Keywords demineralization - dual rinse HEDP - root canal irrigation - sodium hypochlorite -
surface roughness - Triton - tissue dissolution
Introduction
The successful outcome of root canal therapy is determined by the methodology and
efficacy of instrumentation, irrigation, disinfection, and three-dimensional (3D)
obturation of the entirety of the root canal system.[1 ] The intricate structure of the root canal system is of prime importance in this
regard. Complex microanatomical features, including isthmuses, lateral canals, and
apical ramifications, can provide shelter for biofilms and soft tissue debris. These
remnants may subsequently lead to reinfection and act as a nutrient source for microbes
in root canals that have undergone previous treatment.[2 ] As a result, clinicians widely accept that the more intricate the endodontic anatomy,
the greater is the necessity for enhanced methods of chemo-mechanical root canal debridement.
This process heavily depends on the application of irrigation solutions. Various agents
have been recommended for endodontic irrigation. Among these, sodium hypochlorite
(NaOCl) is regarded as the gold standard and a premier choice,[3 ] as it possesses both tissue dissolution and antimicrobial properties.[4 ]
[5 ] The pulpal tissue dissolution is a highly desirable characteristic of any irrigating
solution, as it facilitates both the cleansing of the root canal system and the removal
of necrotic tissue.
Mechanical instrumentation of root canals results in the formation of smear layer
on the walls of the canal. This layer comprises inorganic dentin particles and organic
matter, including pulp tissue, odontoblastic processes, necrotic material, microorganisms,
and their metabolic byproducts.[6 ] These components can accumulate in the intricate structures of the root canal system,
such as fins and ramifications.[7 ]
[8 ] Consequently, along with NaOCl, the use of an additional decalcifying substance
is advised. In the past, this was accomplished by switching between the NaOCl irrigant
and a complementary solution containing ethylenediaminetetraacetic acid (EDTA).[9 ] However, EDTA presents with certain limitations such as its decreased efficacy in
removal of smear layer from apical third of the root canal system[10 ] and its cytotoxicity.[11 ]
Furthermore, it substantially reduces the calcium-phosphate ratio of root dentin,
which subsequently alters the permeability, surface roughness, and microhardness of
the root dentin.[12 ]
Of late, a new dental product containing 1-hydroxyethylidene-1,1-diphosphonic acid
(HEDP), called the Dual Rinse HEDP (DR HEDP), has been introduced to the market by
Medcem GmbH in Weinfelden, Switzerland. This product, supplied in a capsule form,
comprises of 0.9 g of etidronate powder, which is to be mixed with 10 mL of NaOCl
solution (0.5–5.25%) just before use in order to create an all-in-one irrigant.[13 ] This formulation allows dental professionals to employ DR HEDP solution in a “continuous
chelation” method for root canals in the instrumentation phase and through the entire
duration of treatment session.[14 ] Primary benefit of DR HEDP is its ability to function as a single irrigant for both
cleaning and disinfecting root canals, significantly reducing irrigation time.[15 ]
According to Rath et al,[16 ] employing a continuous chelation protocol led to a more uniform organic and inorganic
composition of the dentin surface compared with sequential chelation using NaOCl followed
by EDTA. Earlier research has shown that DR HEDP exhibits antimicrobial properties
similar to NaOCl while being nontoxic.[17 ]
[18 ] Additionally, DR HEDP matches NaOCl in its ability of organic tissue dissolution[19 ] and enhances the adhesion of sealers and cements to root canal dentin.[20 ]
[21 ]
[22 ]
A new all-in-one endodontic irrigation solution called Triton has been introduced
by Brasseler in Savannah, United States. This product is composed of various chemical
compounds, including 1,2,4-butanetricarboxylic acid, 2-phosphono, citric acid, sodium
dodecylbenzenesulfonate, alcohol, polyethylene glycol 4-(tert-octylphenyl) ether,
sodium lauryl sulfate, 2-ethylhexyl sodium sulfate, sodium cumenesulphonate, sodium
hydroxide, and sodium hypochlorite. The solution comes in two separate components,
part A and part B, which are combined by employing an automix technique to create
a single solution prior to application.
