Fixed appliance orthodontics is one of the most often employed methods for correcting malocclusion and enhancing dental function and aesthetics.[1] In the world, it is estimated that a significant amount of the population has malocclusion, which is the irregularity in the alignment of the teeth.[2]  Around 56% to 93% of the population has some kind of malocclusion that needs to be assessed or treated with orthodontics.[3] With the growing popularity of orthodontic therapy, bonded brackets are now utilized for most treatments in which adhesive systems are critical during treatment to help maintain bracket stability, along with reducing the damage to the underlying enamel.[4, 5]

The success of fixed orthodontic treatment is dependent upon the effectiveness of fixed orthodontic bracket bonding materials.[4] Traditional orthodontic cements are mostly resin-based composites, bond well, and are clinically durable, with some drawbacks such as polymerization shrinkage, marginal gaps, and plaque formation around the interface of the brackets and the tooth.[6, 7] These factors lead to microleakage, the penetration of oral fluids, bacteria, and acids between the adhesive-enamel interface, resulting in enamel demineralization and the formation of white spot lesions (WSLs).[8] Severe adverse effects of fixed orthodontic therapy include white spot lesions, which have been reported to occur in about 30% to 70% of orthodontic patients, depending on the length of treatment, oral hygiene habits, and diagnostic techniques employed.[9]

The development of demineralization around orthodontic brackets can be attributed to a combination of factors, including the accumulation of plaque and the shift in the oral microbial population, as well as the extended period of time during which the teeth are exposed to the acidic challenges.[10] The bracket margins are important, and where mineral loss is often seen because of the inability to maintain good oral hygiene.[11] This interface may cause microleakage, which affects the longevity of orthodontic bonding and can provide a good habitat for microbacterial growth and acid diffusion.[12]

Therefore, bioactive adhesives are an emerging and promising alternative to conventional bonding systems because they interact with the surrounding oral environment and can potentially remineralize the tooth enamel.[13] These materials may include calcium phosphate, fluoride-releasing agents, or fillers made of glass, which allow ions to exchange and minerals to deposit.[13] Bioactive adhesives are designed not only to adhere but also to prevent enamel mineral loss, whereas conventional adhesive materials are only mechanical in nature.[13]

Although bonding materials in the field of orthodontics have improved, the ideal bonding would not only ensure adequate retention of brackets, but also maintain the integrity of the enamel during the course of treatment without causing enamel demineralization. It is important to assess the ability of the bioactive adhesives to maintain the microleakage resistance and enamel mineral loss resistance after ageing to determine their clinical relevance and the potential of reducing orthodontic complications. Hence, an in vitro comparative study was done to evaluate microleakage and enamel demineralization around orthodontic brackets bonded with conventional and bioactive adhesives after artificial aging. The purpose of this study was to compare the performance of these different adhesive systems and to see if there is any difference in their ability to provide protection against enamel damage without compromising bracket bonding, and to evaluate whether bioactive adhesives could be superior in these aspects.

