Tooth decay is a significant public health problem worldwide and a significant cause of tooth destruction, dental care needs, and health care costs.[1] Oral diseases affect about 3.5 billion people around the world, or nearly 45% of the world's population.[2] More than 2 billion people have permanent dentition and suffer from untreated dental caries, while about 514 million children have primary dentition and suffer from untreated dental caries.[3] Although preventive and restorative dentistry have progressed, the long-term survival of dental restorations is still difficult due to secondary caries, material degradation, and exposure to changing oral conditions.[4] These challenges have led to the development of restorative materials that not only are mechanically durable but also actively interact with the surrounding dental tissues to reduce demineralization and enhance demineralization.

Bioactive restorative materials have received a great deal of interest due to their capacity to release therapeutic ions, such as fluoride, calcium, and phosphate, to the surrounding environment.[5] These ions can help to remineralize, inhibit demineralization, and maintain the mineral balance at the tooth-restoration interface.[6] The latest generation of bioactive materials, such as glass ionomer cements (GIC), resin-modified glass ionomers (RMG), glass hybrids, giomers, alkasites, and bioactive resin-based restoratives (RBR), have varying chemical compositions and mechanisms of ion release.[7] Tuygunov et al. (2024) emphasized that calcium- and phosphate-releasing glass ionomer formulations possess demineralization potential, while Aliberti et al. (2025) showed that environmental conditions are a major determinant of ion release from restorative materials.[8, 9] In their investigation, fluoride release differed significantly across pH conditions and exposure periods, emphasizing the importance of evaluating bioactive behaviour under clinically relevant conditions.

The oral cavity experiences frequent shifts in pH caused by bacterial acid production, food consumption, and buffering by saliva.[10] Acidic environments can help to break down the reactive fillers and facilitate the release of ions, which can lead to increased demineralizing ion availability and impact material integrity.[11] Under acidic conditions (pH 4.8) the fluoride concentrations reported by Aliberti et al. (2025) were from 1.61 to 7.67 mg/L; for near-neutral conditions (pH 6.8), the range was 1.41 to 5.33 mg/L, but the release of fluoride varied for individual materials.[9] In a similar fashion, Maaly et al. (2025) showed that there were significant differences in the release of fluoride and calcium between Surefil One, Equia Forte HT Fil, and ACTIVA BioACTIVE, and associated differences in surface microhardness.[12] The results indicate that the ion-releasing capacity is dependent on material and that the amount of ions released is not necessarily correlated with superior overall restorative performance.

The use of bioactive restorative materials is ever-growing and gaining momentum in relation to their therapeutic properties, but attaining an optimal balance between ion release and mechanical strength is a significant challenge. The mechanical integrity of the material is an important variable in the oral environment, where repeated chemical challenges are present, and testing only ion release can give an incomplete assessment of the suitability of the material in the clinical setting. It is therefore important to make a direct comparison under controlled pH to see if enhanced ion release is also correlated with either improved or reduced surface hardness.

This study was set out to comparatively investigate the ion-release capacity and surface microhardness of selected bioactive restorative materials in acidic and neutral conditions and to assess the effect of pH on the physicochemical performance of the bioactive restorative materials. The findings could serve as important evidence in choosing restorative materials with both bioactive properties and acceptable mechanical properties, especially in patients who are subjected to frequent acidic challenges.

An in vitro comparative experimental study was conducted at Sharif Medical and Dental College, Lahore, Pakistan. Specimen preparation, immersion procedures, and laboratory testing were performed with the facilities and analytical equipment of the dental materials laboratory. The study period lasted for six months: from 1st December 2025 to 30th May 2026.

The sample size was calculated based on the approach used in the OpenEpi sample size calculator that compares two independent means, as described in the experimental design of Maaly et al. (2025), which assessed ion release and microhardness of three bioactive restorative materials.[12] They studied 90 specimens, divided into three groups of restorative materials with 10 specimens in each group and in each storage-time group. A 95% confidence level, 80% statistical power, equal allocation between acidic and neutral conditions, and an assumed standardized effect size of 0.80 were assumed for the present calculation. These assumptions yielded an estimated minimum of 26 specimens per pH condition for each restorative material. The sample size was expanded to 30 specimens per condition to include potential specimen damage and laboratory testing inaccuracies. Thus, if three materials had been studied, 180 specimens would have been needed, 60 for each material, split equally between the acidic and neutral conditions.

The bioactive properties and suitability for laboratory investigation guided the use of a non-probability purposive sampling technique to select restorative materials. All specimens were prepared with standardized dimensions and randomly distributed among the experimental conditions of acidic or neutral. An equivalent number of specimens was kept in each experimental group to allow comparability and to reduce allocation bias.

Bioactive restorative materials that are commercially available and have proven ion-releasing properties were deemed appropriate for use. Samples with uniform size, smooth surfaces and no visible cracks, voids or structural defects, and complete setting or polymerization were included and were prepared according to the manufacturers' recommendations. Specimens that failed to hold up in preparation and baseline examination were not deemed suitable for experimental testing.

