Comparative evaluation of pH, and calcium and fluoride ion release between two pulp capping agents at different time periods: an in vitro study

Introduction

Vital pulp therapy (VPT) aims to preserve and safeguard healthy pulp tissue, which has been compromised but not yet destroyed due to factors, such as caries, trauma, or restorative procedures [1]. Pulp capping is the use of a biocompatible substance at the base of cavity to stimulate the formation of hard tissue. Capping material affects the prognosis of remaining pulp tissue, along with health status and healing capacity of pulp [1, 2].
Remaining dentine thickness (RDT) influences pulp vitality. According to Fusayama’s research, an inner layer is impacted by bacteria toxins and an outer layer is dentin infected by bacteria [3, 4]. The inner layer should be retained during clinical therapy, since it is physiologically mineralizable, uninfected, and partially demineralized. Consequently, bonding substrate in clini­cal scenario is typically made of affected dentin ra­ther than normal one. Caries-affected dentin (CAD) has less amount of bacterial contamination when compared with caries-infected dentin, and can be remineralized as collagen retains the regular crossband infrastructure. However, it is essential for the restorative material to create the right microenvironment for remineralization by providing a strong interface with dentin [5]. Therefore, a pulp capping restorative material should possesses ideal qualities, such as antibacterial action, bonding efficiency, resistance to microleakage, inducing hard tissue formation, and biocompatibility. Along with these advantages, the property of releasing calcium and fluoride ions is necessary for the remineralization of dentin and enamel [6].
Calcium silicate-based cements are commonly used in clinical settings, such as endodontic treatments and VPT. The effectiveness of these materials largely stems from their bioactivity, characterized by their capability to leach out calcium ions (Ca2+), and generate crystalline deposits similar to apatite upon exposure to phosphate-containing physiological fluids [7, 8]. Indeed, the ionic byproducts of calcium silicates were shown to promote osteogenesis and angiogenesis [9].
Secondary caries is the primary reason for unsuccessful treatment of coronal restoration. Therefore, averting secondary caries is essential in preventing tooth loss, and fluoride plays a significant part in this process [10, 11]. Fluoride ions also stimulate remineralization and suppress oral micro-organisms. The presence of fluoride ions can increase the acid resistance of cavity margin, thus playing a role in integrity of cavity walls [12].
pH of a material has an effect on the stimulation of repair by deposition of mineralized tissue [13]. pH cycling within dental plaque influences the release of fluoride ions from nearby restorations, exerting a significant impact on the fluoride release pattern of these restorations. Plaque pH values above 6 are thought to be safe, 6-5.5 is possibly cariogenic, and 5.5-4 is cariogenic. This pH cycling can be beneficial, since the restorative material is able to “smartly” release more fluoride when pH drops to the cariogenic region [14, 15]. An alkaline pH encourages the healing of dental tissues and remine­ralization [13].
Two of such pulp capping base materials have recently been introduced to the market: TheraBase (BISCO) and Biner LC (Meta Biomed). TheraBase is a dual-cure self-adhesive base/ liner that releases calcium and fluo­ride while offering radiopacity. It bonds to tooth substrate chemically, and continually releases calcium as well as fluoride ions. It also has a property of rechargeability of these ions. Its calcium ion leaching creates a pH alkaline enough to support pulp vitality [16]. Biner LC is a light-curing cavity liner and base material that releases calcium and fluoride, providing radiopacity, and has been specifically designed to be used with adhesives, composites, and traditional restorative materials [1].

Objectives

The aim of the study was to evaluate and compare the ion release and pH of TheraBase and Biner LC over different time periods. The null hypothesis is that no difference in ion release and pH will be observed at all time intervals between the two materials.