The present study was designed as an in vitro comparative experimental study to evaluate the microleakage and enamel demineralization around orthodontic brackets bonded with conventional and bioactive adhesive systems after artificial aging. The study was conducted for 6 months from July to December, 2025 in the Department of Dental Materials. Study samples were extracted human premolar teeth, and orthodontic brackets were bonded with various adhesive systems. Artificial aging tests were conducted to simulate intraoral conditions, and microleakage and demineralization of the enamel around the bracket–adhesive interface were assessed. The sample size was determined using the OpenEpi sample size calculator with the following parameters: 95% confidence interval with 8% margin of error. The sample size estimation was determined by the prevalence of enamel demineralization/white spot lesions found in one previous study conducted by Boersma et al., who reported about 50% prevalence of enamel demineralization with fixed orthodontic appliance therapy. The minimum sample size obtained by using the formula for estimating a single population proportion was 150 orthodontic bracket specimens. The samples were also split into two groups: Group A teeth were bonded with conventional orthodontic adhesive, and Group B teeth were bonded with the bioactive adhesive. The study specimens were selected using a non-probability purposive sampling technique. Human permanent premolars extracted were collected and processed in accordance with the predetermined inclusion and exclusion criteria. Selected teeth were then cleaned of soft tissue and calculus, disinfected in the same way and then kept under standard conditions for further processing. The samples were randomly divided into two experimental groups based on the orthodontic adhesive type. Randomization was used to divide the specimens into 2 experimental groups based on the type of orthodontic adhesive used. The teeth in group A were bonded with a conventional orthodontic adhesive, while the teeth in group B were bonded with a bioactive adhesive. All samples were bonded in the same way and were artificially aged before evaluation of microleakage and enamel demineralization. Extracted human permanent premolars with intact buccal enamel surfaces, without any caries, cracks, fracture, restoration, enamel defects, and previous orthodontic treatment were included in the study. Normal anatomical form and enamel structure of the teeth suitable for bracket bonding were selected. To ensure uniformity of samples, only freshly extracted teeth with no apparent pathological alteration were selected. Preexisting enamel demineralization, white spot lesions, carious lesions, developmental enamel defects, cracks, fractures, and restoration on the bonding surface were not included in the study. Specimens were also excluded if they had abnormal morphology, exposure to bleaching agents previously, or if damage occurred during preparation or handling and shown. Specimens were also excluded if they had abnormal morphology, had been exposed to bleaching agents previously, or had experienced damage during preparation and handling and were shown. Teeth that could not be stored in a standardized manner or did not meet the requirements of the study protocol were excluded from analysis. The collection of data was carried out by following a standard laboratory protocol. Human premolars were extracted, cleaned, disinfected, and stored in a suitable medium until prepared. All the samples were cut into acrylic resin blocks, and buccal surfaces were polished to get a uniform enamel surface. Two different adhesive systems were used to bond the orthodontic brackets on the prepared enamel surface following the manufacturer's instructions. The specimens were divided into two groups: teeth bonded with conventional orthodontic adhesive and teeth bonded with bioactive adhesive. All specimens were bracket-bonded and then artificially aged by thermocycling to simulate the intraoral temperature fluctuation and long-term clinical environment. After aging, the microleakage was evaluated with the dye penetration technique. The samples were stained in staining solution and then cut into sections and analysed under a stereomicroscope to assess dye penetration depth at the enamel–adhesive interface. A predetermined system was used to determine the score for microleakage. Artificially aged specimens were evaluated according to the standardized methods for enamel demineralization around orthodontic brackets. The extent and number of mineral loss and lesion formation surrounding the bracket margins were evaluated and documented for each specimen. To minimize observer error, all measurements were made by a calibrated examiner, and the data that were collected were documented on a structured data collection sheet for statistical analysis. The data collected were analyzed by applying the software known as Statistical Package for Social Sciences (SPSS) version 26. Microleakage data were presented as mean ± SD, and enamel demineralization data were presented as mean. A categorical variable, such as microleakage scores, was given as frequencies and percentages. To determine the normality of data distribution, the Shapiro-Wilk test was used. An independent sample t-test was used for normally distributed continuous variables for comparison between the conventional and the bioactive adhesive groups, while the Mann–Whitney U test was used for non-normally distributed continuous variables or for ordinal microleakage scores. The chi-square test was used to assess the association between adhesive type and categorical outcome. A p-value of ≤0.05 was considered statistically significant.