Current research has shown that Triton outperforms traditional irrigants such as NaOCl
and EDTA in removing debris and smear layers,[23 ] as well as dissolving soft tissues.[24 ] A recent study demonstrated Triton to have enhanced antimicrobial activity and organic
tissue dissolution efficacy when compared with EndoJuice.[25 ]
Till date, literature presents with no studies that have compared the physical and
chemical properties of DR HEDP and Triton. Hence, the objectives of the present ex vivo study were (1) to evaluate the efficacy of DR HEDP and Triton on soft tissue dissolution;
(2) to evaluate the effect of DR HEDP and Triton on surface roughness of root canal
dentin; and (3) to determine the demineralization effect of a DR HEDP and Triton on
root canal dentin.
The null hypotheses tested in the study were: (1) there is no difference in the ability
of DR HEDP and Triton in dissolving soft tissue; (2) there is no difference in the
surface roughness produced by DR HEDP and Triton; and (3) there is no difference in
the demineralization produced by DR HEDP and Triton.
Materials and Methods
Ethical clearance for the use of human extracted teeth for the experiments performed
in the present study was obtained from the institutional review board prior to the
commencement of the study (555/2024).
Sample Size Estimation
The G*Power software (Heinrich Heine University, Dusseldorf, Germany) was used to
estimate sample size. With 95% confidence level, 90% power, standard deviation of
0.15, and mean difference of 0.217, the minimum sample size was calculated to be 10
for each group for all the parameters tested in this study.[19 ]
[26 ]
[27 ]
Soft Tissue Dissolution
Sample Preparation
Forty extracted human single-rooted teeth with fully formed apices containing one
straight single were selected. All the teeth were radiographed and analyzed under
magnifying loupes (EyeMag Smart, Carl Zeiss, Oberkochen, Germany) to verify the presence
of a single canal with closed apex, and the absence of intraradicular resorption,
caries, cracks, or root canal filling. Superficial soft tissues on the root surface
were removed with a curette and the teeth were stored in saline with 0.2% sodium azide
(Millipore Sigma, St. Louis, Missouri, United States) at 4°C until use. The teeth
were decoronated using a diamond disc (Horico Dental, Berlin, Germany) to standardize
the root length. Working length was established by inserting a size 10 K file (Dentsply
Sirona Endodontics, Ballaigues, Switzerland) into each root canal until it was just
visible at the apical foramen and by subtracting 1 mm from the recorded length. Chemo-mechanical
preparation was performed using ProTaper Universal nickel-titanium rotary instruments
(Dentsply Sirona), with a torque control electric motor (X-Smart). The root canals
were instrumented to size F3 (30/6%). Irrigation was performed using 2 mL of a DR
HEDP (Medcem) combined with 3% NaOCl solution (Vista Dental, Racine, Wisconsin, United
States) for 1 minute between each instrument change. One capsule (0.9 g) of DR HEDP
powder was mixed with 10 mL of 3% NaOCl solution. Canals were finally irrigated with
5 mL of distilled water for 1 minute. The canals were dried with sterile paper points
(Dentsply Sirona). Horizontal reference grooves of 0.5 mm depth were first made on
the buccal and lingual surfaces of middle third of each root to serve as a guide for
indicating the standard locations where simulated resorptive cavities were to be made
later in the root canal. Each root specimen was then embedded in Eppendorf tubes (Merck,
Darmstadt, Germany) using a putty impression material (3M ESPE, St. Paul, Minnesota,
United States). Once that material set, the roots were removed from the vials to enable
assembly and reassembly of the specimens. Each root was then sectioned longitudinally
into two halves using a diamond disc (Horico Dental) under water cooling. Semicircular
cavities were then be made on the root canal in the middle third (at the previously
marked reference groove) of each root half using no. 6 round diamond point (Horico
Dental) with slow speed under water cooling. The bur was marked 2 mm from its tip
using a permanent marker, to ensure precise depth of the cavity. The depth of resorptive
defect was standardized to 4 mm, which was confirmed by a digital caliper. The resorptive
cavities were then irrigated with 5 mL of DR HEDP (Medcem) combined with 3% NaOCl
(Vista Dental) for 1 minute to remove the smear layer. Finally, the cavities were
flushed with 5 mL of distilled water for 1 minute and dried using paper points (Dentsply
Sirona) ([Fig. 1 ]).
