Effect of Low-Temperature Intracanal Sodium Hypochlorite on Root Surface Temperature Reduction and Organic Matter Dissolution: An In Vitro Study

Effect of Low-Temperature Intracanal Sodium Hypochlorite on Root Surface Temperature Reduction and Organic Matter Dissolution: An In Vitro Study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Effect of Low-Temperature Intracanal Sodium Hypochlorite on Root Surface Temperature Reduction and Organic Matter Dissolution: An In Vitro Study Marcos Felipe Iparraguirre Nunovero, Marco Antônio Hungaro Duarte, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7811504/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 31 Jan, 2026 Read the published version in Scientific Reports → Version 1 posted 14 You are reading this latest preprint version Abstract Introduction: This study aimed to evaluate the effect of low-temperature intracanal sodium hypochlorite (NaOCl) irrigation on root surface temperature reduction and its ability to dissolve organic matter. Methods Twenty-five mandibular premolars were accessed and instrumented to size 40.05. Final irrigation protocols with 20 mL of 2.5% NaOCl, at two different temperatures: room temperature (control) and 2.5°C (experimental), were applied. Initial and minimum root surface temperatures (last 4 mm of the root) were recorded using a digital thermometer. For tissue dissolution analysis, glass capillaries filled with catgut were attached to the cervical and apical thirds of twenty 3D-printed maxillary incisors and weighed before and after the same irrigation protocols. Data was statistically analyzed. Results Low-temperature NaOCl irrigation led to a significantly greater reduction in root surface temperature (p 0.05). Conclusion Low-temperature NaOCl effectively reduced root surface temperature while maintaining its tissue-dissolving capacity compared to the control group. Clinical relevance: Cold sodium hypochlorite effectively reduces external root surface temperature while maintaining its ability to dissolve organic tissue, providing clinicians with a practical and efficient intracanal cryotherapy strategy for minimizing postoperative inflammation without compromising chemo-mechanical preparation. Health sciences/Health care Health sciences/Medical research Endodontics Cryotherapy Sodium hypochlorite Tissue dissolution Temperature Figures Figure 1 Introduction The biomechanical preparation of the root canal system involves the use of manual or mechanical instruments combined with auxiliary chemical irrigants. Sodium hypochlorite (NaOCl) is widely used in endodontic therapy due to its bactericidal properties and ability to dissolve organic tissue [ 1 , 2 ]. However, its antibacterial activity and tissue-dissolving capacity depend on its concentration, volume, and contact time [ 3 , 4 ]. High concentrations of NaOCl may be toxic to periapical tissues and can cause severe irritation if extruded beyond the apex [ 4 , 5 ]. To minimize these risks, negative pressure irrigation techniques have been introduced in endodontics to improve irrigant delivery in the apical region while reducing the risk of extrusion [ 6 ]. These methods allow for increased irrigant flow and more effective debridement compared to conventional syringe irrigation [ 1 ]. Cryotherapy has been used therapeutically since around 3000 B.C. to moderate inflammation. Its application expanded over the centuries and was even recommended by Hippocrates in Ancient Greece [ 7 – 9 ]. Cryotherapy works by extracting heat from warmer tissues to a cooler environment, thereby reducing inflammation and blood flow, inhibiting neural receptors, and lowering metabolic activity [ 9 , 10 ]. In dentistry, cryotherapy is commonly employed following oral surgical procedures—such as extractions and implant placement—to reduce pain and swelling, as well as in the management of temporomandibular joint disorders [ 8 ]. Despite its frequent use, strong evidence supporting its precise mechanism of action remains limited due to lack of standardization regarding application time, duration, method, and the cooling agent used [ 7 , 11 ]. Cryotherapy has gained attention in endodontics as a low-cost, simple, and non-toxic technique. Several randomized clinical trials have investigated its efficacy in relieving postoperative pain following root canal treatment [ 10 , 12 , 13 ]. One method of delivering cryotherapy to inflamed periradicular tissues is through intracanal irrigation with cold solutions, such as chilled saline, after canal instrumentation [ 7 , 9 , 10 ]. This approach has been shown to reduce the external root surface temperature by more than 10°C, potentially producing anti-inflammatory effects on periapical tissues [ 7 , 13 – 16 ]. NaOCl has demonstrated similar antibacterial activity across a range of temperatures [ 16 ]. While heating the solution has been shown to enhance its tissue-dissolving capacity [ 17 ], there are no studies assessing its performance at lower temperatures. Therefore, the present study aimed to determine whether continuous irrigation with low-temperature NaOCl (2.5°C) could reduce the external temperature of the apical 4 mm of the root surface, and to evaluate the organic matter–dissolving capability of NaOCl at low temperature compared to room temperature. Materials and Methods Root Surface Temperature One hundred extracted human mandibular premolars were evaluated via digital radiographs in buccolingual and mesiodistal projections. Twenty-five teeth with Vertucci Type I canal configuration, apical curvature less than 10° and with an anatomical foramen of tooth (assessed during root canal treatment) < #40 K type instrument. Crowns were sectioned to standardize root length at 15 mm. The specimens were immersed in 5% NaOCl for 30 minutes, followed by rinsing in distilled water to prevent dehydration. Access cavities were prepared, and canals were preflared using