The study included 150 specimens equally divided into two groups according to the adhesive system used. A total of 75 specimens were tested in each of the conventional and bioactive adhesive groups for a balanced evaluation of the microleakage and enamel demineralization results after artificial aging. (Table 1)

Artificial aging showed that specimens bonded with bioactive adhesive exhibited a significantly lower microleakage between the enamel and the adhesive. This difference was found to be statistically significant, supporting the sealing ability and resistance to interfacial leakage through the bioactive adhesive system. (Table 2)

Microleakage assessment revealed that conventional adhesive specimens had a higher incidence of more severe microleakage, while bioactive adhesive specimens had the majority showing no or minimal dye penetration. This difference indicated a greater integrity of the margins and less degradation of the adhesive interface in the bioactive group. (Table 3)

Measurement of enamel demineralization showed significantly less mineral loss around brackets bonded with bioactive adhesive. The bioactive group exhibited less severity of demineralization and less lesion development than conventional adhesive, suggesting a protective effect of the bioactive group with regard to the enamel surface during orthodontic treatment. (Table 4)

Conventional adhesive samples had higher percentages of moderate or severe enamel changes compared to bioactive adhesive samples, which had higher percentages of minimal or no demineralization. This difference in adhesive systems was statistically significant. (Table 5)

There was a strong positive correlation between microleakage and enamel demineralization, indicating that the greater the leakage of the interface between the adhesive and tooth structure, the more enamel demineralization occurred after aging. This finding indicated that controlling microleakage may play an important role in preventing enamel damage around orthodontic brackets. (Table 6)

The results of the overall comparison revealed the superiority of bioactive adhesive over the conventional orthodontic adhesive in terms of reduction of microleakage and enamel demineralization. These results indicated that, besides the mechanical retention, bioactive materials could provide other preventive benefits. (Table 7).

Table 1. Baseline Distribution of Study Specimens According to Adhesive System Used

Table 2. Comparative Evaluation of Microleakage Depth at Enamel–Adhesive Interface After Artificial Aging

Table 3. Severity-Based Distribution of Microleakage Scores Among Study Groups

Table 4. Comparison of Enamel Demineralization Around Orthodontic Brackets Following Artificial Aging

Table 5. Distribution of Enamel Demineralization Severity Between Conventional and Bioactive Adhesive Groups

Table 6. Relationship Between Microleakage and Enamel Demineralization After Aging

Table 7. Overall Performance Comparison of Adhesive Systems

The effect of conventional and bioactive orthodontic adhesive on microleakage and enamel demineralization after artificial aging was evaluated in the present in vitro study. The results indicated that the bioactive adhesive had significantly lower microleakage and enamel mineral loss than conventional adhesive when compared with specimens bonded with the bioactive adhesive. This means that if bioactive components are incorporated, this could be a way to enhance the sealing properties of orthodontic adhesives and offer further enamel protection during orthodontic therapy.

Our results on the reduction of microleakage with enhanced adhesive performance are in agreement with the results from the comparative evaluation of various orthodontic adhesive systems by Masarykova et al. (2023), which found that there was a wide range of microleakage after thermal cycling. They stressed that the composition of the adhesive and the quality of the interface between the adhesive and the bracket are significant factors in determining leakage patterns surrounding orthodontic brackets. As in the previous study, the present study showed that the selection of the adhesive is a very important factor in reducing fluid and bacterial penetration at the interface between enamel and adhesive.[14]

The lower enamel demineralization in the bioactive adhesive group was similar to the results reported by Ali et al. (2024), who assessed a bioactive composite material for orthodontic bracket bonding and found it to be more resistant to enamel demineralization than conventional materials. The main reason for the protection was the presence of active ingredients that are able to liberate beneficial ions and stimulate remineralization to the margins of brackets. The present results also confirm the notion that orthodontic bonding materials can be used not only as bonding agents, but also as preventive materials against the formation of white spot lesions.[15]

This marked decrease in enamel mineral loss was also confirmed by Yang et al. (2022), who examined a calcium silicate-based orthodontic adhesive and found that it neutralized and remineralized acids. They concluded that bioactive fillers may alter the local environment of orthodontic brackets by increasing mineral deposition and decreasing mineral dissolution in the enamel during the cariogenic challenge. This effect could be attributed to these mechanisms, which likely contributed to the reduced demineralization scores that we found with bioactive adhesive.[16]

