Effect of intracanal and extracanal heating on pulp dissolution property of continuous chelation irrigant.

Context: Extracanal and intracanal heating of sodium hypochlorite (NaOCl) improve its pulp dissolution, but limited literature is available on its effect as a combined single irrigant with etidronate. Aim: The aim of this study is to compare the effect of temperature on the effectiveness of NaOCl and continuous chelation protocol on the time required for the dissolution of vital and necrotic pulp. Materials and
Methods: Dissolution time of 120 standardized bovine (buffalo) pulp fragments, divided into 12 subgroups based on tissue type (vital/necrotic), irrigant (NaOCl/continuous chelation), and temperature (extracanal, intracanal, and nonheated irrigant), was noted. Conical glass tips mimicking the root canal were considered specimen containers. About 0.2 ml of irrigant corresponding to the irrigation protocol was taken in them and then pulp samples were added to it. Samples were observed using loupes under 2.5 X magnification. Dissolution time was recorded using a stopwatch. The study was approved by the Institutional Ethical Committee (SDC/2019/591). Statistical Analysis Used: Two-way analysis of variance; statistical product and service solutions version 25. The level of significance was set at P < 0.05.
Results: Time for pulp dissolution by continuous chelation mixture was significantly more as compared to NaOCl alone in all subgroups. Pulp tissue dissolution for both vital and necrotic pulp was improved by the increase in temperature of both irrigants and dissolution time was more for necrotic than vital tissue. Pulp tissue dissolution was significantly better by intracanal heating as compared to extracanal heating.
Conclusion: Although intracanal heating of continuous chelation mixture improves its pulp dissolution capacity significantly as compared to extracanal heating and nonheating protocol but pulpal dissolution capacity of nonheated 5% NaOCl still remains significantly better as compared to intracanal and extracanal heated continuous chelation mixture. Copyright:

INTRODUCTION

Success in endodontic therapy is mainly dependent on the removal of the vital and necrotic pulp tissue, microorganisms, and their toxins. However, certain root canal spaces such as delta loop, isthmus, accessory canals, and dentinal tubules remain inaccessible and often remain uninstrumented and may be a reason for endodontic treatment failure due to residual infected organic or inorganic tissue.[1] Pulp tissue can be eliminated from the inaccessible areas by utilizing the solvent ability of endodontic irrigants.[2]

There are a variety of irrigants available; however, sodium hypochlorite (NaOCl) remains the irrigant of choice to date because of its excellent antimicrobial properties, vital, and necrotic tissue dissolving ability, and flushing action.[3] Its chemical efficacy can be enhanced by altering its concentration, temperature[4], and pH.[5]

Preheated NaOCl solution has a greater ability to dissolve pulp tissue and cleanse the root canal.[6] Woodmansey[7] reported that hypochlorite at boiling temperature can disintegrate the pulp tissue at a speed of 210 times higher as compared to room temperature. NaOCl can be warmed by preheating it in syringes. However, the preheated solutions stabilize the body temperature within seconds.

Iandolo et al.[8] proposed intracanal heating of NaOCl for better removal of smear layer and observed that it is possible to obtain higher temperature for a longer time with this technique as opposed to extracanal heating.

NaOCl has a limited effect on the inorganic component of the smear layer which is removed by ethylenediaminetetraacetic acid, thus making it indispensable for root canal treatment; however, it not only has a harmful erosive effect on dentine but has raised environmental concerns as well. This led to the introduction of biocompatible chelator etidronate, which has a less erosive effect on dentine. Lottanti et al.[9] and Bedir et al.[10] showed that etidronate could be used in combination with NaOCl without affecting its antimicrobial or proteolytic properties. Hence, the combination of NaOCl and 1-hydroxyethylidene-1,1-bisphosphonate, or etidronate was proposed to be used as a single irrigant in the biomechanical preparation[11] which lead to the concept of continuous chelation in 2005.

Although studies by Zehnder et al.[11] and Ulusoy et al.[12] states that 1-hydroxyethane-1,1-diphosphonic acid (HEDP) scarcely influences the short-term loss of NaOCl properties but there are studies that have observed the negative impact of HEDP on pulp dissolution capacity of NaOCl.[1314]

Furthermore, literature lacks in studies evaluating the influence of the type of pulpal tissue and method of warming of NaOCl on the pulp dissolution properties of continuous chelation protocol, especially that of Twin Kleen, which was introduced by Maarc Dental in 2018, hence this study was conducted, and the null hypothesis tested was that the use of intracanal (100°C) and extracanal (65°C) heating has no effect on the time required for dissolution of vital and necrotic pulp by ultrasonically agitated NaOCl and a mixture of NaOCl and etidronate.

