UNIVERSIDADE ESTADUAL PAULISTA

“JÚLIO DE MESQUITA FILHO”

Instituto de Ciência e Tecnologia

Campus de São José dos Campos

ORIGINAL ARTICLE DOI: https://doi.org/10.4322/bds.2024.e4316

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in

any medium, provided the original work is properly cited.

Assessment of Dual Rinse Combined with Sodium Hypochlorite

Irrigating Solution on Post-Instrumentation Pain and Bacterial Load

Reduction: A Randomized Clinical Trial

Avaliação do Dual Rinse associado à solução irrigadora de hipoclorito de sódio na dor pós-instrumentação e na redução da

carga bacteriana: um ensaio clínico randomizado

Laila Zakaria Ismail ABDEL-HAMID1 , Randa Mohamed EL BOGHDADI1 , Shaimaa Ismail GAWDAT1,2 , Sherifa Tarek SALEM3 ,

Noha Salah SOLIMAN3 

1 - Cairo University, Faculty of Dentistry, Department of Endodontics. Cairo, Egypt.

2 - New Giza University, Faculty of Dentistry, Department of Endodontics. Giza, Egypt.

3 - Cairo University, Faculty of Medicine, Department of Clinical and Chemical Pathology. Cairo, Egypt.

How to cite: Abdel-Hamid LZI, El Boghdadi RM, Gawdat SI, Salem ST, Soliman NS. Assessment of Dual Rinse combined with sodium

hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial. Braz Dent Sci.

2024;27(4):e4316. https://doi.org/10.4322/bds.2024.e4316

ABSTRACT

Objective: The study compared the efcacy of Dual Rinse combined with sodium hypochlorite in comparison to

sodium hypochlorite irrigating solution on post instrumentation pain incidence, reduction of the load of intracanal

bacteria and periapical MMP-9 expression in patients with pulp necrosis. Material and Methods: Thirty-four

patients diagnosed with a necrotic mandibular premolar having a single root canal were included. Following

conrmation of the diagnosis, patients were randomly allocated into one of two groups of 17 patients each

(intervention group; Dual Rinse + 2.5% NaOCl, control group; 2.5% NaOCl irrigation). Standard endodontic

treatment was done in two visits. Post-instrumentation pain was assessed at 6, 12, 24 hours and 48 hours using

Heft-parker visual analogue scale (Heft parker VAS). The intracanal bacterial levels assessed pre- and post-

instrumentation through counting of CFU/mL. Periapical MMP-9 levels were assessed post-instrumentation and

pre-obturation by ELISA. All data collected from patients was statistically analyzed. Results: Regarding post-

instrumentation pain incidence, there was no statistically signicant difference detected among tested irrigants at

all time intervals (p>0.05). There was a signicant reduction in the count of intracanal bacterial in both groups

after instrumentation, but without any signicant difference between tested irrigants (p>0.05). Within a week,

there was a considerable drop in the periapical MMP-9 levels without any discernible differences between the

two irriagnts (p>0.05). Conclusion: The addition of Dual Rinse to 2.5% NaOCl solution did not result in any

apparent increase in the incidence of post-instrumentation pain or in periapical MMP-9 levels. Both irrigating

solutions were almost equally successful in lowering the amount of bacteria in primary infected root canals while

maintaining the antibacterial activity of the NaOCl.

KEYWORDS

Bacterial load reduction; Dual rinse in sodium hypochlorite; Irrigation; Matrix metalloproteinase (MMP-9);

Post-instrumentation pain.

RESUMO

Objetivo: O estudo comparou a ecácia do Dual Rinse associado ao hipoclorito de sódio com a solução irrigadora

de hipoclorito de sódio na incidência de dor pós-instrumentação, na redução da carga bacteriana intracanal e na

expressão periapical de MMP-9 em pacientes com necrose pulpar. Material e Métodos: Trinta e quatro pacientes

diagnosticados com necrose pulpar em pré-molar inferior com um único canal radicular foram incluídos. Após a

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

INTRODUCTION

Root canal procedures are commonly

associated with pain during the rst 24 hours.

The incidence of postoperative discomfort is

reported to be up to 40% and then gradually

declines [1]. Many factors affect the occurrence of

postoperative pain such as: mechanical, chemical

and microbial factors which are responsible for

periradicular irritation [1,2]. The primary objective

of endodontic therapy is to totally eradicate

bacteria along with their byproducts from root

canals. It is considered as a difcult task since the

root canal anatomy is complex, which makes the

entire removal of microorganisms challenging.

All tissue remnants and debris cannot be removed

using only root canal instruments. Thus, to

achieve proper disinfection of the root canals,

irrigants play an important role to overcome

the limitations of instrumentation [3]. Sodium

hypochlorite (NaOCl) is the most commonly

utilized irrigant for root canal disinfection due

to its excellent antibacterial action on gram

positive, gram negative planktonic bacteria and its

ability to effectively remove the bacterial biolm.

In addition to its proteolytic action with organic

tissue dissolving ability [4]. However, the smear

layer’s inorganic component and accumulated

hard tissue debris created during instrumentation

cannot be removed by NaOCl solution alone.

As a result, it is usually recommended to apply

ethylenediaminetetraacetic acid (EDTA) and

NaOCl in an alternating manner [5]. However,

combination of EDTA with NaOCl was found

to eliminate the active chlorine in the NaOCl,

thus impairing its antibacterial action, and the

excessive use of EDTA leads to dentine erosion [6].

Therefore, using EDTA as a nal irrigant prior to

obturation is advised.

Etidronic acid, also known as HEDP or

1-hydroxyethane 1,1-diphosphonic acid is

biocompatible with NaOCl and has a mild

chelating action [7] hence the idea of “continuous

chelation” was introduced [8], which involved

the use of NaOCl in combination with 9%

Etridonic acid. The main clinical advantage

of mixing HEDP and NaOCl during irrigation,

is the application’s ease and time saving over

ushing root canals with two irrigating agents

alternatively. The chemistry of Dual Rinse HEDP

is based on sodium etidronate, the HEDP salt is

added to the NaOCl for clinical use just before

the treatment begins, in order to provide in

order to provide single root canal irrigant that

combines proteolytic and chelating properties.

It is considered to be the rst HEDP material

advocated to be used in root canal treatment.

It comes in the form of capsules with 0.9 grams

of powdered etidronate, which needs to be

combined with 10 mL of NaOCl prior to use. As a

result, an irrigant mixture containing about 9%

HEDP and active chlorine is created [9].

