Natural nanoparticles versus the gold standard in direct pulp capping: a randomized clinical trial

Abstract

This study compared the efficacy of nanopropolis and nanocurcumin as direct pulp-capping agents to that of mineral trioxide aggregate (MTA) in recently erupted permanent molars, assessing the clinical and radiographic outcomes over six months. Thirty six permanent molars were randomly allocated to three groups (n = 12 each): MTA (control), nanopropolis and nanocurcumin. The study followed a double-blind design. Clinical and radiological evaluations assessed pain, percussion sensitivity and swelling preoperatively and one week, three months and six months afterwards. At one week, the nanocurcumin group showed significantly higher pain levels than the MTA and nanopropolis groups. The pain scores equalised by three months and remained stable until six months in all groups. Percussion sensitivity and swelling occurred in 8.3% of cases in the MTA group, 25% in the nanopropolis group and 16.7% in the nanocurcumin group. Nanopropolis and nanocurcumin provided clinical and radiographic outcomes comparable to MTA in direct pulp capping of recently erupted permanent molars. While MTA showed slightly superior attributes, the natural nanomaterials present viable alternatives. This study supports the potential use of bioactive nanomaterials derived from natural sources in vital pulp therapy, though longer-term investigations are warranted.

Introduction

Vital pulp therapy preserves pulpal tissues that have been compromised by deep caries, trauma, previous restorative procedures or iatrogenic causes (Hanna et al. 2020; Shallal-Ayzin et al. 2021). Direct pulp capping is a technique in which a small mechanical or carious pulpal exposure is covered with a protective material that provides a tight bacterial seal. This technique may be performed when pulpal exposure occurs through noninfected dentine and the tooth has a history of reversible pulpitis (Zou et al. 2024).

Calcium hydroxide was the gold-standard pulp capper for exposed pulp in permanent dentition. However, calcium hydroxide alone does not have adequate sealing ability and may predispose the pulp surface to inflammation, necrosis and tunnel defects in the newly created dentinal bridge (Alqahtani et al. 2023). Mineral trioxide aggregate (MTA), which forms calcium hydroxide upon setting, has overcome these limitations (Chen et al. 2018).

MTA consists of tricalcium silicate, dicalcium silicate, tetracalcium aluminoferrite, tricalcium aluminate and gypsum. It has low solubility, high biocompatibility and good sealing and reparative dentine-forming ability (Saikia et al. 2023). It also stimulates the migration, adhesion and attachment of undifferentiated cells, thus facilitating the formation of a dentine bridge, although with additional handling, costs, and setting time (Alsubait et al. 2014; Zhu et al. 2014).

Recently, several nanomaterials have been developed as promising pulp-capping alternatives. These nanomaterials have stable structures, improved mechanical properties, decreased crack propagation, improved bonding to the tooth structure and excellent antibacterial properties (Parirokh et al. 2018). Metallic nanoparticles have also been shown to have superior antimicrobial effects, but they may have unfavourable side effects, so there has been a change in research direction towards natural nanoparticle products, including nanopropolis and nanocurcumin (Deshmukh et al. 2022). Propolis has superb antimicrobial, antioxidant, immunomodulatory and anti-inflammatory properties that are beneficial for caries prevention, root canal disinfection and direct and indirect pulp capping. It may also function as a good storage medium for avulsed teeth and a nurturing medium for accelerating surgical wound healing because it neutralises microbial infections, inflammatory processes and pulpal necrosis and stimulates reparative dentine formation (Alghutaimel et al. 2024; Azmoudeh and Nazeri 2023). Curcumin also exhibits antioxidant and anti-inflammatory properties (Hameeda et al. 2020).

Post-operative pain after pulp capping arises from nociceptor activation by inflammatory mediators, though anti-inflammatory agents can help reduce it (Shallal-Ayzin et al. 2021). Although studies highlight the antibacterial and anti-inflammatory properties of nanopropolis and nanocurcumin, clinical evidence for their efficacy in direct pulp capping is limited. This study evaluates their effectiveness in young permanent molars, testing the null hypothesis that they perform similarly to MTA.