MATERIALS AND METHODS

The study was approved by the Institutional Ethical Committee (SDC/2019/591). Freshly extracted bovine (buffalo) teeth were collected from meatpacking plant of Meerut and stored at–20°C for 1 h. The teeth were thawed to room temperature, longitudinally sectioned, and the pulp was extirpated. One hundred twenty pulp samples were standardized by weight (6 ± 0.5 milligrams each) using analytical balance and volume (5 mm × 3 mm × 1.5 mm [mm]) to limit the effect of these parameters on dissolution time.

Poststandardization, the pulp samples were randomly divided into two groups – V (Vital n = 60) and N (Necrotic n = 60). About 60 fresh bovine pulp tissue samples were incubated at 36°C for 2 days to simulate necrotic pulp. Samples in each group were further distributed into six subgroups of 10 samples each (n = 10) depending on the irrigation protocol as follows:

Group 1 included 5% NaOCl (H) and vital pulp (V). It was further divided into three groups based on the heating of the irrigant, i.e., HVN: Nonheated NaOCl, HVI: Intracanal heated NaOCl, and HVE: Extracanal heated NaOCl.

Group 2 included 5% NaOCl (H) and necrotic pulp (N). It was further divided into HNN: Nonheated NaOCl, HNI: Intracanal heated NaOCl, and HNE: Extracanal heated NaOCl.

Group 3 included 5% NaOCl (H) and etidronic acid (E) and vital pulp (V). It was further divided into HEVN: Nonheated continuous chelation mixture, HEVI: Intracanal heated continuous chelation mixture, and HEVE: Extracanal heated continuous chelation mixture.

Group 4 included 5% NaOCl (H) and etidronic acid (E) and necrotic pulp (N). It was further divided into HENN: Nonheated continuous chelation mixture, HENI: Intracanal heated continuous chelation mixture, and HENE: Extracanal heated continuous chelation mixture.

According to the manufacturer's recommendation, the solution for continuous chelation protocol was freshly prepared by mixing 10 ml of 5% NaOCl with two capsules of Twin Kleen for 60 s just before use.

Conical glass tips mimicking root canal (1 ml volume) were considered as specimen containers having 15 mm length and 7 mm upper and 5 mm lower diameter. About 0.2 ml of 5% NaOCl or 5% NaOCl and etidronate mixture corresponding to the irrigation protocol was taken in them, and then vital or necrotic pulp samples were added to it.

Subgroup E – Extracanal heating of the irrigant was done by immersing the irrigant syringe into a water bath set at 65°C.

Subgroup I-Intracanal heating of the irrigant was done by placing the Calamus heated plugger of small size 40/.025 (Dentsply Maillefer, Switzerland) at 100°C with apico-coronal movements to within 3 mm short of specimen container tip for 5 s.

Ultrasonic activation was performed with 25/02 ultrasonic tip (Ultra X, Eighteeth Medical) immersed in the irrigating solutions to a depth of 5 mm away from the pulp specimens, without touching them for 15 s/min. Irrigant was replenished after every 2 min till complete dissolution (observed under loupes having 2.5 × magnification). Time was recorded using a stopwatch. Complete dissolution was considered when no tissue fragment was visible in the solution.

The endpoint for the complete dissolution was visually assessed by two observers to eliminate observer bias and in case of difference in opinion, the mean value was recorded.

The data were analyzed statistically using two-way analysis of variance using the Statistical Product and Service Solutions version 25 (IBM Corp. Armonk, New York, USA); the level of significance was set at P < 0.05.

RESULTS

Intergroup and intragroup comparisons showed statistically significant differences (P < 0.05) in the mean time required for pulp dissolution among all the groups [Tables 1 and 2].

Table 1

Time taken (minutes) for pulp dissolution with their mean, standard deviation, maximum and minimum time scores

Group name	Parameters	HVN	HVI	HVE
NaOCl and vital pulp (Group 1)	Mean time	18.600	11.700	14.200
	SD	1.174	0.823	1.317
	Maximum time	20	13	16
	Minimum time	17	11	12
Group name	Parameters	HNN	HNI	HNE
NaOCl and necrotic pulp (Group 2)	Mean time	40.700	21.000	27.900
	SD	0.823	1.633	0.738
	Maximum time	42	24	29
	Minimum time	40	19	27
Group name	Parameters	HEVN	HEVI	HEVE
NaOCl+etidronic acid and vital pulp (Group 3)	Mean Time	192.200	128.300	157.600
	S.D.	2.573	0.949	1.506
	Maximum time	196	130	160
	Minimum time	189	127	155
Group name	Parameters	HENN	HENI	HENE
NaOCl+etidronic acid and necrotic pulp (Group 4)	Mean Time	483.100	291.300	350.100
	SD	3.071	2.319	2.470
	Maximum time	487	295	347
	Minimum time	479	288	354