Several in vitro studies [10-16] and scoping

review of laboratory studies [17] have shown

promising findings when HEDP was mixed

with NaOCl. The combination of HEDP with

NaOCl was not found to impair the antibacterial

conrmação do diagnóstico, os pacientes foram alocados aleatoriamente em um dos dois grupos de 17 pacientes

cada (grupo de intervenção; Dual Rinse + NaOCl 2,5%, grupo de controle; irrigação com NaOCl 2,5%). O

tratamento endodôntico padrão foi realizado em duas visitas. A dor pós-instrumentação foi avaliada em 6, 12,

24 horas e 48 horas usando a escala visual analógica de Heft Parker (EVA de Heft Parker). Os níveis bacterianos

intracanais foram avaliados antes e depois da instrumentação por meio da contagem de CFU/mL. Os níveis

periapicais de MMP-9 foram avaliados por ELISA pós-instrumentação e pré-obturação. Todos os dados coletados

dos pacientes foram analisados estatisticamente. Resultados: Com relação à incidência de dor pós-instrumentação,

não foi detectada diferença estatisticamente signicativa entre os irrigantes testados em todos os intervalos de

tempo (p>0,05). Houve uma redução signicativa na contagem de bactérias intracanais em ambos os grupos

após a instrumentação, mas sem nenhuma diferença signicativa entre os irrigantes testados (p>0,05). Em uma

semana, houve uma queda considerável nos níveis periapicais de MMP-9 sem nenhuma diferença perceptível

entre os dois irrigantes (p > 0,05). Conclusão: A adição do Dual Rinse à solução de NaOCl a 2,5% não resultou

em nenhum aumento aparente na incidência de dor pós-instrumentação ou nos níveis periapicais de MMP-9.

Ambas as soluções irrigadoras foram quase igualmente bem-sucedidas na redução da quantidade de bactérias

nos canais radiculares infectados primários, mantendo a atividade antibacteriana do NaOCl.

PALAVRAS-CHAVE

Redução da carga bacteriana, Dual Rinse em hipoclorito de sódio, Irrigação, Matriz metaloproteinase (MMP-9),

Dor pós-instrumentação.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

action of NaOCl in terms of eradication of

Enterococcus faecalis

within biofilms and in

the dentinal tubules [10-12], this antibacterial

efcacy was not diminished even in the presence

of the smear layer [13]. The same combination

of irrigants during canal instrumentation were

associated with signicantly less hard-tissue debris

deposition [14,15]. The use of this irrigant mix as

a nal rinse did not decalcify the canal walls and

was capable of adequate smear layer removal [16].

Chelating agents such as HEDP when

combined with NaOCl have shown to signicantly

reduce debris deposition, but might have potential

undesirable effects as they may excessively erode

dentine in the apical portion of the canal, thus

widening the apical foramen and causing irrigant

extrusion, thereby increasing postoperative

discomfort and/or inammatory changes within

the periapical tissues [18]. Furthermore, it is

possible that HEDP reacts slowly when combined

with NaOCl and limits its antibacterial action [9].

After thorough systematic online search,

articles published on the use of HEDP in combination

with NaOCl are mainly in-vitro studies and only one

clinical trial was found measuring postoperative

pain and bacterial reduction after using HEDP in

combination with NaOCl versus NaOCl alone [19].

Since clinical studies are the gold standard of

interventional trials as they produce the highest

level of evidence, effectively aid in clinical decision-

making regarding the best intervention for the

patient’s condition, and provide the most successful

clinical outcomes for the patient’s satisfaction.

Thus, the aim of this study was to

compare the efcacy of Dual Rinse combined

with sodium hypochlorite in comparison to

sodium hypochlorite irrigating solution on post

instrumentation pain incidence, reduction of

the load of intracanal bacteria and periapical

MMP-9 expression in patients with pulp necrosis.

The null hypothesis presumed that there was no

difference in postinstrumentation pain, bacterial

reduction and periapical MMP-9 expression

after irrigation using Dual Rinse combined with

sodium hypochlorite versus sodium hypochlorite.

MATERIAL & METHODS

Ethical approval and protocol registration

The study proposal was approved by the

Research Ethics Committee of the Faculty of

Dentistry in Cairo university with approval

number (REF No:19-7-61) and registered in

Clinical Trials Registry of the US (ClinicalTrials.

gov NCT04035330). The reporting of this study

was done in accordance with the CONSORT

guidelines.

Trial design and study settings

The study was a prospective randomized

controlled blinded clinical trial, with two

parallel groups and an allocation ratio 1:1.

The research was carried out in compliance with

the 2013 Helsinki Declaration guidelines. Each

patient gave their written informed consent and

were given a copy of it. This study was conducted

in the outpatient endodontic department clinic

inside Faculty of Dentistry, Cairo University,

during the period from May November 2020 to

March 2022 by the same operator.

Sample size calculation

According to results of a previous

investigation [20], the anticipated mean difference

in pain score at 48 hours using Heft parker VAS

scale between the two groups was 20. Using

power 80% and 5% signicance level a total of

15 patients in each group (total 30 patients) in

the two groups were studied. To compensate for

losses during follow-up, the number was raised

once more to a total sample size of 17 per group

(drop-out rate 15%). PS: Power and Sample Size

Calculation Software Version 3.1.2 was used to

calculate the sample size (Vanderbilt University,

Nashville, Tennessee, USA).

Eligibility criteria

ASA I or II healthy male or female patients,

18–50 years old, with necrotic permanent

mandibular premolar teeth were enrolled in this

trial. Clinically the diagnosis was established

according to following criteria; lack of spontaneous

pain and radiographically through widening of

the periodontal membrane space or a periapical

radiolucency within range of 2 mm. Patients

with compromised medical conditions, pregnant

women, history of intolerance to any medication,

and those who received analgesics or antibiotics

during the last 24 hours were excluded. Other

exclusion criteria included teeth associated with

any acute pain and/or swelling, pocket depth

more than 5mm, mobility more than grade I,

previously treated teeth, non-restorable teeth.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

Incompletely formed roots, signs of vertical root

fracture, external or internal root resorption,

calcication or perforation were also excluded.

Clinical diagnosis

All patient information, including their

name, phone number, address, their medical

and dental history along with history of chief

complaint was recorded. Extraoral and intraoral

clinical examination of the suspected tooth

by percussion, palpation, pulp testing and

radiographic examination were precisely

performed. The diagnosis of necrotic mandibular

premolar teeth was confirmed by the chief

complaint’s history of reporting no discomfort

in response to heat or cold, negative response to

the electric pulp testing, where the contralateral

tooth served as a control to verify the previous

diagnostic tests, and radiographic assessment

using intra-oral sensor plate and Digora software

(Soredex, Helsinki, Finland) showed lower

premolar with slight widening in the periodontal

membrane space or a periapical radiolucency

within 2 mm.

Randomization, allocation concealment and

blinding

Participants included thirty-four patients

who met the inclusion criteria. They were

randomly assigned to one of two different

irrigation regimens. Software (http://www.

random.org/) was used to create a computerized

random sequence. For the allocation concealment,

the operator placed eight folded numbered papers

in sealed envelopes and these papers were later

dragged by the patients. The investigator, the

participants, and the assessors were blinded.

