Assessment of the Antibacterial Effect and Post-operative Pain Following Various Techniques of Intracanal Heating of Sodium Hypochlorite: A Randomized Clinical Trial
Phase: PHASE1
Conditions: Necrotic Teeth
Sponsor: Misr University for Science and Technology

 
 
 
Assessment of the Antibacterial Effect and Post-operative Pain 
Following Various Techniques of Intracanal Heating of Sodium 
Hypochlorite: A Randomized Clinical Trial 
ﺗﻘﯿﯿﻢ اﻟﺘﺄﺛﯿﺮ اﻟﻤﻀﺎد ﻟﻠﺒﻜﺘﯿﺮﯾﺎ وأﻟﻢ ﻣﺎ ﺑﻌﺪ اﻟﺠﺮاﺣﺔ ﻟﻠﺘﻘﻨﯿﺎت اﻟﻤﺨﺘﻠﻔﺔ ﻟﺘﺴﺨﯿﻦ 
ھﯿﺒﻮﻛﻠﻮرﯾﺖ اﻟﺼﻮدﯾﻮم داﺧﻞ ﻗﻨﺎة اﻟﺠﺬر: ﺗﺠﺮﺑﺔ ﺳﺮﯾﺮﯾﺔ ﻋﺸﻮاﺋﯿﺔ 
 
A Thesis Protocol  
Submitted for Partial Fulfillment of the Requirements of the 
  Master Degree in Dental Science  
in Endodontics 
Misr University for Science and Technology, College of Oral and Dental Surgery 
Submitted By 
Alaa Eldin Yasser Mohamed Salah Eldin 
Teaching assistant, Endodontics division, Conservative Surgery Department, 
College of Oral and Dental Surgery, Misr University for Science and Technology 
Bachelor’s Degree of Dentistry  (2019) 
Main Supervisor: 
Prof. Dr. Mahmoud Hassan Mohamed 
Professor of Endodontics, College of Oral and Dental Surgery, Misr University for 
Science and Technology and Al-Azhar University 
CO-supervisors: 
Dr. Rehab Ali Farag  
Lecturer of Endodontics, College of Oral and Dental Surgery, Misr University for 
Science and Technology 

 
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Dr. Shaimaa Mohamed Seleim  
Lecturer of Micribiology, Microbiology and Immunology Department, Faculty of 
Medicine, Cairo University 
 
 
Date:  November, 5, 2025

 
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1
 
1- Research Question: 
Is the antibacterial effect of sodium hypochlorite irrigation and the postoperative 
pain affected by heating using either Ultrasonic or System-B activation or a 
combination of both? 
 
 

 
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2- Rationale for conducting the study: 
Intracanal heating of sodium hypochlorite has been suggested as a method of 
activation of root canal irrigation in in-vitro studies. This clinical trial aims to 
evaluate the antibacterial effect of sodium hypochlorite heated inside the root canal 
using various activation techniques. 
 
 

 
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3- Introduction & background:   
 
Root canal treatment success depends on removal of necrotic and vital pulp 
tissues and bacteria and bacterial byproducts from root canal system (Dioguardi 
et al., 2018). 
Mechanical cleansing and removal of necrotic or vital pulp tissue lead to 
formation of a layer of debris called “ Smear layer” which is made of organic and 
inorganic substances that should be removed from root canal system with the 
help of irrigants (Vaz Braga Pintor et al., 2016). 
The presence of complex anatomy like isthmi and anastomoses makes chemical 
cleansing of root canal system difficult because they are filled with the smear 
layer (Torabinejad et al., 2002). 
The ideal features of root canal irrigants include cleansing, lubrication of 
endodontic instruments and root canal system, dissolution of inorganic and 
organic tissues, antimicrobial properties, absence of cytotoxicity without 
alteration of dental microstructure. Sodium hypochlorite (NaOCl) has been the 
most used irrigant in endodontics due to its antimicrobial and tissue dissolving 
action (Zehnder, 2006). 
Due to presence of complex anatomy inside root canal system, activation of 
irrigation is essential to improve cleaning such complex system. Ultrasonic 
activation is presently the most popular method for activation of irrigation. It 
delivers the irrigant farther into areas of the root canal system untouched by 
instruments and it improves the mechanical cleaning by increasing the wall shear 
stress (Boutsioukis and Arias-Moliz, 2022). 
 