Fig. 1 Schematics illustrating the stages of specimen preparation for the analysis of soft
tissue dissolution: (A ) intact single-rooted tooth; (B ) decoronation; (C ) simulated resorptive cavities; and (D ) reassembled root halves maintaining canal patency.
Soft Tissue Preparation
Freshly extracted shrimp meat was employed for the experiment to simulate pulp tissue.
The tissue samples used were standardized in weight and placed in the resorptive cavities
of both halves of each root. Prior to weighing, each piece of tissue was blotted on
Whatman filter paper (Merck, Darmstadt, Germany), dried, and then weighed using a
precision balance placed in an airtight container (AT 261, Mettler, Greifensee, Switzerland).
Both root halves were then reassembled using a light-curing resin barrier (OpalDam,
Ultradent Products, South Jordan, Utah, United States). Patency was maintained by
placing an F3 ProTaper gutta-percha point (Dentsply Sirona) between the canals of
the two root sections. The apex of each root was sealed with sticky wax to simulate
a closed-end system. A plastic tube of 3 mm height was glued at the cementoenamel
junction (CEJ) of each root to act as a reservoir for the irrigating solution. Each
root was then placed in its respective Eppendorf vial, and the samples were randomly
divided using a sequence generator (
www.random.org
) into four groups (n = 10) based on the irrigation regimen described below.
Irrigation Techniques
The irrigant volume was standardized to 5 mL, and the irrigation time was 5 minutes
for all samples.
Group 1: 3% NaOCl solution (Vista Dental) combined with DR HEDP (Medcem)
Group 2: Triton (Brasseler)
Group 3: 3% NaOCl (Vista Dental)
Group 4: Distilled water (control)
In the DR HEDP group, one capsule of DR HEDP (0.9 g) was dissolved in 10 mL of the
3% of NaOCl (Vista Dental) solution and used immediately. In the Triton group, using
the automix technique, the solutions from both components (A and B) were aspirated
using a syringe as recommended by manufacturer and were used immediately. All irrigating
solutions were introduced into the canal using a 30-gauge side-vented stainless steel
needle (Vista Dental). The needle tip was inserted 1 mm short of the working length
in each canal. After the irrigation protocols, the canals in groups 1, 2, and 3 were
flushed with 5 mL distilled water for 1 minute to remove any precipitate that may
have formed. The roots were then removed from the Eppendorf vials, split into two
halves, and the meat tissue from the resorptive cavities was reweighed. Before weighing,
all meat specimens were blotted dry on Whatman filter paper (Merck). All measurements
were performed by a single blinded investigator who was unaware of the test solutions
used. The percentage of tissue weight loss or gain was calculated by subtracting the
posttreatment weight from the initial weight, dividing by the original weight, and
multiplying by 100 ([Fig. 2 ]).
Fig. 2 Flowchart of the methodology employed for soft tissue dissolution.
Demineralization Analysis
Demineralization Analysis
Sample Preparation
Forty intact human single-rooted permanent mandibular first premolars, with fully
formed apices and single straight canals were obtained from individuals aged between
18 and 30 years. The teeth were extracted for orthodontic reasons. The rationale for
selecting a relatively young age group (orthodontic patients) is to avoid the confounding
variable of excessive calcification of root canal dentin. All the samples were radiographed
and examined under magnifying loupes (EyeMag Smart) to verify canal anatomy and exclude
teeth with resorption, calcification, caries, cracks, or previous endodontic treatment.
Superficial soft tissue remnants on the root surfaces were removed using a curette,
and the teeth were stored in saline with 0.2% sodium azide (Millipore Sigma) at 4°C
until use.
Irrigation Techniques
Samples were randomly divided into four groups (n = 10) using the aforementioned technique. Chemo-mechanical preparation was performed
using ProTaper Universal NiTi rotary instruments (Dentsply Sirona) with an electric
motor (X-Smart, Dentsply). Root canals were instrumented to F3 (30/0.06). Between
each file, 2 mL of the experimental irrigant solution was delivered for 1 minute,
followed by 5 mL of final rinse with the same irrigant solution. Irrigants were introduced
using a 30-gauge side-vented needle (Vista Dental) inserted 1 mm short of the working
length. The irrigant effluent from each sample was collected in a test tube placed
beneath the Eppendorf tube holding the specimen. These aliquots were prepared for
atomic absorption spectrometry (AAS) to quantify calcium ion concentration.