The assistant prepared 25 mL of the irrigant in an

amber bottle and gave them to the investigator

who was carrying out the root canal treatment,

such that the irrigants cannot be distinguished

from one another.

Root canal clinical procedures and sampling

First visit

The teeth were anaesthetized by 2%

mepivacaine hydrochloride with epinephrine

1:100,000 (Mepecaine-L; Alexandria Company

for Pharmaceuticals, Alexandria, Egypt) then

isolated using rubber dam (Dental Dam;

Sanctuary Dental, UK). The eld was swabbed

with 30% hydrogen peroxide and 5.25% NaOCl

to thoroughly disinfect all surfaces. Access cavity

was prepared by a sterile round and an Endo Z

bur (Dentsply Maillefer, Ballaigues, Switzerland)

under rubber dam isolation. After completion of

the access, the pulp chamber and the operational

field were cleansed and sanitized once more

in the manner described above. Sodium 5%

thiosulfate was then used to neutralize the

NaOCl. Conrmation of canal patency was carried

out with hand K-les (MANI Inc; Utsunomiya,

Japan) size #10 or #15. Using an electronic

apex locator (Root ZX; J. Morita, Irvine, CA,

USA), the working length was determined then

radiographically veried at 1 mm short of the

radiographic apex. The canals were enlarged to

size #20. Collection of the

pre-instrumentation

bacterial sample S-1

was done by flushing

the canal with saline and a sterile paper point

#15 or #20 (Meta Biomed Co., Ltd, Korea) was

then inserted for at least one minute to absorb

the uid inside the canal within 1 mm from the

radiographic apex. This was done 3-4 times.

Then, paper points were aseptically transferred

to tubes that contain 2 mL of sterile thioglycollate

broth (Merck, Darmstadt, Germany). The patients

were randomly divided into 2 groups as follows;

control group (2.5% NaOCl) and intervention

group (Dual Rinse + 2.5% NaOCl).

Preparation of irrigation

25 mL of 2.5% NaOCl irrigation was prepared

for the control group by mixing 12.5 mL of 5%

NaOCl with 12.5 mL of distilled water. In the

intervention group, preparation of 25 mL of 2.5%

NaOCl in a similar manner as in control group and

to prepare 9% Dual Rinse HEDP directly before

treatment; 10 mL NaOCl was added with each

Dual Rinse HEDP capsule containing 0.9 grams

of the etidronate powder to create a combined

irrigant. A total of 2.5 Dual Rinse HEDP capsules

were used to prepare a total volume of 25 mL of

the allocated irrigant. Mechanical preparation

was done according to the manufacturer’s

recommendations utilizing M Pro rotary files

sizes 18/.09, 20/.04, 25/.06, 35/.04 (Innovative

materials and Devices; Shanghai, China) and

an endodontic X smart motor followed by using

#40 hand K-les for apical gauging. Irrigation

was done with a disposable plastic syringe and

a 30 gauge side vented needle (Steri irrigation

tips; Diadent, Chungcheongbuk-do, Korea) that

reached 1 mm less than the working length,

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

the canals were irrigated thoroughly using the

appropriate irrigating solution at a rate of 5 mL

for 1 min. Equal volume of irrigation was applied

to all root canals (5 mL prior to instrumentation,

5 mL between each subsequent le, and 5 mL as

the last ush after root canal instrumentation,

for a total volume of 25 mL). Sterile paper points

were used to dry the canal and 5 mL of 5% sodium

thiosulfate solution was ushed into the canal,

followed by addition of 5 mL distilled water for

inactivation of NaOCl prior to obtaining the post-

instrumentation bacterial sample (S-2) which was

collected in a similar way as S-1 sample. The rst

periapical sample (post-instrumentation) (PS-1)

was collected after instrumentation, by inserting

sterile #20 paper point for one minute, past the

apical foramen by 2mm and this process was

repeated 3-4 times. Then, the paper points were

put into a sterile 2 mL microcentrifugation tube

that contained sterile saline solution before

being quickly transported to an 80 °C freezer for

further use. The access cavities were sealed by

glass ionomer (Medil, Promedica) without the

use of intracanal medication. The patients were

scheduled for recall after 1 week. Treatment

of all participants was completed in two visits

and the patients recorded their degree of post-

instrumentation pain on Heft parker VAS at 6,12,

24 and 48 hours. Ibuprofen 400 mg (Novartis

Pharma S.A.E., Pathion Inc., Ontario, Canada)

was prescribed in case of severe pain.

Second visit

The canals were re-entered and ushed with

saline under rubber dam isolation. To obtain the

second periapical sample (pre-obturation) (PS-2),

canals were disinfected in a sample collected in a

similar manner to PS-1. Following the sampling,

the nal ushing of the canals was done in the

control and intervention groups with 5 mL of

(2.5% NaOCl- 17% EDTA), (2.5% NaOCl- 9%

Dual Rinse HEDP) respectively. Passive ultrasonic

irrigant activation was done for 60 seconds using

a P5 Satalec ultrasonic device with an Irrisafe tip

#25, zero taper. Gutta-percha master cones with a

#40, 0.04 taper were tted to the working length

which was veried with a radiograph. Obturation

of the canals was done with the modied single

cone technique and a resin-based sealer (ADseal,

Meta Biomed Co. Ltd, Korea) using the 0.04 taper

gutta percha cones (Meta Biomed Co. Ltd, Korea)

with auxillaries. Access cavities were sealed with

composite resin (3M, ESPE, Filtek).

Post-instrumentation pain assessment

The patients recorded their degree of post-

instrumentation pain on Heft parker VAS at 6,12,

24 and 48 hours. Patients were asked to place

a mark anywhere on the horizontal VAS that is

170mm long. The distance between point zero and

the patient’s mark was measured by the operator

using a ruler so that their pain level was assigned

as the following: No pain; = 0mm, mild pain;

>0 mm and ≤ 54 mm, moderate pain; > 54 mm

and <114 mm and severe pain (≥114 mm).

Microbiological analysis

Isolation and identication of aerobic and

anaerobic bacteria): Pre and post treatment

root canal samples in both groups were cultured

for qualitative and quantitative assessment of

microbial infection. For isolation of anaerobic

bacteria, the samples were streaked with a

metronidazole disc (5 μg, Oxoid, Basingstoke,

UK) on neomycin blood agar, blood agar as

well as phenyl ethyl alcohol agar. Anaerobic

incubation of the cultured plates was done in a

chamber at 37 °C for 3 days with an automatic,

microprocessor-controlled system for the

cultivation of anaerobic bacteria. In parallel,

specimens were plated aerobically on blood

agar, chocolate and MacConkey agar (BD, Becton

Dickinson, Heidelberg, Germany) and incubated

aerobically at 37 °C for 48 hours. 10-fold serial

dilutions up to 1/104 were made in thioglycollate

broth in order to quantify the number of bacteria.