HVN: Non-heated NaOCl, HVI: Intracanal heated NaOCl, HVE: Extracanal heated NaOCl, HNN: Non-heated NaOCl, HNI: Intracanal heated NaOCl, HNE: Extracanal heated NaOCl, HEVN: Non-heated continuous chelation mixture, HEVI: Intracanal heated continuous chelation mixture, HEVE: Extracanal heated continuous chelation mixture, HENN: Non-heated continuous chelation mixture, HENI: Intracanal heated continuous chelation mixture, HENE: Extracanal-heated continuous chelation mixture, SD: Standard deviation, NaOCl: Sodium hypochlorite

Table 2

Two-way analysis of variance - F-test for comparing difference in time taken for dissolution of pulp

Pair of groups	Probable values of two-way ANOVA-F test
	Nonheated AND Intracanal heating	Intracanal heating and extracanal heating	Nonheated and extracanal heating
NaOCl + vital pulp and NaOCl + necrotic pulp	0.0002*	0.0019*	0.0034*
NaOCl + vital pulp and NaOCl + etidronic acid + vital pulp	0.0012*	0.0005*	0.0007*
NaOCl + vital pulp and NaOCl + etidronic acid + necrotic pulp	0.0008*	0.0006*	0.0002*
NaOCl + necrotic pulp and NaOCl + etidronic acid + vital pulp	0.0004*	0.0017*	0.0011*
NaOCl + necrotic pulp and NaOCl + etidronic acid + necrotic pulp	0.0005*	0.0007*	0.0033*
NaOCl + etidronic acid + vital pulp and NaOCl + etidronic acid + necrotic pulp	0.0001*	0.0003*	0.0002*

*A significant difference between subgroups at 0.05 level of significance, i.e., P<0.05. NaOCl: Sodium hypochlorite, ANOVA: Analysis of variance

Pulp tissue dissolution for both vital and necrotic pulp was improved by an increase in temperature of both NaOCl and continuous chelation mixture. Time taken by intracanal heated irrigant was significantly less than the time taken for extracanal heated irrigant to dissolve pulp tissue, and time taken by extracanal heated irrigant was significantly less as compared to that of nonheated irrigant, P < 0.05.

The time taken for pulp dissolution by continuous chelation mixture was significantly more as compared to NaOCl alone, P < 0.05.

In both the irrigation protocols, the time taken for dissolution of necrotic tissue was significantly more than vital pulp tissue, P < 0.05.

DISCUSSION

Bovine pulp was selected in the study to represent the organic remnants present in root canals because of its similarity to human pulp tissue and ease of obtaining an adequate amount of tissue which allowed easier standardization.[15]

The[16] incubated fresh tissue at 36°C for 2 days to simulate necrotic tissue. A similar methodology was opted in this study.

After 2 min of contact with organic tissue, most of the activity of NaOCl is lost. To ensure the availability of active molecules and to remove remnants of dissolved tissue, frequent replenishment is therefore essential.[4]

Studies by Tartari et al.[13] and Stojicic et al.[4] have shown better pulp dissolution with ultrasonic activation of NaOCl. Hence, a similar activation protocol was used in this study too. The present investigation was standardized using the same volume of irrigant and time of agitation during each replenishment. The mechanism of passive ultrasonic action has been attributed to acoustic streaming and cavitation. The motion of fluids and irrigants increases by the energy produced from ultrasonic, which improves the contact between dentinal walls and the irrigant, thus accentuating the action of irrigants.

On intergroup comparison, it was seen that necrotic tissue took significantly more time to dissolve than vital pulp tissue. A study by AbouRass and Oglesby[17] found similar results in which fresh rat dermal tissue dissolved more rapidly than necrotic tissue.

The degradation of fatty acids and lipids present in organic tissue resulting in soap and glycerol formation (saponification reaction) explains the mechanism of tissue dissolution. On coming in contact with necrotic tissue, a more rapid fall in chlorine concentration (as compared to normal tissue) and rapidly diminishing potency of NaOCl were observed by Austin and Taylor.[18] Stojicic et al.[4] advocated a frequent irrigant renewal to compensate for this fall. However, despite following this protocol, the ability of NaOCl to dissolve necrotic tissue remained compromised in this study.