ProTaper SX (DentsplySirona, Ballaigues, Switzerland). Irrigation was performed with 2.5% NaOCl, and apical patency was confirmed with a size #15 K-file. Working length (WL) was determined visually by inserting a #15 K-file until its tip was visible at the apical foramen under 10x magnification. Canals were instrumented using Reciproc R40 (VDW, Munich, Germany). Irrigation was performed with 2 mL of 2.5% NaOCl, followed by a final rinse with 17% EDTA for 1 minute. Canals were dried with sterile paper points. Roots were inserted into polypropylene tubes exposing the apical 4 mm and isolated with rubber dam and Top Dam resin (Fig. 1 -A) (FGM Dental Group, Santa Catarina, Brazil). K-type thermocouples (TP-01, Thomas Edison Co, Wenzhou, China; range 50°C–1350°C) connected to a digital thermometer (A-PlusType K RoHS, Thomas Edison Co) were attached to the apical root surface. (Fig. 1 -B) Each specimen underwent two irrigation protocols: Control Group – 20 mL of 2.5% NaOCl at room temperature (25°C), delivered over 5 minutes using the EndoVac system. Both the Microcannula and syringe were at room temperature. Experimental Group – 20 mL of 2.5% NaOCl at 2.5°C, delivered over 5 minutes using the EndoVac system. Both the Microcannula and syringe were pre-cooled and maintained at 2.5°C in a calibrated refrigerator until use. (Fig. 1 -A) Initial and minimum surface temperatures were recorded during each irrigation session. Tissue Dissolution Twenty 3D-printed maxillary central incisors (IM do Brasil, São Paulo, Brazil) were used. Access cavities and canal instrumentation were completed. Two small openings were created on the external root surface—one in the cervical third and one in the apical third—to connect the canal to the external environment. Glass capillaries (0.4 mm internal diameter, 10 mm length) were embedded in the openings using light-cured flowable composite resin to ensure stabilization. Capillaries were filled with 4.0 chromic gut sutures (Bioline, São Paulo, Brazil) [ 2 ]. (Fig. 1 -C) Teeth were randomly divided into two groups, following the same final irrigation protocols as described above (room-temperature or cold NaOCl, both delivered over 5 minutes with the EndoVac system). Capillaries and microcannulas were maintained at the respective temperatures prior to use. (Fig. 1 -D) Capillaries were dried in a hot-air oven at 50°C for 15 minutes and weighed on a precision scale (Shimadzu, Barueri, Brazil) before and after irrigation. Tissue dissolution was calculated by subtracting final weight from initial weight. Statistical Analysis Root Surface Temperature: Data were tested for normality using the Kolmogorov–Smirnov test. Paired t -tests were used to compare initial and minimum temperatures between groups. Tissue Dissolution Data were first tested for normality using the Kolmogorov–Smirnov and Shapiro–Wilk tests. Since final weight values were not normally distributed, the nonparametric Mann–Whitney U test was applied. After confirming the normal distribution of “Initial weight” and “weight loss” variables, they were compared using independent samples T Student test. Results Regarding root surface temperature, the control group exhibited an average decrease of nearly 1°C after irrigation with 2.5% NaOCl at room temperature ( p < 0.05). In contrast, the experimental group, which received NaOCl at 2.5°C, showed a mean temperature reduction of approximately 8°C ( p < 0.05). The temperature drop observed in the experimental group was significantly greater than that in the control group ( p < 0.05) (Table 1 ). Table 1 Protocol, mean and standard deviation (°C) of the initial temperature and the lowest temperature reached during the procedure. Protocol Time point n Media (°C) DP Control Initial 25 24.94 a 1,31 Lowest 25 24.04 b 1,28 Experimental Initial 25 24.81 a 1,02 Lowest 25 16.52 c 2,36 For tissue dissolution, no statistically significant difference was observed between groups, regardless of solution temperature ( p > 0.05) (Table 2 ). Both protocols demonstrated similar ability to dissolve organic matter. Table 2 – Protocol, mean and standard deviation of the weight (mg) before and after the irrigation protocols, and the percentage of dissolution of the analyzed groups. Protocol Weight Media ± DS (mg) % Dissolution Control Initial 50.6 ± 6.48 a 1.8 ± 0.79 A Final 49.7 ± 6.49 a Experimental Initial 46.9 ± 2.81 a 1.71 ± 0.69 A Final 46.1 ± 2.85 a Discussion Cryotherapy aims to elicit physiological responses such as reduced blood flow, neural receptor inhibition, and decreased metabolic activity [ 7 ]. Its adaptation to intracanal application has emerged as a cost-effective, localized therapeutic strategy for managing inflammation and postoperative pain following endodontic treatment [ 9 ]. Intracanal cryotherapy has been indicated to reduce postoperative pain in patients with symptomatic irreversible pulpitis [ 16 , 19 ] or to prevent pain in asymptomatic individuals undergoing root canal treatment [ 14 ]. In both cases, cryotherapy has been shown to reduce the incidence and severity of postoperative pain, particularly during the first few hours after treatment [ 12 , 14 , 19 ]. The most widely recommended cryotherapy protocol in the literature involves final irrigation with 20 mL of cold saline (2°–4°C) for 5 minutes [ 9 , 19 ]. In the present study, cold NaOCl was used instead of saline due to its widespread clinical use and well-established properties, including its antimicrobial activity, tissue-dissolving capacity, and ability to penetrate dentinal tubules [ 2 , 18 – 22 ]. Negative pressure irrigation (EndoVac) was employed to ensure consistent delivery to the working length while minimizing the risk of irrigant extrusion. Using cold NaOCl may simplify clinical protocols by eliminating the need to switch to a different irrigant for cryotherapy. In 2015, Vera et al. reported that cryotherapy with cold saline