The main bacterial species isolated in culture were

identied according to standard microbiological

procedures on the basis of colony morphology,

biochemical properties and Gram stain. The use

of matrix-assisted laser desorption/ionization-

time-of-ight (MALDI-TOF) analysis allowed for

the identication of bacterial morphotypes that

could not be detected using standard techniques.

Analysis of the periapical MMP-9 levels

Quantikine ELISA (Human MMP-9

Immunoassay USA & Canada | R&D Systems,

Inc.Catalog Number SMP900) to quantitatively

determine the concentration of both pro- (92

kDa) and active (82 kDa) human MMP-9.

Sample preparation

The extraction of MMP-9 form the paper

points was done by adding 1 mL PBS then

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

vortexing for 15 seconds, and then centrifugation

for 5 minutes at 3000xg. The supernatant

was used for quantitative measurement of

concentration of human active (82kDa) and Pro-

(92kDa) Matrix Metalloproteinase 9 (MMP-9).

Principle of the assay

It is a quantitative technique that used the

sandwich enzyme immunoassay. A microplate

was precoated with a monoclonal antibody that

was selectively directed against human MMP-

9. Standards and samples were pipetted inside

the wells, followed by binding of immobilised

antibody to MMP-9. Following removal of any

unbound material by washing, the human

MMP-9 polyclonal antibody linked to an enzyme

was added to the wells. The wells were next lled

with a substrate solution, and a colour developed

proportional to the amount of MMP-9 initially

bounded. This was done after a wash to remove

unbound antibody-enzyme reagent. The colour

development was stopped, and the colour

intensity was measured.

Statistical analysis

Categorical data were reported as frequency

and percentage values and Fisher’s exact test

was used for comparisons between the groups.

For comparisons within the same group, pairwise

comparisons using repeated McNemar’s tests

with Bonferroni correction were conducted after

the Cochran q test. Numerical data values were

presented in the form of mean and standard

deviation (SD), they were checked for normality

using Shapiro-Wilk test. VAS data showed non-

parametric distribution so they were analyzed

using Mann-Whitney U test for comparisons

between the groups and Friedman’s test followed

by Nemenyi post hoc test for comparisons within

the same group. Bacterial count data were log

transformed to correct for positive skewness and

with other numerical data they were found to be

normally distributed. For intergroup comparisons,

the independent t-test was employed, and for

intragroup comparisons, repeated ANOVA

measures were conducted followed by the

Bonferroni post hoc test. All tests were conducted

with a signicance level of p<0.05. R statistical

analysis software, windows version 4.1.3 was

used to conduct the statistical analysis [21].

RESULTS

Out of 50 patients meeting the eligibility

criteria, a total of 34 patients met the inclusion

criteria and were enrolled, patients were

randomized into one of two groups (17 patients

each). All patients were included in the analysis

as shown in (Figure 1). No statistically signicant

difference was detected among the tested groups

regarding age, gender, tooth type (rst or second

mandibular premolar) as shown in (Table I).

Figure 1 - CONSORT 2010 Flow diagram of the trial design.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

Post-instrumentation pain

At 6 hours, a higher incidence of post-

instrumentation pain was recorded in the Dual

Rinse + NaOCl group (5.9%) compared to no

pain reported in the NaOCl group. At 12 hours,

both groups showed a similar incidence of post-

instrumentation pain (23.5%), while at 24 hours,

a higher incidence was reported in the NaOCl

group (23.5%) compared to the Dual Rinse +

NaOCl group (17.6%). Finally, at 48 hours, the

incidence of pain was higher in the Dual Rinse

+ NaOCl group (5.9%) compared to no pain

reported in the NaOCl group. The only pain

category reported among participants at any time

intervals was the mild category. There was no

statistically signicant difference detected among

the tested groups at any time interval (p>0.05).

Microbiological analysis

Incidence of bacterial growth

All pre-instrumentation root canal samples

at S-1 showed positive microbial growth in both

groups. Irrigation with Dual Rinse + NaOCl

rendered 47.1% of canals free from bacteria

(8 out of 17 cases), while irrigation with NaOCl

rendered 41.2% of canals free from bacteria (7 out

of 17 cases). There was no signicant difference

between both groups regarding incidence of

aerobic, anaerobic and total bacterial growth at

S-1 and S-2 time periods (p>0.05) (Table II).

Percent % of bacterial load reduction

The mean % reduction for the total bacterial

count was higher in Dual Rinse + NaOCl

group (72.35±40.97) than in the NaOCl group

(68.92±41.31) with no statistically signicant

difference between the two groups (p=0.759)

(Figure 2).

Bacterial species

As determined by MALDI-TOF, the isolated

aerobes were found to belong to four species,

predominated by

Streptococcus mitis

and

Enterococcus faecalis

. The identification of

anaerobic isolates by MALDI-TOF revealed nine

species with the highest frequencies shown for

Fusobacterium, Parvimonas, and Eikenella

There was no significant difference between

both groups in terms of the presence of different

Table I - Mean, standard deviation (SD), frequencies (n), and percentages (%) for comparison of demographic data between the two groups

Parameter NaOCl Dual Rinse + NaOCl p-value

Gender

Male n 6 5

1 ns

% 35.3% 29.4%

Female n 11 12

% 64.7% 70.6%

Age Mean ± SD* 32.94±9.40 35.82±7.02 0.319 ns

Tooth type

Mandibular 1st

Premolar

n 7 6

1 ns

% 41.1% 35.3%

Mandibular 2nd

Premolar

n 10 11

% 58.9% 64.7%

Signiﬁcant (p ≤ 0.05). ns: non-signiﬁcant (p>0.05).

Table II - Frequency and percentage values for incidence of bacterial growth between two groups at S-1 and S-2

Bacterial type Incidence of Bacterial growth at time intervals NaOCl Dual Rinse +NaOCl p-value

Total S-1 n (%) 17 (100.0%) 17 (100.0%) NA

S-2 n (%) 10 (58.8%) 9 (52.9%) 1 ns

Aerobic S-1 n (%) 14 (82.4%) 13 (76.5%) 1 ns

S-2 n (%) 6 (35.3%) 2 (11.8%) 0.225 ns

Anaerobic S-1 n (%) 13 (76.5%) 15 (88.2%) 0.653 ns

S-2 n (%) 4 (23.5%) 7 (41.2%) 0.463 ns

Signiﬁcant (p ≤ 0.05). ns: non-signiﬁcant (p>0.05). N/A: means not applicable

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

types of bacteria at both time intervals and there

was no clear bacterial species selection by either

treatment groups, and similar taxa prevalence

across the two groups (p>0.05) (Table III,

Table IV, Figure 3, Figure 4).

Periapical MMP-9 levels

At the first treatment visit, mean values

of MMP-9 were (217.82±82.94) in the NaOCl

group and (177.87±39.63) in the Dual Rinse +

NaOCl group. These values dropped signicantly

in the second visit reaching mean values

of (92.45±14.66) in the NaOCl group and

(85.88±18.46) in the Dual Rinse + NaOCl group.