AAS Analysis
Calcium levels were determined using a flame atomic absorption spectrometer (iCE 3000
series, Thermo Fisher Scientific Inc., United States) at a wavelength of 422.7 nm.
Due to high calcium concentrations in the test samples, a 10-fold dilution exceeded
the upper detection limit of the instrument. Therefore, all experimental samples were
diluted 100-fold prior to analysis to bring the concentrations within the linear range
(0.05–3.0 mg/L) and simplify calculations. A sensitivity check was performed at 0.5 mg/L,
with an expected absorbance range of 0.25 to 0.50 AU. Background correction was employed.
Standard stock solutions (1000 µg/mL) were diluted to prepare working standards (1,
3, 5, and 10 µg/mL), and a calibration curve was constructed (R
2 = 0.992) ([Fig. 3 ]).
Fig. 3 Flowchart of the methodology employed for demineralization analysis.
Surface Roughness Analysis
Sample Preparation
Twenty extracted human single-rooted teeth were selected based on the same criteria
as mentioned above for the demineralization analysis. Teeth were decoronated at the
CEJ using a diamond bur (Horico Dental) under water coolant. Each root was longitudinally
sectioned into two dentin slices using a diamond saw microtome (Leica SP 1600, Leica
Microsystems, Wetzlar, Germany). Forty dentin segments were horizontally embedded
in autopolymerizing acrylic resin (Meliodent RR, Kulzer, Hanau, Germany), exposing
the root canal dentin surface. The dentin surface of each root segment was polished
with a grinder-polisher, under water using a series of silicon carbide papers with
increasing fineness (500, 800, 1,000, and 1,200 grit). Polishing was performed under
distilled water. Final polishing was accomplished using 0.1 μm alumina suspension
(Ultra-Sol R; Eminess Tec Inc, Monroe, North Carolina, United States) on a rotary
felt disk. The samples were then randomly divided into four groups (n = 10) using the aforementioned technique. Each sample was immersed in a glass plate
containing 15 mL of the respective experimental solution for 6 minutes at room temperature.
All the glass plates were placed on a vibrator to agitate the experimental solutions.
The experimental solutions were renewed after the treatment of each specimen. Thereafter,
the specimens were rinsed with 50 mL of distilled water and blotted dry.
Surface Roughness Analysis
Surface roughness (Ra, µm) was measured using atomic force microscopy (AFM; Veeco,
Santa Barbara, California, United States) in contact mode with silicon nitride tips.
The instrument was calibrated using polyethylene spheres. Scans were performed by
a blinded, trained operator. Three regions (10 × 10 µm2 ) per specimen were imaged, and the average Ra value was computed for each specimen
using image analysis software ([Fig. 4 ]).
Fig. 4 Flowchart of the methodology employed for surface roughness analysis.
Statistical Analysis
Statistical analysis was performed using IBM SPSS Statistics version 26 (IBM Corp.,
Armonk, New York, United States). The normality of data for all three quantitative
parameters (soft tissue dissolution, surface roughness, and demineralization) was
assessed using the Kolmogorov–Smirnov and Shapiro–Wilk tests. One-way analysis of
variance was applied to compare the mean values of the three parameters among the
four experimental groups. Tukey's honest significant difference test was conducted
for post hoc pairwise comparisons to identify intergroup differences. The significance
level was set at p < 0.05. All values were reported as mean ± standard deviation, along with 95% confidence
intervals.
Results
Soft Tissue Dissolution
[Fig. 5 ] summarizes the soft tissue dissolution data for the control and experimental groups.
Preirrigation tissue weights were not significantly different across the experimental
and control groups (p = 0.859). Postirrigation tissue weights varied significantly among the experimental
groups. DR HEDP (36.49 ± 2.77%) and NaOCl alone (35.57 ± 1.80%) demonstrated the highest
tissue dissolution with no significant difference between them (p = 0.589). Triton (29.50 ± 1.37%) exhibited significantly less tissue dissolution
compared with both DR HEDP and NaOCl alone groups (p < 0.001). Distilled water, the negative control, showed no tissue dissolution effect
(1.67 ± 0.15%).