reduced root surface temperatures by up to 14.3°C over 4 minutes [ 7 ]. In contrast, the present study observed a maximum reduction of 8.3°C with cold NaOCl. This difference may be explained by a higher ambient temperature in the current study (25°C vs. ~23°C), as well as the potential exothermic nature of NaOCl’s tissue-dissolving reaction. NaOCl degrades fatty acids through saponification, producing soap and glycerol, which may release heat and slightly limit the cooling effect during irrigation [ 23 ]. A 2024 study by de Oliveira et al. evaluated tissue dissolution in simulated complex anatomies and found that ultrasonic activation enhanced NaOCl's ability to dissolve organic matter [ 2 ]. This might be explained by the increased efficiency of heated NaOCl, resulting from an accelerated reaction rate and improved irrigant flow due to ultrasonic agitation [ 2 , 17 ]. In contrast, the present study showed no significant difference in tissue dissolution between cold and room-temperature NaOCl. This discrepancy may be attributed to the much greater amount of organic material used by Oliveira et al. (> 700 mg per specimen) compared to the amount used in the present study (< 51 mg per specimen). To our knowledge, no previous studies have evaluated the tissue-dissolving ability of NaOCl at low temperatures in simulated complex anatomies. Within the limitations of this in vitro study, our findings suggest that negative pressure irrigation with cold NaOCl can reduce external root surface temperature by over 8°C and maintain this reduction for a sufficient duration to potentially elicit periapical physiological effects. Furthermore, low-temperature NaOCl demonstrated a comparable tissue-dissolving capacity to that of room-temperature NaOCl. Conclusion Intracanal irrigation with 2.5% sodium hypochlorite at 2.5°C significantly reduced external root surface temperature without compromising its ability to dissolve organic matter, when compared with room-temperature NaOCl. This suggests that cold NaOCl may serve as an effective alternative for intracanal cryotherapy. Declarations Author contributions Conceptualization: M.I.N, E.C. Methodology: M.I.N., P.G.T., U.X.S. Data collection and curation: M.H.D., M.I.N., P.G.T. Analysis and interpretation: U.X.S., V.P.W., B.C.C., M.H.D. Visualization: U.X.S., V.P.W., E.C. Writing – Original Draft Preparation: M.I.N., E.C. Writing – Reviewing and editing: E.C., B.C.C., M.H.D., V.P.W., U.X.S. Supervision: V.P.W., U.X.S., E.C. All authors reviewed and approved the final manuscript. Funding This study was funded by the “Conselho Nacional de Desenvolvimento Científico e Tecnológico” (CNPq, Brasilia, Brazil) as part of a doctoral research incentive grant. (Number:1610162020-0) Data Availability The dataset generated and analyzed during the current study, including the table of extracted articles, are available from the corresponding author upon request. Ethical approval This study was approved by the local ethics committee (protocol: 5.024.031). Competing Interests The authors declare they have no competing interest related to this study. References Mancini M, Cerroni L, Iorio L, et al. (2013) Smear layer removal and canal cleanliness using different irrigation systems: SEM evaluation. J Endod 39(11):1456–1460. de Oliveira Neto RS, de Souza Lima LA, Titato PCG, et al. (2023) Effectiveness of a new endodontic irrigation system for removing smear layer and dissolving simulated organic matter. Clin Oral Investig 28(1):1–6. Moorer WR, Wesselink PR (1982) Factors promoting the tissue dissolving capability of sodium hypochlorite. Int Endod J 15(4):187–96. Stojicic S, Zivkovic S, Qian W, et al. (2010) Tissue dissolution by sodium hypochlorite: effect of concentration, temperature, agitation, and surfactant. J Endod 36(9):1558–1562. Zehnder M. Root canal irrigants (2006) J Endod 32(5):389–398. Haapasalo M, Shen Y, Wang Z, Gao Y (2014) Irrigation in endodontics. Br Dent J 216(6):299–303. Vera J, Ochoa J, Vazquez-Carcaño M, et al. 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(2024) Intracanal cryotherapy with or without foraminal enlargement. Sci Rep 14(1):19905. Nandakumar M, Nasim I (2020) Cryotreated NaOCl and postoperative pain: RCT. J Conserv Dent Endod 23(2):131–136. Lalfakawmi S, Gupta A, Duraisamy AK, et al. (2024) Impact of Cryotreated and Warm Sodium Hypochlorite on Postoperative Pain in Teeth with Symptomatic Irreversible Pulpitis: A Randomized Controlled. J Endod. 50(11):1543–1550. de Hemptinne F, Slaus G, Vandendael M, Jacquet W, De Moor R J, Bottenberg P (2015) In vivo intracanal temperature evolution during endodontic treatment after the injection of room temperature or preheated sodium hypochlorite. J Endod 41(7):1112-1115. Mokhtari H, Milani AS, Zand V, et al. (2023) Temperature of NaOCl and postoperative pain. Clin Exp Dent Res 9(5):859–867. Vera J, Ochoa J, Romero M, et al. (2018) Cryotherapy in symptomatic apical periodontitis: multicenter RCT. J Endod 44(1):4–8. Karataş E, Ayaz N, Uluköylü E, et al. (2021) Effect of NaOCl temperature on postoperative pain. J Appl Oral Sci 29:1–8. Clegg MS, Vertucci FJ, Walker C, et al. (2006) Irrigant effects on apical biofilms. J Endod 32(5):434–437. Wong DT, Cheung GS (2014) NaOCl penetration into dentinal tubules. J Endod 40(6):825–829. Estrela C, Estrela CR, Barbin EL, et al. (2002) Mechanism of action of sodium hypochlorite. Br Dent J 13:113–117. Sun M, Liu S, Zhang Y, et al. (2019) Thermodynamics of saponification reactions. J Mol Liq 280:252–258. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 31 Jan, 2026 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 07 Dec, 2025 Reviewers agreed at journal 06 Dec, 2025 Reviewers agreed at journal 03 Dec, 2025 Reviews received at journal 03 Dec, 2025 Reviews received at journal 02 Dec, 2025 Reviewers agreed at journal 01 Dec, 2025 Reviewers agreed at journal 01 Dec, 2025 Reviewers agreed at journal 01 Dec, 2025 Reviewers agreed at journal 01 Dec, 2025 Reviewers invited by journal 01 Dec, 2025 Editor invited by journal 10 Oct, 2025 Editor assigned by journal 09 Oct, 2025 Submission checks completed at journal 09 Oct, 2025 First submitted to journal 08 Oct, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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1","display":"","copyAsset":false,"role":"figure","size":465775,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eA\u003c/strong\u003e: Irrigation protocol using the EndoVac system for the Control and Experimental groups, performed on a polypropylene platform isolated with a rubber dam. \u003cstrong\u003eB\u003c/strong\u003e: Thermocouple of the digital thermometer adapted to the apical 4 mm of the root surface. \u003cstrong\u003eC\u003c/strong\u003e: Prototyped incisor with capillaries filled with catgut to simulate an anatomically complex area. \u003cstrong\u003eD\u003c/strong\u003e: Irrigation protocol using the EndoVac system for the Control and Experimental groups in the prototyped incisor with pre-attached capillaries.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7811504/v1/b43b6fde2c87708f775327b3.png"},{"id":101692467,"identity":"03003952-cbc1-4fae-8b6a-da88db2650aa","added_by":"auto","created_at":"2026-02-02 16:19:46","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1160620,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7811504/v1/097e51f8-03ed-44a8-81f5-d6df4db0f5a7.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Effect of Low-Temperature Intracanal Sodium Hypochlorite on Root Surface Temperature Reduction and Organic Matter Dissolution: An In Vitro Study","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe biomechanical preparation of the root canal system involves the use of manual or mechanical instruments combined with auxiliary chemical irrigants. Sodium hypochlorite (NaOCl) is widely used in endodontic therapy due to its bactericidal properties and ability to dissolve organic tissue [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. However, its antibacterial activity and tissue-dissolving capacity depend on its concentration, volume, and contact time [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eHigh concentrations of NaOCl may be toxic to periapical tissues and can cause severe irritation if extruded beyond the apex [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. To minimize these risks, negative pressure irrigation techniques have been introduced in endodontics to improve irrigant delivery in the apical region while reducing the risk of extrusion [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. These methods allow for increased irrigant flow and more effective debridement compared to conventional syringe irrigation [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eCryotherapy has been used therapeutically since around 3000 B.C. to moderate inflammation. Its application expanded over the centuries and was even recommended by Hippocrates in Ancient Greece [\u003cspan additionalcitationids=\"CR8\" citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Cryotherapy works by extracting heat from warmer tissues to a cooler environment, thereby reducing inflammation and blood flow, inhibiting neural receptors, and lowering metabolic activity [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIn dentistry, cryotherapy is commonly employed following oral surgical procedures\u0026mdash;such as extractions and implant placement\u0026mdash;to reduce pain and swelling, as well as in the management of temporomandibular joint disorders [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Despite its frequent use, strong evidence supporting its precise mechanism of action remains limited due to lack of standardization regarding application time, duration, method, and the cooling agent used [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eCryotherapy has gained attention in endodontics as a low-cost, simple, and non-toxic technique. Several randomized clinical trials have investigated its efficacy in relieving postoperative pain following root canal treatment [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. One method of delivering cryotherapy to inflamed periradicular tissues is through intracanal irrigation with cold solutions, such as chilled saline, after canal instrumentation [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. This approach has been shown to reduce the external root surface temperature by more than 10\u0026deg;C, potentially producing anti-inflammatory effects on periapical tissues [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan additionalcitationids=\"CR14 CR15\" citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eNaOCl has demonstrated similar antibacterial activity across a range of temperatures [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. While heating the solution has been shown to enhance its tissue-dissolving capacity [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], there are no studies assessing its performance at lower temperatures. Therefore, the present study aimed to determine whether continuous irrigation with low-temperature NaOCl (2.5\u0026deg;C) could reduce the external temperature of the apical 4 mm of the root surface, and to evaluate the organic matter\u0026ndash;dissolving capability of NaOCl at low temperature compared to room temperature.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eRoot Surface Temperature\u003c/h2\u003e\u003cp\u003eOne hundred extracted human mandibular premolars were evaluated via digital radiographs in buccolingual and mesiodistal projections. Twenty-five teeth with Vertucci Type I canal configuration, apical curvature less than 10\u0026deg; and with an anatomical foramen of tooth (assessed during root canal treatment) \u0026lt; #40 K type instrument. Crowns were sectioned to standardize root length at 15 mm. The specimens were immersed in 5% NaOCl for 30 minutes, followed by rinsing in distilled water to prevent dehydration.