There was no statistically signicant difference

in the ability of any of both irrigants to lower

periapical MMP-9 levels (p>0.05).

DISCUSSION

One of the fundamental goals of endodontic

therapy is controlling and reducing pain during

and particularly after root canal treatment.

Table III - Frequency (n) and % values for bacterial species at S1and S2 in NaOCl group

Group Bacterial species S1 S2 p-value

Aerobic

Strept mitis/oralis

8 (47.1%) 2 (11.8%)

0.710ns

Enterococcus faecalis

5 (29.4%) 1 (5.9%)

Staph hominus

5 (29.4%) 3 (17.6%)

Klebsiella oxytoca

1 (5.9%) 0 (0.0%)

Anaerobic

Fusobacterium spp

5 (29.4%) 1 (5.9%)

0.973ns

Parvimonas micra

2 (11.8%) 0 (0.0%)

Eikenella corrodens

3 (17.6%) 1 (5.9%)

Treponema denticola

3 (17.6%) 1 (5.9%)

Capnocytophagiagingivalis

1 (5.9%) 0 (0.0%)

Lactobacillus paracasei

1 (5.9%) 0 (0.0%)

Lactobacillus salivarius

3 (17.6%) 1 (5.9%)

Aggregatibacter actinomycetemcomitans

0 (0%) 0 (0%)

Prevotella melaninogenica

0 (0%) 0 (0%)

Figure 2 - Mean values of percent % reduction of aerobic, anaerobic

and total bacterial count for the two groups.

Figure 3 - Bar chart showing percentage of bacterial species at S1

and S2 in NaOCl group.

Figure 4 - Bar chart showing percentage of bacterial species at S1

and S2 in Dual rinse + NaOCl group.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

Although the exact cause of postoperative pain is

unknown, it has been linked to various causes, of

which the microbiological aspect is one of most

important factors. Periapical inammation is one

of the most prevalent causes of pain experienced

after RCT as a result of microorganisms remaining

within the root canal or debris extruded apically

in the periapical tissues that cause irritation [22].

The complete eradication of bacteria from

infected root canals through effective irrigation

and instrumentation is a prerequisite for effective

endodontic therapy [23]. Several antimicrobial

agents have been used in endodontics to remove

debris, smear layer, necrotic pulp tissue and to

ensure proper eradication of microorganisms.

Thus, for proper disinfection and smear layer

removal, the combination of NaOCl with a

chelating agent such as EDTA represented the

gold standard. To counteract the problems with

the use of EDTA, it has been suggested to use

the mild chelator HEDP, which has short-term

compatibility with NaOCl [7].

Several in-vitro investigatons [10-16] on

the usage of NaOCl and HEDP together as a root

canal irrigant had promising results. However, it

should be considered that most of these studies

are in-vitro and that those based on evidence are

still lacking.

The present study included thirty-four

participants with necrotic pulp and slight

widening or small periapical radiolucency within

2mm. Large periapical radiolucencies were not

included in the study because they may suggest

a longer-lasting root canal infection and may also

be a sign of extra-radicular infection or cystic

transformation, both of which have the potential

to negatively affect the success rate [24,25].

2.5% NaOCl was used in the current

investigation because it is less cytotoxic than

5.25% NaOCl. Lower NaOCl concentrations

were related to less frequent post-endodontic

discomfort than higher concentrations used in

previous randomized clinical studies treating

nonvital pulps and both concentrations had

similar antibacterial effectiveness [26-29].

In order to prevent the possibility that

intracanal medicament to increase the likelihood

of post-instrumentation pain, bacterial load

reduction and MMP-9 expression, no intracanal

medication was used. Several randomized clinical

trials investigated the effects of irrigants and

irrigation devices without the use of intracanal

medications to avoid their chemical effects,

particularly when extruded [27,30,31].

The results of this study revealed that the

demographic information (age, gender and

tooth type of participants) of the two groups

did not signicantly differ between them. Thus,

provided that the two groups had similar baseline

characteristics which indicate a successful

randomization, removing any confounding

factors and enabling adequate comparison of the

intervention’s impact on the two groups.

Pain assessment by the patients was done at

6, 12, 24 and 48 hours. In order to be sure that

the effects of the local anesthetic worn off, pain

Table IV - Frequency and % values for bacterial species at S1and S2 in Dual rinse+ NaOCl group

Group Bacterial species S1 S2 p-value

Aerobic

Strept mitis/oralis

5 (29.4%) 0 (0%)

0.451 ns

Enterococcus faecalis

5 (29.4%) 2 (11.8%)

Staph hominus

2 (11.8%) 0 (0.0%)

Klebsiella oxytoca

1 (5.9%) 0 (0.0%)

Anaerobic

Fusobacterium spp

5 (29.4%) 2 (11.8%)

0.836 ns

Parvimonas micra

4 (23.5%) 2 (11.8%)

Eikenella corrodens

2 (11.8%) 1 (5.9%)

Treponema denticola

1 (5.9%) 1 (5.9%)

Capnocytophagiagingivalis

1 (5.9%) 0 (0.0%)

Lactobacillus paracasei

0 (0.0%) 0 (0.0%)

Lactobacillus salivarius

1 (5.9%) 1 (5.9%)

Aggregatibacter actinomycetemcomitans

3 (17.6%) 0 (0%)

Prevotella melaninogenica

2 (11.8%) 0 (0%)

Signiﬁcant (

≤ 0.05) ns: non-signiﬁcant (p>0.05).

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

assessment began six hours postoperatively [32].

According to Pak and White [33] the incidence

and degree of postoperative pain reported

by patients is significant in the first day and

diminishes signicantly to low levels throughout

the course of the rst two days, thus a maximum

of 48 hours was chosen.

The majority of the patients showed no

postoperative pain after Dual Rinse + NaOCl or

NaOCl irrigation alone, and if pain occurred, it

was in the mild pain category with no signicant

difference between both irrigants. This came in

agreement with a prior randomized clinical trial

by Ballal et al. [19].

Regarding the effectiveness of the irrigating

solution on intracanal bacterial growth, both

NaOCl and Dual Rinse + NaOCl groups revealed

a considerable decrease in incidence of bacterial

growth after instrumentation in comparison to

pre-instrumentation values. This is consistent

with results from prior clinical trials that

investigated the effectiveness of irrigation and

instrumentation for disinfecting and reducing

intracanal bacteria [34-37].

Dual Rinse + NaOCl irrigation rendered 47%

of the canals free from bacterial growth, while

NaOCl irrigation rendered 41% of canals free from

bacteria, with no signicant difference between

the two irrigating solutions. These results show

that when both irrigants are combined fresh,

HEDP does not reduce the antibacterial activity

of NaOCl, which is highly advantageous in clinical

settings.

The results of our study are in agreement

with a randomized clinical trial by Ballal et al.