Fig. 5 Mean percentage of soft tissue dissolution in different experimental groups. Bars
represent the mean % of tissue loss observed with DR HEDP, Triton, NaOCl, and distilled
water. Error bars indicate standard deviation.
Demineralization Effect
[Fig. 6 ] summarizes the demineralization data for the control and experimental groups. Samples
treated with Triton™ released the highest amount of calcium ions (135.61 ± 3.99 mg/L),
compared with DR HEDP (49.66 ± 3.05 mg/L) and NaOCl alone (26.06 ± 3.18 mg/L) groups
(p < 0.001). There was no significant difference between DR HEDP and NaOCl alone groups
(p > 0.05). Samples treated with distilled water showed the least amount of calcium
ion release (36.39 ± 2.96 mg/L).
Fig. 6 Mean calcium ion concentration (mg/L) in different experimental groups. Bars depict
the mean (Ca2+ ) concentration measured for Triton, DR HEDP, NaOCl, and distilled water. Error bars
represent standard deviation.
Surface Roughness
[Fig. 7 ] summarizes the surface roughness data for the control and experimental groups. Triton
demonstrated the highest mean surface roughness (191.37 ± 3.95 nm), compared with
the DR HEDP (104.25 ± 3.02 nm) and NaOCl alone (101.24 ± 4.06 nm) groups (p < 0.001). There was no significant difference between the DR HEDP and NaOCl alone
groups (p = 0.244). Distilled water showed the least surface roughness (36.39 ± 2.96 nm). [Fig. 8 ] demonstrates the 3D AFM images of dentin surfaces after treating with DR HEDP, Triton,
NaOCl, and distilled water.
Fig. 7 Mean surface roughness in different experimental groups. Bars represent the mean
surface roughness observed with DR HEDP, Triton, NaOCl, and distilled water. Error
bars indicate standard deviation.
Fig. 8 Three-dimensional atomic force microscopic images of root canal dentin surfaces after
treatment with the experimental agents. Triton had maximum rough surface compared
with DR HEDP and NaOCl. Distilled water showed minimal surface roughness. (A ) Triton; (B ) DR HEDP; (C ) NaOCl; (D ) distilled water.
Discussion
Previous literature evaluated the individual performance of DR HEDP and Triton when
they were used as root canal irrigants.[13 ]
[25 ]
[28 ] However, no comparative studies have evaluated the effectiveness of these two all-in-one
irrigants in tissue dissolution, surface roughness, and their demineralizing potential.
The findings of the present study demonstrated that Triton had less soft tissue dissolution
ability, compared with NaOCl alone and the combination of NaOCl with HEDP (DR HEDP).
Hence, the first null hypothesis that “there is no difference in the ability of DR
HEDP and Triton in dissolving soft tissue” has to be rejected. This result supports
the earlier work by Ballal et al,[17 ] who reported that etidronate does not interfere with NaOCl's proteolytic effect
when used in continuous chelation. Novozhilova et al[29 ] observed that etidronate-containing irrigants maintained stable chlorine levels
and dissolution efficacy over time, further validating the superior soft tissue dissolution
efficacy of DR HEDP. The poor soft tissue dissolution performance of Triton could
be attributed to its complex formulation, which includes acidic and surfactant components
that could reduce the availability of active chlorine when premixed, thereby compromising
its proteolytic activity. Deepa et al[24 ] reported that while Triton is an effective tissue dissolving agent, its activity
under standard conditions may be less than that of traditional NaOCl-based irrigants.