Materials and methods

This interventional, randomised, prospective clinical trial, conducted at the *** University, with ethical approval number, RECO6U/33–2022 and its meeting held on November 12, 2022 and was registered online at www.clinicaltrials.gov, and all data regarding the aim of the study, number of patients enrolled, inclusion and exclusion criteria, starting participants, outcomes and estimated trial completion dates were recorded. Sample size (n = 12) per group, was computed using G*Power (α = 0.05, β = 0.2, power = 80%, d = 0.55), as previously reported (Nasri et al. 2022). The materials used included MTA (Dentsply Maillefer, Tulsa, USA), nanopropolis (Nano Gate, Cairo, Egypt), nanocurcumin (Nano Gate, Cairo, Egypt), glass ionomer and composite.

Materials preparation

Following a previous study (Zaleh et al. 2022), The nanoparticle synthesis protocol for the propolis began with 100 g of raw propolis dissolved in 500 mL of 80% ethanol. This solution underwent continuous agitation via hot plate stirrer (MSH- 20 A; witeg Labortechnik, Wertheim, Germany) for seven days, followed by filtration. The filtrate was subsequently diluted with distilled water at a 1:10 ratio to precipitate propolis particles. Ultrasonication was performed for 20–30 min, and then propolis nanoparticles were generated before loading them into a plastic syringe.

Curcumin-loaded zinc oxide nanoparticles were synthesised by first dispersing 0.925 g of nano-ZnO powder in 50 mL of ethanol, followed by 30 min of simultaneous stirring and sonication. Following the addition of 0.075 g of native curcumin, the solution was agitated overnight in a sealed vessel (Nano Gate, Cairo, Egypt) and then dried for 24 h at 60 °C before loading them into a plastic syringe.

Participant selection

Thirty-six patients referred to a teaching hospital were selected for this study. Subjects were aged 7–13 years and presented with permanent molars with mature or immature apex, exhibiting reversible pulpitis with pinpoint pulpal exposure not exceeding 2 mm without systemic pathologies. The probe was carefully positioned at the site of pulp exposure to determine its diameter. Exclusion criteria were irreversible pulpitis, sustained pulpal haemorrhage exceeding 10 min post-exposure, pulpal necrosis, percussion sensitivity, aberrant mobility, tissue inflammation, sinus tract formation, non-restorable dentition, radiographic evidence of resorption, calcified canals, inter-radicular osseous deterioration or coreesponding pathosis. The patient did not need to take analgesic. We refrained from applying the vitality test as children may give false results. The challenge with vitality testing in such cases lies in the potential for false-positive or false-negative results, particularly in teeth with immature apices, where nerve responsiveness may not correlate directly with pulp health. Therefore,it remains contingent upon patient cooperation and the inherent limitations of available pulp sensibility assessments. All the patients’ parents provided written informed consent after being given details of the treatment’s benefits/risks, number of visits and the expected duration of the research. All consent forms were in Arabic.

Procedure

The investigation comprised 36 samples allocated randomly into three groups of 12 samples each, which were differentiated by the pulp-capping agent used: the control group received direct MTA application, the first experimental group underwent direct propolis nanoparticle application and the second experimental group received direct curcumin nanoparticle application. To ensure randomisation, the participants were assigned codes shuffled electronically. Group membership was double-blinded.

The procedure began with the administration of local anaesthesia. The cavity outline was then prepared using a high-speed round burr with irrigation, and carious soft dentine in the vicinity of the pulp was removed with spoon excavator. Once the pulp was exposed and determined to meet the inclusion criteria, characterised by transient odontalgic responses to thermal stimuli without spontaneous exacerbation, absence of nocturnal discomfort, well-circumscribed mild to moderate pain that abated promptly upon stimulus withdrawal, normal pulpal sensibility testing parameters, and radiographic absence of periapical pathosis—despite the paradoxical presence of minimal pulpal exposure (≤ 2 mm) that traditionally connotes irreversible inflammatory compromise, the pulp tissue was disinfected with 2.5% sodium hypochlorite (NaOCl), and haemostasis was achieved by pressure over the exposed pulp using cotton pellets moistened with 2.5% NaOCl for 10 min (Suhag et al. 2019). Persistent hemorrhagic pulps (> 10 min) suffices to exclude the treated tooth and operate pulpotomy as indicated.