[19], which previously showed that irrigation

with a mixture of Dual Rinse HEDP and 2.5%

NaOCl irrigant resulted in the lack of culturable

bacteria in half of the sampled canals, as opposed

to 40% when 2.5% NaOCl was only used.

Our study results are also supported by ndings of

previous clinical trials [35,38] in which irrigation

with 2.5% NaOCl rendered 40-60% of the root

canals free from cultivable bacteria.

The rationale for the antibacterial efcacy

obtained with the Dual Rinse could be explained

by the action of continuous chelation that prevents

the formation of a smear layer, allowing greater

NaOCl penetration into the dentinal tubules,

promoting bacterial reduction. This rationale is

in alignment with previous ndings from other

invitro studies [4,15,16]. Another possible

rationale that the continuous chelation approach

was able to allow for the irrigant to penetrate

and disturb the bacterial biolm matrix, allowing

for improved NaOCl contact with the bacteria,

increasing its antibacterial efcacy. This has been

reported in previous in-vitro studies [10,11].

Percent reduction of intracanal bacteria for

Dual Rinse + NaOCl irrigation was more than

NaOCl group but with no signicant difference

between the two irrigating solutions. The lack

of a significant difference between the two

irrigating solutions in a clinical situation could

be linked to the regular and abundant irrigation

volume [39]. This can enhance the benets of

mechanical preparation and eradication of the

bacteria regardless of the irrigant type utilized.

In this study, there was no evident selection

of either aerobic or anaerobic taxa in any group.

The microorganisms detected inside the root

canals are consistent with ndings from previously

published studies about teeth with primary

asymptomatic apical periodontitis [19,35,40,41]

where the Streptococcus mitis and

Enterococcus

faecalis

were found as the main aerobic taxa.

Staphylococcus

was also identified but less

frequently. The main anaerobic taxa identied was

Fusobacterium

species, while other anaerobic taxa

identied but less frequent than

Fusobacterium

were

Parvimonas micra, Treponema denticola,

Capnocytophagiagingivalis

Prevotella

melaninogenica

. The microbial composition

observed in our study aligns with previous

research findings, which have consistently

reported the prevalence of Gram-negative

anaerobic bacteria in primary untreated root

canal infections with necrotic pulp [19,42,43].

The periapical MMP-9 expression was

measured post-instrumentation and pre-

obturation using periapical fluid analysis.

MMP-9 was chosen as it was demonstrated to

be crucial in apical periodontitis through the

initiation of osteoclastic bone resorption- [44].

MMP-9 is also crucial in inducing pain through

proinammatory cytokines that modulate and

increase expression of MMP-9 by nerve cells

following peripheral nerve injury [45].

The results of periapical levels MMP-9 came

in agreement with a previous study [19] which

showed no statistically significant difference

between both irrigants at both time intervals on

the periapical MMP-9 levels.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

There was a statistically signicant reduction

in MMP-9 levels from post-instrumentation levels

to the pre-obturation levels measured one week

after the rst visit in both groups. This result

is consistent with earlier research that found a

substantial decrease in periapical MMP-9 levels

between the two visits [46,47] and in another

study [19] in which Dual Rinse + NaOCl as well

as NaOCl irrigation alone signicantly reduced

the MMP-9 expression from baseline levels when

teeth were sampled 1 week after the cleaning and

shaping procedure. The antimicrobial activity of

the irrigants may be responsible for the decrease

in MMP-9 levels, which was confirmed by

microbiological analysis.

The clinical significance of the present

work showed that Dual Rinse seems to be a

promising root canal irrigant with no fear of loss

of antibacterial efcacy of NaOCl and no apparent

increase in post-operative pain at clinical levels

or histological levels as determined by levels of

inammatory marker MMP-9.

CONCLUSION

The present study accepted the null

hypothesis. There was no increase in the

incidence of post-instrumentation pain by

the addition of Dual Rinse to 2.5% NaOCl.

The antimicrobial efficacy of the NaOCl was

not affected, with no increase in periapical

MMP-9 and no obvious periapical inammation

in the form of postoperative pain. Both irrigating

solutions successfully reduced bacterial counts

from primary infected root canals, without any

signicant difference between them.

Author’s Contributions

LZIAH: Methodology, Investigation,

Writing – Original Draft Preparation. RMEB:

Conceptualization, Writing – Review &

Editing, Supervision. SIG: Conceptualization,

Methodology, Writing – Review & Editing.

STS: Investigation, Writing – Review & Editing.

NSS: Investigation, Writing – Original Draft

Preparation.

Conict of Interest

No conicts of interest declared concerning

the publication of this article.

Funding

The authors declare that no nancial support

was received.

Regulatory Statement

Every procedure used in the study complied

with the 1964 Helsinki Declaration and its later

2013 revisions, as well as the ethical guidelines

established by Cairo University’s institutional

research council approval number (REF No:19-

7-61).

REFERENCES

1. Alves ACG, Cabau L, Pavan NNO, Tookuni IVM, Viana BAS, Stulp

P,etal. Post-obturation pain and its relationship with reference

time and other risk factors. Braz Dent Sci. 2022;25(3):e3519.

http://doi.org/10.4322/bds.2022.e3519.

2. El Mubarak AH, Abu-bakr NH, Ibrahim YE. Postoperative pain

in multiple-visit and single-visit root canal treatment. J Endod.

2010;36(1):36-9. http://doi.org/10.1016/j.joen.2009.09.003.

PMid:20003932.

3. Almeida G, Marques E, De Martin AS, da Silveira Bueno CE,

Nowakowski A, Cunha RS. Influence of irrigating solution on

postoperative pain following single-visit endodontic treatment:

randomized clinical trial. J Can Dent Assoc. 2012;78:c84.

PMid:22985896.

4. Zehnder M. Root canal irrigants. J Endod. 2006;32(5):389-98.

http://doi.org/10.1016/j.joen.2005.09.014. PMid:16631834.

5. Nair PN, Henry S, Cano V, Vera J. Microbial status of apical root

canal system of human mandibular first molars with primary

apical periodontitis after “one-visit” endodontic treatment. Oral

Surg Oral Med Oral Pathol Oral Radiol Endod. 2005;99(2):231-52.

http://doi.org/10.1016/j.tripleo.2004.10.005. PMid:15660098.

6. Grawehr M, Sener B, Waltimo T, Zehnder M. Interactions of

ethylenediamine tetraacetic acid with sodium hypochlorite in

aqueous solutions. Int Endod J. 2003;36(6):411-7. http://doi.

org/10.1046/j.1365-2591.2003.00670.x. PMid:12801288.

7. Zehnder M, Schmidlin P, Sener B, Waltimo T. Chelation in root

canal therapy reconsidered. J Endod. 2005;31(11):817-20. http://

doi.org/10.1097/01.don.0000158233.59316.fe. PMid:16249726.