The dissolution susceptibility of necrotic and vital pulp tissue may be different because of their structural and compositional difference. The loss in vascular supply and water content in necrotic tissues, according to Wang et al.[19] would probably lead to structural changes such as shrinkage, decrease in thickness, and stiffening leading to a much denser pulpal structure that might be resistant to dissolution.

However, there are contrasting opinions. Hasselgren et al.[20] in their study observed that necrotic tissues dissolved more easily as compared to that of fresh samples. This may be attributed to the vascularity, which could have resisted the action of NaOCI. The differences observed with these previous studies may be attributed to the difference in tissue type, the surface area of contact, the difference in experimental design, and methodology.

The results of the present study were in accordance with Tartari et al.[13] and Vyavahare et al.[14] which showed that the association of etidronate with NaOCl negatively interfered with the dissolution ability of NaOCl. Hypochlorite ion (OCl−), which prevails in alkaline solutions is related to a greater tissue dissolution capacity, according to Macedo et al.[21] When compared to NaOCl, the mixture of NaOCl, and etidronate is more acidic, which might account for lesser dissolution ability due to reduced OCl− ion present in the mixture. In addition, Zehnder et al.[22] emphasized the free available chlorine loss in NaOCl by etidronate, which continues over time and is dose-dependent.

On the contrary, no significant changes in the tissue dissolution ability of NaOCl alone as compared to continuous chelation mixture were reported by Ulusoy et al.[12] This disparity could be due to basic compositional differences in etidronate supplied by different manufacturers.

Increased effectiveness of NaOCl has been achieved by altering its concentration, time of exposure, pH, and temperature. Increased irrigant temperatures can be achieved by either preheating solutions outside the canal or by intracanal heating.[7]

Extracanal heating of NaOCl in the present study was performed using a water bath set at 65°C as it is thought to provide controlled temperature settings compared to kettle and bottle warmers used in previous studies.[23] Rossi – Fedele and De Figueiredo[23] observed that heating NaOCl enhances the pulp dissolution activity but reaches a plateau around 60°C–75°C. This could be because, after a particular time or temperature increase, the available chlorine becomes exhausted.

Intracanal heating was performed at the lowest temperature setting of the Calamus plugger (100°C) by heating the irrigant inside the specimen container tip. Maximum free chlorine is liberated by NaOCl at boiling temperature (96°C–120°C). Hence, no additional benefit is observed by heating above this temperature. In addition, heating above this increases the risk of jeopardizing periradicular tissues by either heat transfer through dentine or accidental extrusion.[24]

The results of this study revealed that pulp dissolution was significantly faster in samples treated with intracanal and extracanal heated NaOCl and continuous chelation mixture as compared to nonheated irrigants. Heating the NaOCl might have made its molecules more agitated, which increased collisions with organic matter, and consequently, its dissolution.[6] In addition, the renewal of the mixture would have kept the free available chlorine content high.

As compared to extracanal heating, intracanal heating had significantly increased pulp dissolution capacity. Prewarmed irrigant usually stabilizes to body temperature, whereas intracanal heating maintains temperature for longer periods; this enhances irrigants efficacy due to increased reaction rate, chemical interaction with the tissues, and increase in the release of free available chlorine.

The null hypothesis was thus rejected. Intracanal and extracanal heating of irrigant significantly decreased the time of dissolution of vital and necrotic pulp tissue with both NaOCl and continuous chelation mixture. However, the pulp dissolution capacity of even nonheated NaOCl was significantly more as compared to extracanal and intracanal heated continuous chelation mixture. Thus, emphasizing that both modes of heating could not compensate for the excess time required in pulp dissolution by continuous chelation mixture.

Due to limitations in obtaining human pulp, this study was performed with bovine pulp tissue since their characteristics are comparable to those of human pulp tissue. However, structural differences do exist.

In addition, while applying these results to clinical situations, there are certain factors to be kept in mind. Pulp being a closed structure is limited by the dentinal walls, which may have a dampening effect on the ultrasonic activation protocols.

However, future studies with a larger sample size and better clinical simulation would be required to validate the findings of this study.

CONCLUSION

Within the limitations of this study, it can be concluded that warming of continuous chelation mixture significantly increases its pulp dissolution ability, with intracanal heating being significantly better as compared to extracanal heating. However, increasing the temperature could not compensate for the loss in pulp dissolution property of continuous chelation irrigation when compared to NaOCl.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.