.M. (2021). Comparative evaluation of different chitosan species and derivatives as candidate biomaterials for oxygen-loaded nanodroplet formulations to treat chronic wounds. Mar. Drugs, 19.","DOI":"10.3390\/md19020112"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1198","DOI":"10.1016\/j.joen.2008.07.001","article-title":"Confocal laser scanning microscopy is appropriate to detect viability of Enterococcus faecalis in infected dentin","volume":"34","author":"Zapata","year":"2008","journal-title":"J. Endod."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"644","DOI":"10.1111\/j.1365-2591.2011.01867.x","article-title":"Bacterial localization and viability assessment in human ex vivo dentinal tubules by fluorescence confocal laser scanning microscopy","volume":"44","author":"Parmar","year":"2011","journal-title":"Int. Endod. J."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1380","DOI":"10.1016\/j.joen.2011.06.018","article-title":"A new noninvasive model to study the effectiveness of dentin disinfection by using confocal laser scanning microscopy","volume":"37","author":"Ma","year":"2011","journal-title":"J. Endod."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1111\/j.1601-1546.2005.00135.x","article-title":"Eradication of endodontic infection by instrumentation and irrigation solutions","volume":"10","author":"Haapasalo","year":"2005","journal-title":"Endod. Top."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Mandras, N., Pasqualini, D., Roana, J., Tullio, V., Banche, G., Gianello, E., Bonino, F., Cuffini, A.M., Berutti, E., and Alovisi, M. (2020). Influence of Photon-Induced Photoacoustic Streaming (PIPS) on root canal disinfection and post-operative pain: A randomized clinical trial. J. Clin. Med., 9.","DOI":"10.3390\/jcm9123915"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"281","DOI":"10.1067\/moe.2002.126163","article-title":"Endodontic infections: Concepts, paradigms, and perspectives","volume":"94","author":"Siqueira","year":"2002","journal-title":"Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endodontol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1134","DOI":"10.1016\/j.joen.2011.04.013","article-title":"Biofilm dissolution and cleaning ability of different irrigant solutions on intraorally infected dentin","volume":"37","author":"Bramante","year":"2011","journal-title":"J. Endod."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"108","DOI":"10.4103\/JCD.JCD_165_19","article-title":"Evaluation of the actual chlorine concentration and the required time for pulp dissolution using different sodium hypochlorite irrigating solutions","volume":"22","author":"Iandolo","year":"2019","journal-title":"J. Conserv. Dent."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"e29","DOI":"10.1016\/j.dental.2012.10.002","article-title":"Incorporation of antimicrobial agents can be used to enhance the antibacterial effect of endodontic sealers","volume":"29","author":"Gjorgievska","year":"2013","journal-title":"Dent. Mater."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Komiyama, E.Y., Lepesqueur, L.S., Yassuda, C.G., Samaranayake, L.P., Parahitiyawa, N.B., Balducci, I., and Koga-Ito, C.Y. (2016). Enterococcus species in the oral cavity: Prevalence, virulence factors and antimicrobial susceptibility. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0163001"},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Mandras, N., Alovisi, M., Roana, J., Crosasso, P., Luganini, A., Pasqualini, D., Genta, E., Arpicco, S., Banche, G., and Cuffini, A. (2020). Evaluation of the bactericidal activity of a hyaluronic acid-vehicled clarithromycin antibiotic mixture by confocal laser scanning microscopy. Appl. Sci., 10.","DOI":"10.3390\/app10080761"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"38","DOI":"10.1159\/000510198","article-title":"Biofilm Models for the Evaluation of Dental Treatment","volume":"29","author":"Eick","year":"2021","journal-title":"Monogr. Oral Sci."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1016\/j.joen.2015.08.035","article-title":"Effect of a Benzalkonium Chloride Surfactant-Sodium Hypochlorite Combination on Elimination of Enterococcus faecalis","volume":"42","author":"Baron","year":"2016","journal-title":"J. Endod."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1376","DOI":"10.1016\/j.joen.2012.06.035","article-title":"Effectiveness of endodontic disinfecting solutions against young and old Enterococcus faecalis biofilms in dentin canals","volume":"38","author":"Wang","year":"2012","journal-title":"J. Endod."}],"container-title":["Journal of Clinical Medicine"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2077-0383\/11\/6\/1650\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T22:37:26Z","timestamp":1760135846000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2077-0383\/11\/6\/1650"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,3,16]]},"references-count":36,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2022,3]]}},"alternative-id":["jcm11061650"],"URL":"https:\/\/doi.org\/10.3390\/jcm11061650","relation":{},"ISSN":["2077-0383"],"issn-type":[{"value":"2077-0383","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,3,16]]}}}