In the nanopropolis and nanocurcumin groups, propolis was injected from a pre-loaded syringe onto a glass slab, then transported to the exposure site with a metal carrier, while in the MTA group, ProRoot MTA powder was mixed as per instructions and applied using a metal carrier. In nanoproplis and nanocurcumin Groups, collagen sponge (Guangdong Victory Biotech, china) was cut in dimensions of 2.5 × 2.5 and was placed as a barrier after the material. To ensure optimal application of nanopropolis and nanocurcumin, the consistency of the mixture should be carefully observed before placement. The nanoparticle solution should be well-dispersed and homogenous to maximise bioavailability and therapeutic efficacy. Utilising sterile glass slab provides a smooth surface for even distribution, preventing clumping and ensuring a controlled amount of material is applied. Additionally, the collagen sponge serves as a mechanical barrier, preventing premature material loss and maintaining localised effects by creating a stable microenvironment for tissue healing and regeneration. Using a well-calibrated metal carrier allows for precise delivery to the exposure site, reducing the risk of contamination or excess material placement. As MTA requires moisture for proper setting, maintaining a slightly damp field without excessive fluid contamination is essential for successful application. Proper adaptation and condensation techniques ensure complete coverage of the exposure site, promoting effective sealing and tissue response.

For composite resin palcement as a final restoration, all cavity walls were firstly itched by phosphoric acid 37% according to manufacturer instructions, followed by All-Bond Bisco universal adhesive (BISCO, Inc. Schaumburg, U.S.A) was applied according to manufacturer instructions. Composite layers were eventually applied and light cured incrementaly at 20_second each. Finishing and polishing were performed, then recalling for patients for follow-ups.

Assessments and follow-up

Preoperative and follow-up exams at 1 week, 3 months, and 6 months assessed pain, swelling, and tenderness using the VAS scale. Treatment outcome assessment involved clinical and radiographic evaluations at baseline and follow-up intervals of 3, 6, and 12 months. Standardised periapical radiographs were taken using parallel technique with beam alignment devices and evaluated by two calibrated examiners for abdormalities. Treatment was considered to have failed if clinical symptoms of irreversible pulpitis or necrosis, radiographic evidence of pathosis, or requirement for additional endodontic intervention was detected. Failed cases were documented with specific causes and time points of failure. Participants’ guardians were advised to administer analgesics (325 mg acetaminophen tablets) to their children on experiencing pain no more than every six hours and to record all administrations. The procedure was successful when the tooth was asymptomatic, functional and responded normally to percussion with no radiographic pathosis. In severely pained patients, especially with persistent swelling and percussion pain, apexogenesis or root canal treatments were performed; these were considered failed treatments. In persisting pain, analgesics were prescribed for 1 week. There was no difference between groups during analgesic prescription. Indicated inferential statistical tests were computed.

Results

Clinical results

Inter- and intra-examiner reliability using Cohen’s kappa coefficient to quantify agreement detected no inconsistencies, while the double-blinding of both patients and evaluators is minimized biases that could influence subjective assessments. The examiners followed pre-established precise definitions of clinical endpoints such as pain perception, swelling, and periapical changes to ensure that self-reported symptoms such as pain on the VAS scale are as accurate and reflective of clinical reality as possible.

Pain score group comparisons

Intergroup analysis of the pain scores revealed no statistically significant differences in the mean pre-operative scores between the groups (MTA: 3.17 ± 0.83; nanopropolis: 2.92 ± 0.67; nanocurcumin: 3.17 ± 0.72; p = 0.608). At one-week post-intervention, the nanocurcumin group experienced significantly higher pain levels (1.5 ± 0.8), followed by the MTA (0.75 ± 0.97) and nanopropolis (0.67 ± 0.98; p = 0.034) groups. By three months, the mean pain scores were no longer significantly different between the groups (MTA: 0.42 ± 0.79; nanopropolis: 0.5 ± 0.67; nanocurcumin: 0.25 ± 0.45; p = 0.637). This persisted through six months (MTA: 0.08 ± 0.29; nanopropolis: 0.25 ± 0.62; nanocurcumin: 0 ± 0; p = 0.337).

Pain score change over time

As shown in Table 1, the mean pain scores in the MTA group decreased from 3.17 ± 0.83 preoperatively to 0.75 ± 0.97 after seven days, 0.42 ± 0.79 after three months and 0.08 ± 0.29 after six months (p < 0.001). The mean pain scores in the nanopropolis group decreased from 2.92 ± 0.67 preoperatively to 0.67 ± 0.98 after seven days, 0.5 ± 0.67 after three months and 0.25 ± 0.62 after six months (p < 0.001). The mean pain scores in the nanocurcumin group decreased from 3.17 ± 0.72 preoperatively to 1.5 ± 0.8 after seven days, 0.25 ± 0.45 after three months and 0.00 ± 0.00 after six months (p < 0.001). Table 2 displays intra-group comparison of pain scores for each group (Friedman test), whilst Table 3 show the computation of Wilcoxon signed Rank test for Pairwise comparison of pain scores.