8. Neelakantan P, Varughese AA, Sharma S, Subbarao CV, Zehnder

M, De-Deus G. Continuous chelation irrigation improves the

adhesion of epoxy resin-based root canal sealer to root dentine.

Int Endod J. 2012;45(12):1097-102. http://doi.org/10.1111/j.1365-

2591.2012.02073.x. PMid:22612994.

9. Zollinger A, Mohn D, Zeltner M, Zehnder M. Short-term storage

stability of NaOCl solutions when combined with Dual Rinse

HEDP. Int Endod J. 2018;51(6):691-6. http://doi.org/10.1111/

iej.12875. PMid:29121393.

10. Novozhilova N, Babina K, Polyakova M, Sokhova I, Sherstneva

V, Zaytsev A,etal. The effect of different compositions and

concentrations of etidronate-containing irrigants on the

antibacterial activity of sodium hypochlorite against

Enterococcus

faecalis

and

Candida albicans.

Dent J. 2024;12(3):46. http://doi.

org/10.3390/dj12030046. PMid:38534270.

11. Campello AF, Rodrigues RCV, Alves FRF, Miranda KR, Brum SC,

Mdala I,etal. Enhancing the intracanal antibacterial effects of

sodium hypochlorite with etidronic acid or citric acid. J Endod.

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

2022;48(9):1161-8. http://doi.org/10.1016/j.joen.2022.06.006.

PMid:35750221.

12. Arias-Moliz MT, Ordinola-Zapata R, Baca P, Ruiz-Linares M, Ferrer-

Luque CM. Antimicrobial activity of a sodium hypochlorite/

etidronic acid irrigant solution. J Endod. 2014;40(12):1999-2002.

http://doi.org/10.1016/j.joen.2014.07.031. PMid:25266466.

13. Morago A, Ordinola-Zapata R, Ferrer-Luque CM, Baca P, Ruiz-

Linares M, Arias-Moliz MT. Influence of smear layer on the

antimicrobial activity of a sodium hypochlorite/etidronic acid

irrigating solution in infected dentin. J Endod. 2016;42(11):1647-

50. http://doi.org/10.1016/j.joen.2016.07.023. PMid:27616541.

14. Paqué F, Rechenberg DK, Zehnder M. Reduction of hard-tissue

debris accumulation during rotary root canal instrumentation

by etidronic acid in a sodium hypochlorite irrigant. J Endod.

2012;38(5):692-5. http://doi.org/10.1016/j.joen.2011.12.019.

PMid:22515905.

15. Aoun C, Rechenberg D-K, Karam M, Mhanna R, Plotino G, Zogheib

C. Effect of continuous chelation irrigation using DualRinse

HEDP+3% NaOCl with or without high-power sonic activation

on debris and smear layer removal. Eur Endod J. 2023;8(2):162-9.

http://doi.org/10.14744/eej.2022.93064. PMid:37010200.

16. Lottanti S, Gautschi H, Sener B, Zehnder M. Effects of

ethylenediaminetetraacetic, etidronic and peracetic acid

irrigation on human root dentine and the smear layer. Int

Endod J. 2009;42(4):335-43. http://doi.org/10.1111/j.1365-

2591.2008.01514.x. PMid:19220516.

17. La Rosa GRM, Plotino G, Nagendrababu V, Pedullà E.

Effectiveness of continuous chelation irrigation protocol in

endodontics: a scoping review of laboratory studies. Odontology.

2024;112(1):1-18. http://doi.org/10.1007/s10266-023-00835-8.

PMid:37433927.

18. Gondim E Jr, Setzer FC, Dos Carmo CB, Kim S. Postoperative

pain after the application of two different irrigation devices in a

prospective randomized clinical trial. J Endod. 2010;36(8):1295-

301. http://doi.org/10.1016/j.joen.2010.04.012. PMid:20647083.

19. Ballal NV, Gandhi P, Shenoy PA, Shenoy Belle V, Bhat V,

Rechenberg DK,etal. Safety assessment of an etidronate in a

sodium hypochlorite solution: randomized double-blind trial. Int

Endod J. 2019;52(9):1274-82. http://doi.org/10.1111/iej.13129.

PMid:30993696.

20. Qazi SS, Manzoor MA, Qureshi RA, Khan HH. Comparison of

postoperative pain- normal saline vs sodium hypochlorite as

irrigants. Pak Oral Dent J. 2005;25(2):177-82.

21. R Core Team. R: a language and environment for statistical

computing [Internet]. Vienna: R Foundation for Statistical

Computing; 2022 [cited 2024 mar 25]. Available from: https://

www.R-project.org/

22. Sipavičiūtė E, Manelienė R. Pain and flare-up after endodontic

treatment procedures. Stomatologija. 2014;16(1):25-30.

PMid:24824057.

23. Haapasalo M, Shen Y, Wang Z, Gao Y. Irrigation in endodontics.

Br Dent J. 2014;216(6):299-303. http://doi.org/10.1038/

sj.bdj.2014.204. PMid:24651335.

24. Nair PN. On the causes of persistent apical periodontitis: a

review. Int Endod J. 2006;39(4):249-81. http://doi.org/10.1111/

j.1365-2591.2006.01099.x. PMid:16584489.

25. Ng YL, Mann V, Gulabivala K. A prospective study of the factors

affecting outcomes of nonsurgical root canal treatment: part 1:

periapical health. Int Endod J. 2011;44(7):583-609. http://doi.

org/10.1111/j.1365-2591.2011.01872.x. PMid:21366626.

26. Verma N, Sangwan P, Tewari S, Duhan J. Effect of different

concentrations of sodium hypochlorite on outcome of primary

root canal treatment: a randomized controlled trial. J Endod.

2019;45(4):357-63. http://doi.org/10.1016/j.joen.2019.01.003.

PMid:30827769.

27. Mostafa M, El-Shrief YAI, Anous WIO, Hassan MW, Salamah FTA,

El Boghdadi RM,etal. Postoperative pain following endodontic

irrigation using 1.3% versus 5.25% sodium hypochlorite in

mandibular molars with necrotic pulps: a randomized double-

blind clinical trial. Int Endod J. 2020;53(2):154-66. http://doi.

org/10.1111/iej.13222. PMid:31563148.

28. Frough-Reyhani M, Ghasemi N, Soroush-Barhaghi M, Amini M,

Gholizadeh Y. Antimicrobial efficacy of different concentration

of sodium hypochlorite on the biofilm of

Enterococcus

faecalis

at different stages of development. J Clin Exp

Dent. 2016;8(5):e480-4. http://doi.org/10.4317/jced.53158.

PMid:27957257.

29. Zand V, Lotfi M, Soroush MH, Abdollahi AA, Sadeghi M, Mojadadi

A. Antibacterial efficacy of different concentrations of sodium

hypochlorite gel and solution on

Enterococcus

faecalis

biofilm.