\u003c/p\u003e\u003cp\u003eAccess cavities were prepared, and canals were preflared using ProTaper SX (DentsplySirona, Ballaigues, Switzerland). Irrigation was performed with 2.5% NaOCl, and apical patency was confirmed with a size #15 K-file. Working length (WL) was determined visually by inserting a #15 K-file until its tip was visible at the apical foramen under 10x magnification. Canals were instrumented using Reciproc R40 (VDW, Munich, Germany). Irrigation was performed with 2 mL of 2.5% NaOCl, followed by a final rinse with 17% EDTA for 1 minute. Canals were dried with sterile paper points.\u003c/p\u003e\u003cp\u003eRoots were inserted into polypropylene tubes exposing the apical 4 mm and isolated with rubber dam and Top Dam resin (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e-A) (FGM Dental Group, Santa Catarina, Brazil). K-type thermocouples (TP-01, Thomas Edison Co, Wenzhou, China; range 50\u0026deg;C\u0026ndash;1350\u0026deg;C) connected to a digital thermometer (A-PlusType K RoHS, Thomas Edison Co) were attached to the apical root surface. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e-B)\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eEach specimen underwent two irrigation protocols:\u003c/p\u003e\u003cp\u003e\u003col\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003e\u003cb\u003eControl Group\u003c/b\u003e \u0026ndash; 20 mL of 2.5% NaOCl at room temperature (25\u0026deg;C), delivered over 5 minutes using the EndoVac system. Both the Microcannula and syringe were at room temperature.\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003e\u003cb\u003eExperimental Group\u003c/b\u003e \u0026ndash; 20 mL of 2.5% NaOCl at 2.5\u0026deg;C, delivered over 5 minutes using the EndoVac system. Both the Microcannula and syringe were pre-cooled and maintained at 2.5\u0026deg;C in a calibrated refrigerator until use. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e-A)\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003c/ol\u003e\u003c/p\u003e\u003cp\u003eInitial and minimum surface temperatures were recorded during each irrigation session.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eTissue Dissolution\u003c/h3\u003e\n\u003cp\u003eTwenty 3D-printed maxillary central incisors (IM do Brasil, S\u0026atilde;o Paulo, Brazil) were used. Access cavities and canal instrumentation were completed. Two small openings were created on the external root surface\u0026mdash;one in the cervical third and one in the apical third\u0026mdash;to connect the canal to the external environment.\u003c/p\u003e\u003cp\u003eGlass capillaries (0.4 mm internal diameter, 10 mm length) were embedded in the openings using light-cured flowable composite resin to ensure stabilization. Capillaries were filled with 4.0 chromic gut sutures (Bioline, S\u0026atilde;o Paulo, Brazil) [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e-C)\u003c/p\u003e\u003cp\u003eTeeth were randomly divided into two groups, following the same final irrigation protocols as described above (room-temperature or cold NaOCl, both delivered over 5 minutes with the EndoVac system). Capillaries and microcannulas were maintained at the respective temperatures prior to use. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e-D) Capillaries were dried in a hot-air oven at 50\u0026deg;C for 15 minutes and weighed on a precision scale (Shimadzu, Barueri, Brazil) before and after irrigation. Tissue dissolution was calculated by subtracting final weight from initial weight.\u003c/p\u003e\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\u003ch2\u003eStatistical Analysis\u003c/h2\u003e\u003cdiv id=\"Sec6\" class=\"Section3\"\u003e\u003ch2\u003eRoot Surface Temperature:\u003c/h2\u003e\u003cp\u003eData were tested for normality using the Kolmogorov\u0026ndash;Smirnov test. Paired \u003cem\u003et\u003c/em\u003e-tests were used to compare initial and minimum temperatures between groups.\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e\n\u003ch3\u003eTissue Dissolution\u003c/h3\u003e\n\u003cp\u003eData were first tested for normality using the Kolmogorov\u0026ndash;Smirnov and Shapiro\u0026ndash;Wilk tests. Since final weight values were not normally distributed, the nonparametric Mann\u0026ndash;Whitney \u003cem\u003eU\u003c/em\u003e test was applied. After confirming the normal distribution of \u0026ldquo;Initial weight\u0026rdquo; and \u0026ldquo;weight loss\u0026rdquo; variables, they were compared using independent samples \u003cem\u003eT\u003c/em\u003e Student test.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eRegarding root surface temperature, the control group exhibited an average decrease of nearly 1\u0026deg;C after irrigation with 2.5% NaOCl at room temperature (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). In contrast, the experimental group, which received NaOCl at 2.5\u0026deg;C, showed a mean temperature reduction of approximately 8\u0026deg;C (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). The temperature drop observed in the experimental group was significantly greater than that in the control group (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eProtocol, mean and standard deviation (\u0026deg;C) of the initial temperature and the lowest temperature reached during the procedure.