 
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NaOCl is used in concentrations between 0.5% - 6%. In high concentrations, it 
may have a faster dissolution capacity, but it is associated with toxicity in cases 
of extrusion, thus, methods to improve the action of less concentrated solutions 
become relevant, such as agitation of the irrigant, constant refreshment and 
volume, and heating of the solution. Using NaOCl solution in high temperature is 
related to an increase in reactivity and, therefore, a reduction in bacterial counts, 
and bacterial biofilm dissolution. Although the dentin acts as a thermal insulator, 
the excessive increase in temperature could dissipate to the periapical tissues, 
causing damage to the bone tissue (Borges et al., 2023). 
Initially, it was suggested to heat NaOCl before inserting it into the canals, but 
this was of little effectiveness (De Hemptinne et al., 2015). 
Woodmansey demonstrated that NaOCl solution can dissolve pulp tissue 210 
times faster at boiling temperature (90-120°C) than at room temperature. 
However, this method remained limited due to the lack of studies confirming its 
safety until Simeone et al. confirmed that when NaOCl solution is heated inside 
the canals at a temperature of 150°C for 10 seconds, the temperature of the 
periodontal tissues does not rise above 42.5°C, which is within the acceptable 
limits for the periodontal tissue (Ahmad Ali et al., 2023) (Woodmansey, 2005) 
(Simeone et al., 2015). 
Rathore et al. in 2020 has stated that intracanal heating to 180°C using System-B 
Heat Source shows better antibacterial efficacy than preheated sodium 
hypochlorite at 60°C. Also it has been suggested to use low concentration NaOCl 
at elevated temperature to report the evidence for better antimicrobial and tissue 
dissolution properties (Govindaraju et al., 2024) (Rathore et al., 2020). 
 

 
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Combination of these two activation methods by ultrasonic agitation of sodium 
hypochlorite followed by intracanal heating may have promising results with 
regards to removal of bacteria (Yared and Al Asmar Ramli, 2020). 
Patients frequently encounter postoperative pain after root canal treatment and 
one of the causes is that NaOCl at higher concentration is irritant to surrounding 
tissue particularly in case of extrusion. Irrigation using lower concentration 
NaOCl was effective in reducing postoperative pain (Rahim et al., 2024). 
Based on the previous studies, studying the effect of irrigation using low 
concentration NaOCl (2.6%) which can be activated to increase its reactivity 
could give predectible antibacterial effect while reducing postoperative pain and 
toxicity associated with extrusion. 
Aim of the study: is to evaluate the antibacterial effect of sodium hypochlorite 
heated inside root canals using various activation techniques. 
 
 
 
 
 
 
 

 
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4- PICO Elements: 
 
Participants:  Patients with asymptomatic single rooted restorable necrotic teeth. 
Intervention 1: Intracanal heating for activation of sodium hypochlorite using  
System-B. 
Intervention 2: Combination of Ultrasonic and System-B for activation of sodium 
hypochlorite. 
Comparator:  Ultrasonic for activation of sodium hypochlorite. 
Outcome: primary: Antibacterial effect 
       
        secondary: Postoperative pain 
Outcomes:  
Prioritization 
of 
Outcome 
Outcome  
Measuring device 
Measuring unit 
Primary outcome 
Antibacterial 
effect 
Colony counting 
CFU/ml 
Secondary 
outcome 
Postoperative pain  Visual Analog 
Scale 
Scoring Scale 
(0-10) 
 
Primary Outcome 
Antibacterial effect is evaluated by counting number of bacterial colonies, samples are 
collected and placed in brain–heart infusion (BHI) broth and subjected to 
microbiological analysis to determine the colony-forming unit (CFU)/ml (Shroff et al., 
2024). 
 