Material and methods

Sample preparation

Cylindrical samples of 6 mm height and 3 mm dia­meter were prepared in moulds. Material was injected into the mould and cured from both sides according to the manufacturer’s instructions for each material. Digital balance was used to weigh the samples, and digital micrometer was employed to measure dimensions to ensure standardization. The samples were suspended in 10 ml deionized water by incorporating dental floss during setting. Then, they were stored at 37°C and 100% relative humidity for 2 hours, 24 hours, 48 hours, 7 days, and 21 days. Solution used for storage was collected and analyzed for measurement of calcium and fluoride ions. Fresh 10 ml distilled deionized water in a new container was used to suspend the samples after drying, and storage was continued till the next time periods.
EDTA titration method was applied to check calcium ion release. EDTA is a chelating agent that can bind to metal ions, such as calcium (Ca2+). By using EDTA in a titration, the amount of EDTA required to bind all calcium ions in a solution can be measured. The endpoint is typically detected with a suitable indicator, which changes color when all calcium ions have reacted with EDTA.
SPADNS spectrophotometric method was used to check fluoride ion release. SPADNS dye creates a colored complex with zirconium ions, typically a reddish-pink color. When fluoride ions are introduced into the solution, they react with zirconium, forming a stable, colorless zirconium-fluoride complex. This reaction reduces the intensity of pink color proportionally to the fluoride ion concentration. The change of color is measured spectrophotometrically, typically at a wavelength of 570 nm.
A total sample size of 30 was considered and calculated using a sample size formula. The samples were divided into 2 groups, which were further divided into 3 subgroups – group 1: TheraBase (n = 15), group 2: Biner LC (n = 15), subgroup 1: Calcium release (n = 5), subgroup 2: Fluoride release (n = 5), subgroup 3: pH (n = 5).
Each subgroup parameter was checked at the following time periods: 2 hours, 24 hours, 48 hours, 7 days, and 21 days.

Statistical analysis

Continuous variable, mean and standard deviation (SD), was obtained. For intragroup comparison at diffe­rent time intervals, ANOVA and Tukey’s post hoc tests were applied. For intergroup comparison, unpaired t-test was used. All statistical tests were performed with 95% CI, while p < 0.05 was considered statistically significant.

Results

The calcium ion release in group 1 was consistently higher than in group 2 at all time periods, and increased gradually over time with the highest release at 21 days interval. Group 2 showed a consistent release at all time intervals. When comparison of calcium ion release was done between group 1 and group 2, it was observed that there was a statistically significant difference in the mean of calcium ion release values (Table 1).
There was a statistically significant change in the fluoride concentration in both group 1 and group 2. Group 1 showed a significantly higher release as compared to group 2 at all intervals. Both groups demonstrated the highest release at 24 hours (Table 2).
The comparison of pH between group 1 and group 2 revealed that there was a statistically significant diffe­rence in the mean of pH values. Group 1 had a higher alkaline mean pH of 10.6, while group 2 showed a more acidic mean pH of 6.3 (Table 3).