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtocol\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eTime point\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003en\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eMedia (\u0026deg;C)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eDP\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eControl\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eInitial\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e24.94\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1,31\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eLowest\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e24.04\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1,28\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eExperimental\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eInitial\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e24.81\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1,02\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eLowest\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e16.52\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e2,36\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eFor tissue dissolution, no statistically significant difference was observed between groups, regardless of solution temperature (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05) (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Both protocols demonstrated similar ability to dissolve organic matter.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003e\u0026ndash; Protocol, mean and standard deviation of the weight (mg) before and after the irrigation protocols, and the percentage of dissolution of the analyzed groups.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtocol\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eWeight\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eMedia\u0026thinsp;\u0026plusmn;\u0026thinsp;DS (mg)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003e% Dissolution\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eControl\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eInitial\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e50.6\u0026thinsp;\u0026plusmn;\u0026thinsp;6.48\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e1.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.79\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eFinal\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e49.7\u0026thinsp;\u0026plusmn;\u0026thinsp;6.49\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eExperimental\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eInitial\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e46.9\u0026thinsp;\u0026plusmn;\u0026thinsp;2.81\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e1.71\u0026thinsp;\u0026plusmn;\u0026thinsp;0.69\u003csup\u003eA\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eFinal\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e46.1\u0026thinsp;\u0026plusmn;\u0026thinsp;2.85\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eCryotherapy aims to elicit physiological responses such as reduced blood flow, neural receptor inhibition, and decreased metabolic activity [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Its adaptation to intracanal application has emerged as a cost-effective, localized therapeutic strategy for managing inflammation and postoperative pain following endodontic treatment [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIntracanal cryotherapy has been indicated to reduce postoperative pain in patients with symptomatic irreversible pulpitis [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e] or to prevent pain in asymptomatic individuals undergoing root canal treatment [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. In both cases, cryotherapy has been shown to reduce the incidence and severity of postoperative pain, particularly during the first few hours after treatment [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe most widely recommended cryotherapy protocol in the literature involves final irrigation with 20 mL of cold saline (2\u0026deg;\u0026ndash;4\u0026deg;C) for 5 minutes [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. In the present study, cold NaOCl was used instead of saline due to its widespread clinical use and well-established properties, including its antimicrobial activity, tissue-dissolving capacity, and ability to penetrate dentinal tubules [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan additionalcitationids=\"CR19 CR20 CR21\" citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Negative pressure irrigation (EndoVac) was employed to ensure consistent delivery to the working length while minimizing the risk of irrigant extrusion. Using cold NaOCl may simplify clinical protocols by eliminating the need to switch to a different irrigant for cryotherapy.\u003c/p\u003e\u003cp\u003eIn 2015, Vera et al. reported that cryotherapy with cold saline reduced root surface temperatures by up to 14.3\u0026deg;C over 4 minutes [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. In contrast, the present study observed a maximum reduction of 8.3\u0026deg;C with cold NaOCl. This difference may be explained by a higher ambient temperature in the current study (25\u0026deg;C vs. ~23\u0026deg;C), as well as the potential exothermic nature of NaOCl\u0026rsquo;s tissue-dissolving reaction. NaOCl degrades fatty acids through saponification, producing soap and glycerol, which may release heat and slightly limit the cooling effect during irrigation [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eA 2024 study by de Oliveira et al. evaluated tissue dissolution in simulated complex anatomies and found that ultrasonic activation enhanced NaOCl's ability to dissolve organic matter [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. This might be explained by the increased efficiency of heated NaOCl, resulting from an accelerated reaction rate and improved irrigant flow due to ultrasonic agitation [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. In contrast, the present study showed no significant difference in tissue dissolution between cold and room-temperature NaOCl. This discrepancy may be attributed to the much greater amount of organic material used by Oliveira et al. (\u0026gt;\u0026thinsp;700 mg per specimen) compared to the amount used in the present study (\u0026lt;\u0026thinsp;51 mg per specimen).