 
 

 
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Secondary Outcome 
Postoperative pain is assessed for each patient using Visual Analog Scale (VAS) with a 
score ranging from 0 (no pain) to 10 (unbearable pain) (Rahim et al., 2024). 

 
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5- Time Frame: 
3 days for postoperative pain follow-up. 
 
 

 
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6- Study design & Methods: 
 
The study design is a parallel randomized controlled clinical trial with allocation 
ratio 1:1:1 
This study is conducted at  Soad Kafafi University Hospital, Clinics of Faculty of 
Dentistry 
 
Sample size: 36 patients divided into 3 groups (12 patients/ group) 
 
Participants’ inclusion criteria:  
For patients: adult male and female patients and medically free. 
For selected teeth: asymptomatic necrotic single rooted teeth, teeth with mature 
root and restorable teeth. 
 
Exclusion criteria:  
For patients: Patients with dental related symptoms ,patients with chronic or 
systemic diseases and pregnant females. 
For selected teeth: multirooted teeth, vital teeth, teeth with incompletely formed 
roots, teeth with internal or external resorption and previously root canal treated 
teeth  
 
 
Intervention Assignment: 
Randomization: 
Randomization will be done according to a check list including the number of 
participants divided into 3 groups according to interventions/Control assessment 
methods. 
The allocation sequence will be generated using (www.random.org). 

 
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Allocation concealment mechanism: 
Allocation concealment is done using sequentially numbered opaque sealed 
envelopes method. 
 
Blinding:  
The participants will be blinded to interventions/control assessment methods. 
Assign numerical codes to samples, ensuring that the experimenters do not know 
which treatment corresponds to which code until the study is completed. 
Materials and methods:  
Preoperative radiograph will be taken for each targeted tooth and vitality test will 
be performed, informed consent will be obtained from patients, preoperative pain 
score will be recorded using VAS, local anasthesia will be administered, rubber 
dam isolation and local disinfection measures applied, then access cavity 
preparation will be performed. Small K #15 file will be used to check canal 
patency, then working length will be determined using electronic apex locater 
and confirmed by a radiograph. 
 
The canals will be enlarged to size 25 K-type file with no irrigation. Then each 
canal will be filled with sterile distilled water using a sterile plastic syringe to 
obtain the first bacteriological sample (S1) before any irrigation. Then a 3 sterile 
paper points size 25 will be placed in the canal for 1 minute. The paper points 
will be immediately transferred into brain–heart infusion (BHI) broth and 
subjected to microbiological analysis. 
 

 
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Patients are randomly divided using computer generated randomization 
(www.random.org) into three groups (n=12/group) according to the irrigant 
activation method : 
Group A (Control group): Activation is done using ultrasonic. 
Group B : Activation is done using System-B. 
Group C: Activation is done using ultrasonic followed by intracanal heating 
using System-B. 
 
The assistant supervisor generates the random sequence and assignes the 
participants to the interventions or control groups. 
Then mechanical instrumentation is performed and canals are irrigated with 2.6% 
NaOCl after each file and the final irrigation protocol is done with activation 
using either ultrasonic or System-B or combination of ultrasonic and System-B 
according to the assigned group. 
 
The ultrasonic activation will be done using ultrasonic tip that reaches 2 mm 
shorter than the working length without touching dentin walls then moved 2-3 
mm in coronal-apical direction for 30 seconds then cycle is repeated 3 times with 
NaOCl refreshment between cycles. 
 
The intracanal heating will be achieved using System-B with temperature set at 
180°C and a suitable tip that reaches 3 mm shorter than working length while not 
touching dentin walls, irrigation is done using 6 ml NaOCl delivered in a syringe 
with side vented needle that reaches 2 mm shorter than working length and 
System-B tip is inserted and activated for 8 seconds then left nonactivated for 10 
seconds, this activation is repeated 10 times and sodium hypochlorite solution is 
refreshed with new solution at each cycle. 
 

 
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Combination of these two activation techniques can be achieved by ultrasonic 
activation followed by intracanal heating using System-B in same manner as 
mentioned in each activation method. 
 