Table 1.

Descriptive statistics of pain scores pre and post operatively and comparison between groups (Kruskal Wallis test)

		Median	Mean	Std. Dev	95% Confidence Interval for Mean		Min	Max	P value
					Lower Bound	Upper Bound
Pre_ Operative	Group I (MTA)	3a	3.17	.83	2.64	3.70	2.00	4.00	.608 ns
	Group II (Nano propolis)	3a	2.92	.67	2.49	3.34	2.00	4.00
	Group III (Nanocurcumin)	3a	3.17	.72	2.71	3.62	2.00	4.00
One Week	Group I (MTA)	1b	.75	.97	.14	1.36	.00	3.00	.034*
	Group II (Nano propolis)	0c	.67	.98	.04	1.29	.00	3.00
	Group III (Nanocurcumin)	2a	1.50	.80	.99	2.01	.00	3.00
Three Month	Group I (MTA)	0a	.42	.79	-.09	.92	.00	2.00	.637 ns
	Group II (Nano propolis)	0a	.50	.67	.07	.93	.00	2.00
	Group III (Nanocurcumin)	0a	.25	.45	-.04	.54	.00	1.00
Six Month	Group I (MTA)	0a	.08	.29	-.10	.27	.00	1.00	.337 ns
	Group II (Nano propolis)	0a	.25	.62	-.14	.64	.00	2.00
	Group III (Nanocurcumin)	0a	.00	.00	.00	.00	.00	.00

Table 2.

Descriptive statistics and intra group comparison of pain scores at different observation times within each group (Friedman test)

Time	Group I (MTA)		Group II (Nano propolis)		Group III (Nanocurcumin)
	Median	Mean ± Std. Dev	Median	Mean ± Std. Dev	Median	Mean ± Std. Dev
Pre_ operative	3a	3.17 ±.83	3a	2.92 ±.67	3a	3.17 ±.72
One Week	1b	.75 ±.97	0b	.67 ±.98	2b	1.50 ±.80
Three Month	0c	.42 ±.79	0b	.50 ±.67	0c	.25 ±.45
Six Month	0c	.08 ±.29	0b	.25 ±.62	0c	.00 ±.00
P value within the same groups	0.000*		0.000*		0.000*

Table 3.

Detailed results of Wilcoxon signed Rank test for pairwise comparison of pain scores

Groups		1 week Versus Pre-operative	3 month versus Pre-operative	6 month Versus Pre-operative	3 month versus 1 week	6 month versus 1 week	3 month versus 6 month
Group I (MTA)	Z	− 2.965	− 2.965	− 3.100	-.850	− 2.060	− 1.414
	P value	.003*	.003*	.002*	.035*	.039*	.157 ns
Group II (Nano propolis)	Z	− 2.989	− 3.114	− 3.126	-.412	− 1.089	− 1.732
	P value	.003*	.002*	.002*	.680 ns	.276 ns	.083 ns
Group III (Nanocurcumin)	Z	− 2.913	− 3.108	− 3.115	− 2.714	− 2.994	− 1.732
	P value	.004*	.002*	.002*	.007*	.003*	.083 ns

Significance level p ≤ 0.05, *significant, ns = non-significant

Post hoc test: within the same column, values sharing the same superscript letter are not significantly different

Percussion and swelling

Percussion and swelling were observed in one case (8.3%) in the MTA group, three cases (25%) in the nanopropolis group and two cases (16.7%) in the nanocurcumin group. These differences were not statistically significant (p = 0.548).

Radiographic changes

Radiographic evaluations at three months had unremarkable findings for all groups. At six months, one case (8.3%) in the MTA group and two (16.7%) in the nanopropolis group exhibited periodontal membrane space widening, while one case (8.3%) in the nanopropolis group and two (16.7%) in the nanocurcumin group manifested periapical radiolucency. These were statistically insignificant (p = 0.627), as shown in Table 4.

Table 4.