\u003c/p\u003e\u003cp\u003eTo our knowledge, no previous studies have evaluated the tissue-dissolving ability of NaOCl at low temperatures in simulated complex anatomies. Within the limitations of this in vitro study, our findings suggest that negative pressure irrigation with cold NaOCl can reduce external root surface temperature by over 8\u0026deg;C and maintain this reduction for a sufficient duration to potentially elicit periapical physiological effects. Furthermore, low-temperature NaOCl demonstrated a comparable tissue-dissolving capacity to that of room-temperature NaOCl.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIntracanal irrigation with 2.5% sodium hypochlorite at 2.5\u0026deg;C significantly reduced external root surface temperature without compromising its ability to dissolve organic matter, when compared with room-temperature NaOCl. This suggests that cold NaOCl may serve as an effective alternative for intracanal cryotherapy.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConceptualization: M.I.N, E.C.\u003c/p\u003e\n\u003cp\u003eMethodology: M.I.N., P.G.T., U.X.S.\u003c/p\u003e\n\u003cp\u003eData collection and curation: M.H.D., M.I.N., P.G.T.\u003c/p\u003e\n\u003cp\u003eAnalysis and interpretation: U.X.S., V.P.W., B.C.C., M.H.D.\u003c/p\u003e\n\u003cp\u003eVisualization: U.X.S., V.P.W., E.C.\u003c/p\u003e\n\u003cp\u003eWriting – Original Draft Preparation: M.I.N., E.C.\u003c/p\u003e\n\u003cp\u003eWriting – Reviewing and editing: E.C., B.C.C., M.H.D., V.P.W., U.X.S.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSupervision: V.P.W., U.X.S., E.C.\u003c/p\u003e\n\u003cp\u003eAll authors reviewed and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by the “Conselho Nacional de Desenvolvimento Científico e Tecnológico” (CNPq, Brasilia, Brazil) as part of a doctoral research incentive grant. (Number:1610162020-0)\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe dataset generated and analyzed during the current study, including the table of extracted articles, are available from the corresponding author upon request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;This study was approved by the local ethics committee (protocol: 5.024.031).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting Interests\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare they have no competing interest related to this study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eMancini M, Cerroni L, Iorio L, et al. (2013) Smear layer removal and canal cleanliness using different irrigation systems: SEM evaluation. J Endod 39(11):1456\u0026ndash;1460.\u003c/li\u003e\n \u003cli\u003ede Oliveira Neto RS, de Souza Lima LA, Titato PCG, et al.\u0026nbsp;(2023) Effectiveness of a new endodontic irrigation system for removing smear layer and dissolving simulated organic matter. Clin Oral Investig 28(1):1\u0026ndash;6.\u003c/li\u003e\n \u003cli\u003eMoorer WR, Wesselink PR (1982) Factors promoting the tissue dissolving capability of sodium hypochlorite. Int Endod J 15(4):187\u0026ndash;96.\u003c/li\u003e\n \u003cli\u003eStojicic S, Zivkovic S, Qian W, et al. (2010) Tissue dissolution by sodium hypochlorite: effect of concentration, temperature, agitation, and surfactant. J Endod 36(9):1558\u0026ndash;1562.\u003c/li\u003e\n \u003cli\u003eZehnder M. Root canal irrigants (2006) J Endod 32(5):389\u0026ndash;398.\u003c/li\u003e\n \u003cli\u003eHaapasalo M, Shen Y, Wang Z, Gao Y (2014) Irrigation in endodontics.\u0026nbsp;Br Dent J 216(6):299\u0026ndash;303.\u003c/li\u003e\n \u003cli\u003eVera J, Ochoa J, Vazquez-Carca\u0026ntilde;o M, et al.\u0026nbsp;(2015) Effect of intracanal cryotherapy on reducing root surface temperature. J Endod 41(11):1884\u0026ndash;1887.\u003c/li\u003e\n \u003cli\u003eFayyad DM, Abdelsalam N, Hashem N (2020) Cryotherapy: a new paradigm of treatment in endodontics. J Endod 46(7):936\u0026ndash;942.\u003c/li\u003e\n \u003cli\u003eGundogdu EC, Arslan H (2018) Effects of various cryotherapy applications on postoperative pain. J Endod 44(3):349\u0026ndash;354.\u003c/li\u003e\n \u003cli\u003eSadaf D, Ahmad MZ, Onakpoya IJ (2020) Effectiveness of intracanal cryotherapy: systematic review and meta-analysis. J Endod 46(12):1811\u0026ndash;1823.\u003c/li\u003e\n \u003cli\u003eKeskin C, \u0026Ouml;zdemir \u0026Ouml;, Uzun İ, G\u0026uuml;ler B (2017) Cryotherapy after root canal treatment. Aust Endod J 43(2):83\u0026ndash;88.\u003c/li\u003e\n \u003cli\u003eHespanhol FG, Guimar\u0026atilde;es LS, Antunes LAA, Antunes LS (2022) Intracanal cryotherapy and postoperative pain: meta-analysis. Restor Dent Endod 47(3):1\u0026ndash;15.\u003c/li\u003e\n \u003cli\u003eAl-Nahlawi T, Hatab TA, Alrazak MA, Al-Abdullah A (2016) Cryotherapy and negative irrigation technique on postendodontic pain. J Contemp Dent Pract 17(12):990\u0026ndash;996.\u003c/li\u003e\n \u003cli\u003eIparraguirre Nu\u0026ntilde;overo MF, Hungaro Duarte MA, et al.\u0026nbsp;(2024) Intracanal cryotherapy with or without foraminal enlargement. Sci Rep 14(1):19905.\u003c/li\u003e\n \u003cli\u003eNandakumar M, Nasim I (2020) Cryotreated NaOCl and postoperative pain: RCT. J Conserv Dent Endod 23(2):131\u0026ndash;136.\u003c/li\u003e\n \u003cli\u003eLalfakawmi S, Gupta A, Duraisamy AK, et al. (2024) Impact of Cryotreated and Warm Sodium Hypochlorite on Postoperative Pain in Teeth with Symptomatic Irreversible Pulpitis: A Randomized Controlled. J Endod. 50(11):1543\u0026ndash;1550.\u003c/li\u003e\n \u003cli\u003ede Hemptinne F, Slaus G, Vandendael M, Jacquet W, De Moor R J, Bottenberg P (2015) In vivo intracanal temperature evolution during endodontic treatment after the injection of room temperature or preheated sodium hypochlorite.\u0026nbsp;J Endod 41(7):1112-1115.