Then the second beacteriologic sample (S2) will be obtained in same manner as 
S1. Then bacteriologic samples will be transferred to microbiology laboratory to 
detect change in bacterial count {Log (CFU/ml)} 
(Mukundan et al., 2024),(Nashaat, 2020),(Iandolo et al., 2018),(Ahmad Ali et al., 
2023) (Shroff et al., 2024) (Yared and Al Asmar Ramli, 2020). 
 
Pain score is recorded for each patient in all groups using Visual Analog Scale 
(VAS) at 6h, 12h, 24h, 48h and 72h postoperative. (Rahim et al., 2024) 
 
 

 
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7-  Statistical plan 
 
Sample size calculation based on bacterial count 
According to a previous study by Mukundan et al (2024) The mean of The mean colony 
count post irrigation for 3 % NaOCl was 258.05 ± 28.61, and for 1 % NaOCl it was 
267.60 ± 30.56.  Using G power statistical power Analysis  program (version 3.1.9.4) for 
sample size determination (Uttley, 2019), A total sample size (n=36; subdivided to 12 in 
each group) will be sufficient to detect a large effect size  (f) = 0.54, with an actual  power 
(1-β error) of 0.8 (80%) and a significance level (α error) 0.05 (5%) for two-sided 
hypothesis test. 
 
Statistical analysis  
Statistical analysis will be performed using a commercially available software program 
(SPSS Chicago, IL, USA).   
Numerical data will be described as mean and standard deviation or as median and range 
as appropriate according to the normality of the data. Data will be compared using 
Kruskal Wallis test or ANOVA test depending on normality. The level of significance 
will be set at P≤0.05. All tests will be two tailed. 
 
 
 
 

 
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8- Ethics consideration:  
 
The present research will be conducted after the approval of the Research Ethics 
Committee (REC) of the Faculty of Dentistry & Ethics Committee of Misr University 
For Science and Technology. It will be conducted on healthy patients with asymptomatic 
single rooted restorable necrotic teeth. 
Ethical considerations regarding patients well-being and confidentiality will be 
undertaken by the researcher and an informed  written consent will be signed by the 
patients before commencing the study explaining all clinical examinations , procedures 
and follow up. ( Attached appendix ) 
 

 
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9- References:  
 
Ahmad Ali, I., Layous, K., Alzoubi, H., 2023. Evaluating the Effectiveness of Different 
Irrigant Activation Techniques in Removing the Smear Layer and Opening the 
Dentinal Canals: A Scanning Electron Microscopic Study. Cureus 15, 1–10. 
https://doi.org/10.7759/cureus.33961 
Borges, M.M.B., de Barros, M.C., de Azevedo Queiroz, Í.O., de Andrade, F.B., Duarte, 
M.A.H., 2023. Effect of two methods of irrigant agitation on the temperature and 
cleanliness of sodium hypochlorite associated or not with a chelator. Brazilian J. 
Oral Sci. 22. https://doi.org/10.20396/bjos.v22i00.8668692 
Boutsioukis, C., Arias-Moliz, M.T., 2022. Present status and future directions – 
irrigants and irrigation methods. Int. Endod. J. https://doi.org/10.1111/iej.13739 
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, 1112–1115. https://doi.org/10.1016/j.joen.2015.02.011 
Dioguardi, M., Di Gioia, G., Illuzzi, G., Laneve, E., Cocco, A., Troiano, G., 2018. 
Endodontic irrigants: Different methods to improve efficacy and related problems. 
Eur. J. Dent. https://doi.org/10.4103/ejd.ejd_56_18 
Govindaraju, L., Shruthi, S.T., Gopal, R., Jenarthanan, S., Rajendran, M.R., 2024. Does 
increase in temperature of sodium hypochlorite have enhanced antimicrobial 
efficacy and tissue dissolution property?-A systematic review and meta-regression. 
J. Conserv. Dent. Endod. 27, 675–684. 
https://doi.org/10.4103/JCDE.JCDE_110_24 
 