Discussion

The efficacy of a filling material capable of delivering calcium and fluoride ions can be measured by its capa­city to release these ions to the neighboring tooth tissue, rather than raising their concentration in saliva. Specifically, the ions must be situated near the tooth’s surface in order to have the greatest impact [6]. Restorative materials, such as glass ionomer cement (GIC) and its derivatives as well as ion-releasing resin composites, possess the ca­pa­city to leach out “therapeutic” ions, but have shown transient failures in preventing secondary caries, primarily as a result of bonding failures and gap formation [5, 17]. In one of the studies, it was observed that the presence of resin decreased the ionic activity [18].
TheraBase is a methacrylate-based dual cure cement composed mainly of Portland cement and barium glass for radiopacity [15]. Biner LC is majorly composed of hydroxy calcium phosphate suspended in urethane dimethacrylate resin [1]. All the parameters were studied over different time periods to check the immediate release at 2 hours, 24 hours, and 48 hours, while the sustained release and material’s longevity were evaluated at 7 days and 21 days. The results showed that TheraBase presented a significantly greater amount of calcium and fluoride release than Biner LC. Moreover, calcium release in TheraBase was seen to gradually increase with time. This was probably due to the utilization of patented (THERA) technology of TheraBase, which allows it to chemically bond to the tooth structure by releasing and recharging the calcium and fluoride ions. It has hydrophilic matrix, thus resulting in an easy exchange of ions. As water goes into the matrix, it reacts, resulting in the release of calcium hydroxide and fluoride ions [15]. The release of calcium ions (Ca2+) and hydro­xyl ions (OH–) from calcium hydroxide holds significant importance when considering biointeractive properties. The release of calcium ions is vital for the process of mineralization due to the promotion of cell migration and differentiation. Furthermore, hydroxyl ions contribute to a high pH level (approximately 12), facilitating enzymatic inhibition of microorganisms. Additionally, hydroxyl ions release alkaline phosphatase, which plays a role in the mineralization process [19].
Portland cement in TheraBase creates calcium silicate cement (CSCs), while the source of calcium in Biner LC is hydroxyapatite. In the hydration reaction in CSCs, leaching out of calcium hydroxide and calcium silicate hydrate gel from these particles occurs. This play a crucial role in the process of healing of the pulp-dentin complex [20, 21]. One probable explanation for the elevated Ca2+ emissions from calcium silicate cements might be linked to the processes involved in setting and hydration [8].
TheraBase resulted in having a fairly alkaline pH, noted at all time periods, while Biner LC showed acidic pH at almost all time intervals. The hydration reactions in CSCs create a hard tissue, inducing microenvironment for an extended period after setting through the generation of an alkaline pH. Additionally, calcium ions play a crucial part in signalling cascades, which take place intracellularly. According to a research on CSC-related mediators of signalling, extracellular receptors of calcium sensing (CaSRs) serve as crucial mediators in the CSC-induced pulp-dentin complex regeneration. CSC triggers various downstream pathways of CaSR, and this cascade mechanism is associated with the regulation of calcium ions and pH levels [22]. Apatite nucleation is accelerated at alkaline pH values, causing the solubility of OH ions to increase and become incorporated into an integral component of the tooth structure [9].
It was observed that TheraBase had a higher fluoride release when compared with Biner LC at all time periods. However, the fluoride release was seen to be constant in both the materials at all interval points. Each material releases fluoride at a different rate due to differences in solubility [6]. The presence of ytterbium fluoride in TheraBase becomes its source of fluoride release. The amount of fluoride that reaches the oral cavity depends more on how well the material leaches fluoride, or can exchange fluoride for other ions in the oral envi­ronment than it does in the fluoride concentration of the substance [23].
As both of the study’s materials continued to release fluoride throughout the testing process at all time pe­riods, they may be helpful in preventing deminera­lization and enhancing remineralization. The release of fluo­ride ions from dental materials has been demonstrated to fluctuate significantly based on the storage medium utilized [24]. It was noted that fluoride released from various restorative materials was increased under severe acidic conditions below pH 4.0. This implies that if an acidic medium was used instead of deionized water in this study, more fluoride might be released. Additionally, it suggests that fluoride release would be increased during bacterial acidogenic attacks, aiding in the prevention of demineralization. Therefore, further investigations are warranted to assess the characteristics of these materials in various storage media, replicating oral cavity conditions [12]. The present study investigated only the release of the ions; further studies are needed to check the rechargeability of the materials.

Conclusions

TheraBase demonstrates superior ion release and a more favorable pH profile compared with Biner LC, making it a promising candidate for bioactive pulp capping in restorative dentistry. Its sustained release of calcium and fluoride ions suggests possible provision of long-term the­rapeutic benefits, positioning it as a viable alternative to traditional materials for pulp capping applications.

Disclosures

1. Institutional review board statement: Not applicable.
2. Assistance with the article: None.
3. Financial support and sponsorship: None.
4. Conflicts of interest: None.

References