When the surface roughness analysis of the irrigants was examined, samples treated
with Triton demonstrated the highest surface roughness when compared with DR HEDP
and NaOCl alone. Hence, the second null hypothesis that “there is no difference in
the surface roughness produced by DR HEDP and Triton” must be rejected. The increased
surface roughness of root dentin caused by Triton can be majorly attributed to the
presence of citric acid and 1,2,4-butanetricarboxylic acid in its formulation. Citric
acid, being a strong chelator, removes smear layer effectively and causes dentin erosion,
which might have increased the surface roughness.[30 ]
[31 ]
[32 ] 1,2,4-Butanetricarboxylic acid is a well-known chelating agent with both phosphonic
acid and carboxylic acid groups, which would have enhanced the effect of Triton in
smear layer removal, thus increasing the surface roughness. The category of surfactant
present in Triton could also contribute to its superior smear layer removal leading
to increased surface roughness. Incorporation of surfactants may improve the efficiency
of an irrigant by decreasing surface tension and improving wettability, thereby enabling
superior contact with the root canal walls. In contrast, DR HEDP exhibited a roughness
profile comparable to NaOCl alone, indicating that etidronate exerts only a gentle
chelating effect[28 ] and does not significantly exacerbate surface degradation. This aligns with previous
research where DR HEDP was found to preserve the organic–inorganic balance of dentin
and enhance adhesive outcomes without overetching.[16 ]
[20 ] Clinically, excessive roughness may compromise the structural integrity of root
dentin and increase susceptibility to crack formation during obturation,[33 ]
[34 ]
[35 ] underscoring the importance of preserving dentin morphology during irrigation.
The decalcification effects of the experimental irrigants revealed that, samples treated
with Triton had the most superior release of calcium ions, compared with both DR HEDP
and NaOCl. Hence, the third null hypothesis that “there is no difference in the decalcification
produced by DR HEDP and Triton” must be rejected. These findings are in accordance
with Castagnola et al,[23 ] who observed marked smear layer removal and deep dentin demineralization following
Triton irrigation. The higher demineralization potential of Triton could be owed to
the acidic pH of citric acid present in its formulation.[36 ]
[37 ] Apart from this, the chelating activity of 1,2,4-butanetricarboxylic acid could
have contributed to the increased demineralization potential of Triton. Conversely,
DR HEDP released approximately half the amount of calcium as Triton, supporting its
categorization as a soft chelator, due to its less aggressive action in terms of demineralization.[28 ] These results are consistent with the longstanding view that etidronate preserves
dentin mineral content while still facilitating sufficient smear layer removal.[13 ]
[14 ] The NaOCl when used alone had minimal dentin decalcification effect due to its poor
chelating efficacy.[38 ]
Sodium hypochlorite is utilized as an endodontic irrigant with the concentrations
ranging from 0.5 to 8.25%, without a universally accepted consensus on the optimum
concentration.[33 ]
[39 ] The current study employed the use of 3% NaOCl as an irrigant since previous studies
showed that more concentrated solutions could affect the mechanical properties of
root canal dentin.[40 ] Apart from this, 3% NaOCl when mixed with HEDP stayed relatively stable for up to
2 hours, with minimal loss of free available chlorine. This presents optimum working
time for clinicians by decreasing the need for chairside mixing.[13 ]
The significance of the present work lies in its capacity to identify the more effective
and less detrimental root canal irrigant that might prolong the survival of root canal-treated
teeth. DR HEDP when used as an all-in-one irrigant provided optimal soft tissue dissolution,
and minimal damage to root canal dentin matrix. In contrast, Triton appeared to be
more aggressive toward dentin. This might potentially affect the structural and mechanical
properties of root canal dentin, a significant consideration in the success of root
canal treatment. By decreasing the structural damage to root dentin matrix, DR HEDP
when used in a continuous chelation approach may potentially enhance the outcome of
root canal therapy. Future research must focus on validating the presented findings
in a clinical setting and explore the long-term follow-ups of these irrigation protocols.
The present work was an ex vivo preliminary investigation into the physical and chemical properties of Triton and
DR HEDP. Results of this study present with a few limitations that may not necessarily
be generalized in clinical situations. First, the soft tissue dissolution model employed
shrimp meat, which is although analogous to human pulp tissue, it lacks the vital
responses and vasculature of in situ pulp tissue. Also, this study also employed single-rooted human teeth with standardized
cavities in the root canals, which may not have provided an overall representation
of the varied root canal anatomies encountered in day-to-day clinical situations.
Second, the evaluation of dentin surface roughness and demineralization was performed
on standardized root slices, which may not capture the regional variations observed
in clinical root canals.
Considering the limitations of the current ex vivo study, it may be concluded that DR HEDP and NaOCl alone produced minimal dentin surface
roughness and demineralization, with superior soft tissue dissolution efficacy compared
with Triton.