 
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Iandolo, A., Amato, M., Dagna, A., Poggio, C., Abdellatif, D., Franco, V., Pantaleo, G., 
2018. Intracanal heating of sodium hypochlorite: Scanning electron microscope 
evaluation of root canal walls. J. Conserv. Dent. 21, 569. 
https://doi.org/10.4103/JCD.JCD_245_18 
Mukundan, D., Jeevanandan, G., Vishwanathaiah, S., Panda, S., Dawood, T., Abutaleb, 
A., Maganur, P.C., 2024. Comparative evaluation of the efficacy of 1% and 3% 
Sodium hypochlorite in reducing the microbial counts in primary teeth root canals 
using Bioluminometer – A randomized clinical trial. Saudi Dent. J. 
https://doi.org/10.1016/j.sdentj.2024.06.006 
Nashaat,  yousra, 2020. Evaluation of the Antibacterial efficacy of newly formulated 
Nano Triple Antibiotic paste with Nano Anti-inflammatory drug as a root canal 
medicament. (A double blind randomized clinical trial). Egypt. Dent. J. 66, 2815–
2824. https://doi.org/10.21608/edj.2020.41856.1245 
Rahim, E., Memon, M., Memon, P., Parveen, S., Qureshi, A.M.N., Shaikh, A.G., 2024. 
Comparison of the Postoperative Pain Following Endodontic Irrigation Using 1.3% 
Versus 5.25% Sodium Hypochlorite in Mandibular Molars with Necrotic Pulps. 
Pakistan J. Heal. Sci. 169–174. https://doi.org/10.54393/pjhs.v5i04.1403 
Rathore, V., Samel, D., Moogi, P., Bandekar, S., Kshirsagar, S., Vyas, C., 2020. 
Antimicrobial efficacy of intracanal and extracanal heated sodium hypochlorite 
against Enterococcus faecalis: An in vitro study. Endodontology 32, 112–117. 
https://doi.org/10.4103/endo.endo_21_20 
Shroff, M., Brave, D., Rathore, V.P.S., Sharma, V., Mehta, J., Thakkar, S.J., 2024. 
Antimicrobial Efficacy of Simvastatin and Double Antibiotic Paste as Intracanal 
Medicaments: A Randomised Clinical Study. Adv. Hum. Biol. 14, 158–162. 
https://doi.org/10.4103/aihb.aihb_141_23 
 

 
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Simeone, M., Valletta, A., Giudice, A., Di Lorenzo, P., Iandolo, A., 2015. The 
activation of irrigation solutions in Endodontics: A perfected technique. G. Ital. 
Endod. 29, 65–69. https://doi.org/10.1016/j.gien.2015.08.005 
Torabinejad, M., Handysides, R., Khademi, A.A., Bakland, L.K., 2002. Clinical 
implications of the smear layer in endodontics: A review. Oral Surg. Oral Med. 
Oral Pathol. Oral Radiol. Endod. 94, 658–666. 
https://doi.org/10.1067/moe.2002.128962 
Uttley, J., 2019. Power Analysis, Sample Size, and Assessment of Statistical 
Assumptions—Improving the Evidential Value of Lighting Research. LEUKOS - J. 
Illum. Eng. Soc. North Am. 15, 143–162. 
https://doi.org/10.1080/15502724.2018.1533851 
Vaz Braga Pintor, A., Rotter Marins dos Santos, M., Masterson Ferreira, D., Barcelos, 
R., Guimarães Primo, L., Cople Maia, L., Professor Rodolpho Paulo Rocco, R., 
2016. Does Smear Layer Removal Influence Root Canal Therapy Outcome? A 
Systematic Review, The Journal of Clinical Pediatric Dentistry. 
Woodmansey, K.F., 2005. Intracanal heating of sodium hypochlorite solution: an 
improved endodontic irrigation technique. Dent. Today 24, 114, 116. 
Yared, G., Al Asmar Ramli, G., 2020. Antibacterial Ability of Sodium Hypochlorite 
Heated in the Canals of Infected Teeth: An Ex Vivo Study. Cureus. 
https://doi.org/10.7759/cureus.6975 
Zehnder, M., 2006. Root Canal Irrigants. J. Endod. 
https://doi.org/10.1016/j.joen.2005.09.014