Materials used in restorative practices that do not have the ion-releasing characteristics relevant to the study were not considered conventional restorative materials. Other specimens with incomplete polymerization, irregular dimensions, cracks on the surface, porosity, contamination, or manufacturing defects were also removed. Specimens that cracked, were contaminated, or damaged during preparation or handling were rejected for further assessment.

The methodology used was adapted from Maaly et al. (2025).[12] Three bioactive restorative materials were chosen, and the standardized cylindrical specimen with a diameter of 10 mm and thickness of 3 mm was prepared using the Teflon mould. The materials were manipulated and placed in the moulds and then set or polymerized as directed by each manufacturer's instructions. Mylar strips and glass slides were used to obtain uniform, smooth surfaces and minimize the formation of air bubbles. The specimens were then inspected for structural flaws and placed in storage at 37°C prior to baseline testing. The two experimental subgroups of each restorative material were based on the immersion medium. The acidic subgroup was dipped in a buffered solution at pH 4.5 and the neutral subgroup in a solution at pH 7.0. All specimens were individually placed in a labeled container with 5 mL of the storage solution and incubated at 37°C for the duration of the experiment.

The time of immersion was 7, 14, and 21 days, and the release of ions was measured. The storage solutions were taken at different intervals and stored in labelled containers, and new solutions of the same pH were supplied throughout the experiment to ensure consistency. Fluoride and calcium ion concentrations of the collected solutions were analyzed by ion chromatography using suitable calibration standards. The concentrations of ions were reported in mg/L, and cumulative ion release was computed as the sum of the amounts of ions collected in the various collection periods.

The Vickers microhardness testing machine was used to measure surface microhardness at baseline and after the specified immersion periods. Each specimen was indented at three different points with a load of 300 g for 15 s. Vickers hardness number (VHN) was taken as the average of the three readings. The change in microhardness from baseline was calculated to determine the effect of acidic and neutral conditions on material stability. Observations were recorded on a data collection sheet, which was structured so that it could be analyzed statistically.

Data collected were analysed in IBM SPSS Statistics. The fluoride release, calcium release, and surface microhardness were quantitative variables and expressed as mean and standard deviation. The statistical testing method for the distribution of numerical variables was selected as the Shapiro–Wilk test. A two-factor factorial analysis of variance (ANOVA) was used to determine differences in ion release and microhardness of the different restorative materials and environmental pH conditions. The linear mixed-effects model was used to evaluate the changes over time and interactions among the restorative material, the pH, and the immersion time when repetition was used on the same specimens. Post hoc pairwise comparisons were conducted with appropriate multiple comparison correction. The relationship between the cumulative ion release and changes in the surface microhardness was investigated using Pearson's correlation coefficient when the assumptions of parametric correlation were met. A p-value less than 0.05 was regarded as statistically significant.

A total of 180 specimens were included in the study, comprising three bioactive restorative materials: glass hybrid, alkasite, and giomer. The 60 specimens collected from each of the materials were split equally between acidic (pH 4.5) and neutral (pH 7.0) and included 30 specimens in each of the subgroups (Table 1).

The release of fluoride ions was also found to be cumulative, with the amount released rising consistently with time from day 7 to day 21 for all the restorative materials, irrespective of the pH levels. The release of fluoride was greater in the acid environment than in the neutral environment. The highest fluoride release was found in glass hybrid, followed by alkasite, while giomer showed the lowest release during the observation period (Table 2).

The release of calcium ions was also found to be time-dependent and occurred more in acidic conditions than in neutral conditions. Alkasite showed the highest cumulative calcium release, followed by glass hybrid and giomer. There were differences in the release rates of the three materials at each of the assessment times (Table 3).

The 21-day immersion period in all the test groups showed a progressive decrease in surface microhardness. The reductions were higher in acidic conditions, especially in the glass hybrid specimens. The surface microhardness of Alkasite was the highest, and the percentage reduction was the lowest, which shows comparatively higher hardness deterioration resistance to the tested conditions (Table 4).

The effects of restorative material and environmental pH on the fluoride release, calcium release, and percentage change in surface microhardness were found to be statistically significant based on 2-way ANOVA (all p<0.001). There were also significant interactions between material type and pH for all three outcomes, which means that the size of the pH-related effect was dependent on the type of material in each case (Table 5).