Descriptive statistics and comparison of frequency of clinical and radiographic changes in different groups (Chi square test)

	Percussion and swelling		Radiographic changes
			3 months		6 months			P (intragroup between 3 & 6 months)
	Yes	No	Yes	No	WPL	PR	No
Group I (MTA)	1 (8.3%)	11 (91.7%)	0	12 (100%)	1 (8.3%)	0	11 (91.7%)	0.307 ns
Group II (Nano- propolis)	3 (25%)	9 (75%)	0	12 (100%)	2 (16.7%)	1 (8.3%)	9 (75%)	0.064 ns
Group III (Nanocurcumin)	2 (16.7%)	10 (83.3%)	0	12 (100%)	0	2 (16.7%)	10 (83.3%)	0.139 ns
P value (between groups)	0.548 ns		1 ns		0.627 ns

WPL widening of periodontal membrane space, PR periapical radiolucency

Significance level p ≤ 0.05, ns = non-significant

Discussion

The efficacy of pulp capping as a minimally invasive vital pulp therapy hinges on bacterial management and reparative dentinogenesis of incompletely formed molar roots, and MTA has become the gold standard due to its biocompatibility and regenerative properties (Shallal-Ayzin et al. 2021) over time. Propolis, meanwhile, provides anti-inflammatory action through lipoxygenase pathway inhibition and TGF-β1 production, facilitating odontoblast differentiation and collagen synthesis, while nanocurcumin offers greater bioavailability, stability and wound-healing properties than conventional curcumin, matching MTA’s anti-inflammatory efficacy (Shi et al. 2019). Dentin sialophosphoprotein (DSPP) expresses odontoblast differentiation, with evidence that nano-MTA significantly upregulates DSPP transcription within 24–72 h following application, which correlates with enhanced mineralisation. Studies have also shown that the calcium silicate composition of nano-MTA particles (50–100 nm) facilitates sustained calcium ion release, triggering DSPP expression (Parolia et al. 2010).

Clinical evaluation at three and six months revealed comparable success rates among MTA, propolis, and curcumin treatments. Propolis’s efficacy stems from its antibacterial properties and flavonoid-mediated immune modulation. This aligns with Kusum et al.’s findings of equivalent success rates among MTA, propolis, and Biodentine at three and six months (Kusum et al. 2015). However, statistical analysis indicated MTA’s superior clinical performance over propolis, corroborating previous clinical, radiographic, and histological evidence. The nano-scale properties of these materials, particularly surface area and ion release kinetics, significantly influence these molecular responses. Smaller particle sizes (30–100 nm) generally demonstrate enhanced cellular uptake and more rapid initiation of signalling cascades than conventional materials. MTA was superior, but propolis remained a viable alternative (Chen et al. 2020).

The non-sginificant results between curcumin and MTA groups in this study echoes previous findings (Hugar et al. 2017; Hugar et al. 2017) Also, Kumar et al. (Kumar et al. 2017) used turmeric powder for pulpotomy of permanent teeth and reported a success rate of 93.4% after 6 months. Prabhakar et al. (Prabhakar et al. 2019) used curcumin compared with MTA efficacy in rats. Both formed dentinal bridge inflammation, and laid new tissue organisation, confirming curcumin’s anti-inflammatory and antimicrobial properties (Mandrol et al. 2016). Thus, nano-propolis and nanocurcumin perform comparably to MTA in young permanent pulp capping.

Conclusion

Nano-propolis and nano-curcumin demonstrate reasonable outcomes, equivalent to MTA. This suggests that natural materials could be viable alternatives to synthetic ones. Limitations of this study include the small sample size, potential bias due to the absence of long-term follow-up, and the exclusion of irreversible pulpitis cases, which may limit generalisability. Additionally, the subjective nature of pain assessments using the VAS scale may have introduced variability. Furthermore, while the study controlled for immediate post-treatment responses, long-term outcomes such as pulp regeneration or failure rates require further investigation.

Authors’ contributions

All Authors contributed equally. All authors of this manuscript approved of this publication.

Funding

None.

Data availability

Available upon request.

Declarations

Ethical approval

This research was authorized by the Research Ethics Committee (REC) of the Faculty of Dental Medicine, 6 October University, code number RECO6U/8–2023.

Clinical trial number

This clinical trial was registered online onto ClinicalTrials.gov. The registry identification number was NCT06029023. 

Consent for publication

Obtained.

Consent to participate

Obtained.

Competing interests

All of the authors declare that they have no competing interests.