\u003c/li\u003e\n \u003cli\u003eMokhtari H, Milani AS, Zand V, et al. (2023) Temperature of NaOCl and postoperative pain. Clin Exp Dent Res 9(5):859\u0026ndash;867.\u003c/li\u003e\n \u003cli\u003eVera J, Ochoa J, Romero M, et al. (2018) Cryotherapy in symptomatic apical periodontitis: multicenter RCT.\u0026nbsp;J Endod 44(1):4\u0026ndash;8.\u003c/li\u003e\n \u003cli\u003eKarataş E, Ayaz N, Uluk\u0026ouml;yl\u0026uuml; E, et al.\u0026nbsp;(2021) Effect of NaOCl temperature on postoperative pain. J Appl Oral Sci 29:1\u0026ndash;8.\u003c/li\u003e\n \u003cli\u003eClegg MS, Vertucci FJ, Walker C, et al. (2006) Irrigant effects on apical biofilms. J Endod 32(5):434\u0026ndash;437.\u003c/li\u003e\n \u003cli\u003eWong DT, Cheung GS (2014) NaOCl penetration into dentinal tubules. J Endod 40(6):825\u0026ndash;829.\u003c/li\u003e\n \u003cli\u003eEstrela C, Estrela CR, Barbin EL, et al.\u0026nbsp;(2002) Mechanism of action of sodium hypochlorite. Br Dent J 13:113\u0026ndash;117.\u003c/li\u003e\n \u003cli\u003eSun M, Liu S, Zhang Y, et al. (2019) Thermodynamics of saponification reactions. J Mol Liq 280:252\u0026ndash;258.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Endodontics, Cryotherapy, Sodium hypochlorite, Tissue dissolution, Temperature","lastPublishedDoi":"10.21203/rs.3.rs-7811504/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7811504/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cb\u003eIntroduction:\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThis study aimed to evaluate the effect of low-temperature intracanal sodium hypochlorite (NaOCl) irrigation on root surface temperature reduction and its ability to dissolve organic matter.\u003c/p\u003e\u003cp\u003e\u003cb\u003eMethods\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTwenty-five mandibular premolars were accessed and instrumented to size 40.05. Final irrigation protocols with 20 mL of 2.5% NaOCl, at two different temperatures: room temperature (control) and 2.5\u0026deg;C (experimental), were applied. Initial and minimum root surface temperatures (last 4 mm of the root) were recorded using a digital thermometer. For tissue dissolution analysis, glass capillaries filled with catgut were attached to the cervical and apical thirds of twenty 3D-printed maxillary incisors and weighed before and after the same irrigation protocols. Data was statistically analyzed.\u003c/p\u003e\u003cp\u003e\u003cb\u003eResults\u003c/b\u003e\u003c/p\u003e\u003cp\u003eLow-temperature NaOCl irrigation led to a significantly greater reduction in root surface temperature (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Both room-temperature and low-temperature NaOCl showed similar organic matter dissolution capabilities, with no significant difference between them (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e\u003cp\u003e\u003cb\u003eConclusion\u003c/b\u003e\u003c/p\u003e\u003cp\u003eLow-temperature NaOCl effectively reduced root surface temperature while maintaining its tissue-dissolving capacity compared to the control group.\u003c/p\u003e\u003cp\u003e\u003cb\u003eClinical relevance:\u003c/b\u003e\u003c/p\u003e\u003cp\u003eCold sodium hypochlorite effectively reduces external root surface temperature while maintaining its ability to dissolve organic tissue, providing clinicians with a practical and efficient intracanal cryotherapy strategy for minimizing postoperative inflammation without compromising chemo-mechanical preparation.\u003c/p\u003e","manuscriptTitle":"Effect of Low-Temperature Intracanal Sodium Hypochlorite on Root Surface Temperature Reduction and Organic Matter Dissolution: An In Vitro Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-12-04 08:47:09","doi":"10.21203/rs.3.rs-7811504/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-12-07T17:35:19+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"296249973320781908528592736711956200248","date":"2025-12-06T13:00:11+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"97466685883226840821543759878897328740","date":"2025-12-03T16:14:53+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-12-03T13:08:46+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-12-02T05:42:44+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"262600035883359591707837346677478288847","date":"2025-12-01T17:04:03+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"24384927756081952485620356784161796106","date":"2025-12-01T16:10:50+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"141383058663111041736033781073009581797","date":"2025-12-01T12:30:28+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"224075526758739738258023570959052184839","date":"2025-12-01T12:30:22+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-12-01T12:18:13+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-10-10T14:07:58+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-10-09T12:15:04+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-10-09T12:14:35+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-10-08T23:52:33+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"22e53af6-8e5e-4256-9fe0-9376d264234c","owner":[],"postedDate":"December 4th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":59042669,"name":"Health sciences/Health care"},{"id":59042670,"name":"Health sciences/Medical research"}],"tags":[],"updatedAt":"2026-02-02T16:19:39+00:00","versionOfRecord":{"articleIdentity":"rs-7811504","link":"https://doi.org/10.1038/s41598-026-37704-7","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2026-01-31 15:58:16","publishedOnDateReadable":"January 31st, 2026"},"versionCreatedAt":"2025-12-04 08:47:09","video":"","vorDoi":"10.1038/s41598-026-37704-7","vorDoiUrl":"https://doi.org/10.1038/s41598-026-37704-7","workflowStages":[]},"version":"v1","identity":"rs-7811504","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7811504","identity":"rs-7811504","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}