The results of Pearson correlation analysis indicated a moderate negative correlation between the cumulative fluoride release and percentage change in microhardness (r = −0.399, p<0.001). The release of calcium ions exhibited a weak, non-significant correlation with the change in microhardness (r = −0.081; p = 0.280) based on the pooled data, indicating that there was no clear linear relationship between the two in the overall data. (Table 6)

Table 1. Distribution of specimens by restorative material and immersion condition

Table 2. Cumulative fluoride ion release (mg/L) over time by material and pH

Table 3. Cumulative calcium ion release (mg/L) over time by material and pH

Table 4.  Vickers surface microhardness (VHN) and percentage change

Table 5. Two-way ANOVA of day-21 outcomes by material and pH

Table 6. Correlation between day-21 cumulative ion release and microhardness change

The present results showed that release of ions and surface microhardness for the bioactive restorative materials differed significantly under acidic and neutral conditions. The release of fluoride was highest for glass hybrid, and calcium release was highest for alkasite, with good microhardness retention. These findings were similar to those of Di Lauro et al. (2023), who observed fluoride and calcium release from glass ionomer and alkasite restoration materials in various pH levels.[13] In a similar study, Kashiyani et al. (2024) found that fluoride was released more from acidic than neutral media, while conventional glass ionomer released more fluoride than Cention N.[14] Aliberti et al. (2025) showed that pH, temperature, and immersion duration significantly affected the release of ions from the restorative materials, with different directions and magnitudes of fluoride release depending on the formulation.[15] These comparisons help to explain the need for studying the environmental conditions that affect the evaluation of ion-releasing materials.

The cumulative concentrations of fluoride and calcium release gradually increased throughout the experimental period and were higher under acid conditions. A similar study was conducted by Banic Vidal et al. (2023); fluoride release from the modern-day restorative materials they studied was found to be significantly different, with EQUIA Forte HT showing the maximum fluoride release.[16] Llancari-Alonzo et al. (2024) found that there were also notable differences between the amounts of fluoride released from various restorative materials and different periods of observation, with Cention N exhibiting comparatively high fluoride release in early time periods and Ketac Universal releasing comparatively high fluoride release in later time periods.[17] In addition, Aliberti et al. (2025) found that under acidic conditions there was an increase in fluoride release, which was especially seen for Riva Light Cure HV in a separate study that included paediatric materials.[9] All of these studies show that the release of ions is affected by the composition of the material, the length of the immersion time, and the pH of the surrounding environment.

The surface microhardness decreased after immersion, especially in the acidic solution, and the hardness of alkasite was comparatively better. Birant and Gümüştaş (2024) reported that the Vickers microhardness of Cention N with a no thermal aging group was significantly higher than all other bioactive restorative materials.[18] Likewise, Shetty et al. (2026) reported that there were differences among the materials in mechanical aging, in which Cention N showed a comparatively low degree of mechanical aging throughout the storage period.[19] Maaly et al. (2025), however, found that the Surefil one and Equia Forte HT Fil had higher microhardness values after water storage, while ACTIVA BioACTIVE had lower.[12] This difference from the present findings may be related to variations in restorative formulations, storage media, and experimental conditions.

The statistical analysis indicated significant effects of material type and pH on ion release and microhardness. The microhardness change was negatively correlated with fluoride release, and the calcium release was not statistically correlated. However, Maaly et al., 2025, showed a positive correlation between fluoride release and hardness of the tested materials and a negative correlation between calcium release and hardness of the tested materials, implying that the associations can be different for the various materials tested.[12] The sensitivity of the restorative performance to environmental conditions was also shown by Kim (2022), who reported that the bonding behaviour and microleakage of ion-releasing restoratives were influenced by environmental pH.[20] However, the relationships between the ion release and mechanical wear observed in the laboratory should not be taken as proof of a cause-and-effect relationship.

Overall, the results indicated that there was no single bioactive material that performed superiorly in relation to the different outcomes and that there was variation in the release of ions and ability to soften the surface. The same conclusions can be drawn from previous studies, which also show that the mechanical stability does not necessarily increase with an increase in ion release. Thus, the results highlight the need for a comparative study of bioactive potential and mechanical properties at various environmental conditions. But, the in vitro design, short immersion time, and lack of clinical parameters, including salivary dynamics, thermal changes, and mastication forces, make direct clinical interpretation limited. Additional experimental research with verified measurements and extended observation periods is needed to determine clinical significance of these relationships.

LIMITATIONS

The present study had certain limitations. It did not completely match the in vivo conditions that the oral cavity would experience, such as saliva flow, bacterial activity, food variations, and masticatory forces. The study was limited to only three bioactive restorative materials, as well as only 21 days of immersion time, which reduced the time for which the materials could be evaluated in terms of long-term material performance. Moreover, the continuous acidic, neutral pH did not mimic the acidic, neutral pH changes in the oral cavity. Other clinically relevant properties such as wear resistance, bond strength, and marginal integrity were not studied. Thus, more long-term experimental and clinical studies are needed to determine the clinical significance of these observations.

Bioactive restorative materials showed differences in surface microhardness and ion release at various pH levels when used within the limitations of this study. High release of fluoride and calcium was related to acidic conditions while higher reduction in microhardness was also observed at that condition. The release of fluoride was the highest with glass hybrid, while the release of calcium was higher and the microhardness was retained better in alkasite. These results indicated that the material composition and the environmental pH affected the restorative performance. Therefore, the selection of bioactive restorative materials should take into account the ion releasing ability and mechanical stability.

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