Iran Endod J. 2016;11(4):315-9. http://doi.org/10.22037/

iej.2016.11.PMC5069909. PMid:27790262.

30. Ng YL, Glennon JP, Setchell DJ, Gulabivala K. Prevalence of and

factors affecting post-obturation pain in patients undergoing

root canal treatment. Int Endod J. 2004;37(6):381-91. http://

doi.org/10.1111/j.1365-2591.2004.00820.x. PMid:15186245.

31. Nagendrababu V, Gutmann JL. Factors associated with

postobturation pain following single-visit nonsurgical root

canal treatment: a systematic review. Quintessence Int.

2017;48(3):193-208. http://doi.org/10.3290/j.qi.a36894.

PMid:27669726.

32. Singh RD, Khatter R, Bal RK, Bal CS. Intracanal medications versus

placebo in reducing postoperative endodontic pain--a double-

blind randomized clinical trial. Braz Dent J. 2013;24(1):25-9.

http://doi.org/10.1590/0103-6440201302039. PMid:23657409.

33. Pak JG, White SN. Pain prevalence and severity before, during,

and after root canal treatment: a systematic review. J Endod.

2011;37(4):429-38. http://doi.org/10.1016/j.joen.2010.12.016.

PMid:21419285.

34. McGurkin-Smith R, Trope M, Caplan D, Sigurdsson A. Reduction

of intracanal bacteria using GT rotary instrumentation, 5.25%

NaOCl, EDTA, and Ca(OH)2. J Endod. 2005;31(5):359-63. http://

doi.org/10.1097/01.don.0000145035.85272.7c. PMid:15851929.

35. Siqueira JF Jr, Rocas IN, Paiva SS, Guimaraes-Pinto T, Magalhaes

KM, Lima KC. Bacteriologic investigation of the effects of sodium

hypochlorite and chlorhexidine during the endodontic treatment

of teeth with apical periodontitis. Oral Surg Oral Med Oral Pathol

Oral Radiol Endod. 2007;104(1):122-30. http://doi.org/10.1016/j.

tripleo.2007.01.027. PMid:17499528.

36. Xavier AC, Martinho FC, Chung A, Oliveira LD, Jorge AO, Valera

MC, et al. One-visit versus two-visit root canal treatment:

effectiveness in the removal of endotoxins and cultivable

bacteria. J Endod. 2013;39(8):959-64. http://doi.org/10.1016/j.

joen.2013.04.027. PMid:23880258.

37. Elbattawy EM, El Shafei J, Mehanna N, Ahmed G, Ghareeb A,

Montasser K. The effect of irrigating solutions on postoperative

pain and intensity of bacteremia following single visit root canal

treatment. Sylwan. 2019;16(11):236-60.

38. Rôças IN, Provenzano JC, Neves MA, Siqueira JF Jr. Disinfecting

effects of rotary instrumentation with either 2.5% sodium

hypochlorite or 2% chlorhexidine as the main irrigant: a

randomized clinical study. J Endod. 2016;42(6):943-7. http://

doi.org/10.1016/j.joen.2016.03.019. PMid:27142579.

39. Carvalho MC, Zuolo ML, Arruda-Vasconcelos R, Marinho

ACS, Louzada LM, Francisco PA, et al. Effectiveness of

XP-Endo Finisher in the reduction of bacterial load in oval-

shaped root canals. Braz Oral Res. 2019;33:e021. http://doi.

org/10.1590/1807-3107bor-2019.vol33.0021. PMid:31508722.

40. Ercan E, Ozekinci T, Atakul F, Gul K. Antibacterial activity of

2% chlorhexidine gluconate and 5.25% sodium hypochlorite

in infected root canal: in vivo study. J Endod. 2004;30(2):84-

Braz Dent Sci 2024 Oct/Dec;27 (4): e4316

Abdel-Hamid LZI et al.

Assessment of Dual Rinse combined with sodium hypochlorite irrigating solution on post-instrumentation pain and bacterial load reduction: a randomized clinical trial

Abdel-Hamid LZI et al. Assessment of Dual Rinse combined with sodium hypochlorite

irrigating solution on post-instrumentation pain and bacterial

load reduction: a randomized clinical trial

7. http://doi.org/10.1097/00004770-200402000-00005.

PMid:14977302.

41. Soares JA, Pires DR Jr. Influence of sodium hypochlorite-

based irrigants on the susceptibility of intracanal microbiota

to biomechanical preparation. Braz Dent J. 2006;17(4):310-

6. http://doi.org/10.1590/S0103-64402006000400009.

PMid:17262145.

42. Hommez GM, Verhelst R, Claeys G, Vaneechoutte M, De

Moor RJ. Investigation of the effect of the coronal restoration

quality on the composition of the root canal microflora in

teeth with apical periodontitis by means of T-RFLP analysis.

Int Endod J. 2004;37(12):819-27. http://doi.org/10.1111/j.1365-

2591.2004.00884.x. PMid:15548272.

43. Rocas IN, Siqueira JF Jr. Frequency and levels of candidate

endodontic pathogens in acute apical abscesses as

compared to asymptomatic apical periodontitis. PLoS

One. 2018;13(1):e0190469. http://doi.org/10.1371/journal.

pone.0190469. PMid:29293651.

44. Corotti MV, Zambuzzi WF, Paiva KB, Menezes R, Pinto LC, Lara

VS, et al. Immunolocalization of matrix metalloproteinases-2

and -9 during apical periodontitis development. Arch

Oral Biol. 2009;54(8):764-71. http://doi.org/10.1016/j.

archoralbio.2009.04.013. PMid:19497558.

45. Ji RR, Xu ZZ, Wang X, Lo EH. Matrix metalloprotease regulation

of neuropathic pain. Trends Pharmacol Sci. 2009;30(7):336-40.

http://doi.org/10.1016/j.tips.2009.04.002. PMid:19523695.

46. Wahlgren J, Salo T, Teronen O, Luoto H, Sorsa T, Tjäderhane L.

Matrix metalloproteinase-8 (MMP-8) in pulpal and periapical

inflammation and periapical root-canal exudates. Int Endod

J. 2002;35(11):897-904. http://doi.org/10.1046/j.1365-

2591.2002.00587.x. PMid:12453017.

47. Alptekin NO, Ari H, Haliloglu S, Alptekin T, Serpek B, Ataoglu T. The

effect of endodontic therapy on periapical exudate neutrophil

elastase and prostaglandin-E2 levels. J Endod. 2005;31(11):791-

5. http://doi.org/10.1097/01.don.0000158010.43884.59.

PMid:16249720.

Laila Zakaria Ismail Abdel-Hamid

(Corresponding address)

Cairo University, Faculty of Dentistry, Department of Endodontics, Cairo, Egypt.

Email: laila.zakaria@dentistry.cu.edu.eg; laila.zakaria1990@gmail.com

Date submitted: 2024 Mar 25

Accept submission: 2024 July 29