Kamber et al (2021) conducted a systematic review and found that preventive bonding materials and modified adhesive systems might help to decrease the prevalence and severity of enamel demineralization throughout fixed orthodontic treatment. This evidence is confirmed by the present study, which showed that bioactive adhesive systems were more effective in preserving enamel than conventional resin-based adhesive systems. The review also highlighted the differences between materials, indicating that the clinical effectiveness can vary based on the composition of the adhesive used and the stability of the material over time.[17]

Our study found that there is a correlation between microleakage and enamel demineralization, which has been reported on in previous studies that indicated that bacterial accumulation, acid diffusion, and mineral loss occur beneath orthodontic brackets. The same study by Masarykova et al., (2023) concluded that microleakage at the enamel–adhesive interface can also lead to white spot lesions and secondary caries around orthodontic appliances. The high correlation found in the present study again emphasizes the need for maintaining an effective adhesive seal.[14]

Demircioglu et al. (2023) assessed the effect of the various adhesive systems on enamel demineralisation and concluded that the composition of the adhesive systems had a significant impact on the preservation of enamel mineral around orthodontic brackets. Their results confirm the current observation that modifications of adhesive materials with protective properties can help to prevent enamel degradation under cariogenic conditions. The differences among the studies may be attributed to different types of adhesives, demineralization methods, and assessment methods.[18]

The impact of various bracket adhesive systems on enamel demineralization was also examined by Hennig et al. (2023), who found that enamel changes following orthodontic bonding are influenced by material properties. Like the current results, they found that mechanical retention is the main advantage offered by conventional adhesives, while newer bioactive materials may offer other biological advantages.[19]

The present results corroborate those obtained by Khalifa et al. (2023), who observed that fluoride-containing orthodontic adhesive modifications provided better remineralizing efficiency and enamel protection. They found that the addition of mineral-releasing components can improve enamel resistance during orthodontics. This is in line with the better performance of the bioactive adhesive seen in our study.[20]

Abbassy et al (2021) investigated fluoride bioactive glass materials surrounding orthodontic brackets and their protective effect against enamel demineralization. They found that bioactive materials could serve as reservoirs for mineral ions, which would help to decrease the dissolution of the enamel and to promote remineralization.[21] The present study confirms these observations in the use of these bioactive materials directly as orthodontic bonding agents.

Recent evidence also indicates that other preventative methods, like remineralizing agents, can affect the results of bracket bonding. Domantaitė and Trakinienė (2023) showed that remineralizing interventions can alter the dentine conditions without compromising the bonding properties. The results of these studies were consistent with the present result, indicating that the proper future orthodontic material should have both mechanical bond strength and enamel-protective properties.[22]

The results of the present study are in line with the recent findings from 2021–2026, which have shown that the bioactive orthodontic adhesives have potential benefits over conventional adhesives in reducing microleakage and enamel demineralization. Fixed orthodontic therapy is primarily used for the treatment of orthodontic, facial, and dental deformities; while conventional adhesives are still clinically effective for brackets, bioactive systems could be a more preventive approach by tackling one of the most prevalent complications of fixed orthodontic therapy, namely, enamel white spot lesion formation. Additional clinical trials are needed to prove that these laboratory benefits result in better long-term orthodontic outcomes.

Limitations

Some limitations in this study need to be taken into account when interpreting the results. The experimental environment was not entirely representative of the complex oral environment, such as saliva flow, bacterial activity, dietary influences, and variations in oral hygiene. The artificial aging procedures were designed to mimic clinical conditions, but may not reflect long-term intraoral stresses during orthodontic treatment. In addition, the study assessed microleakage and demineralization of enamel with lab methods, and additional clinical research with extended follow-up periods is needed to confirm these results in actual orthodontic patients.

It was found that in the present study, bioactive orthodontic adhesive attains better performance than the conventional adhesive and reduces the microleakage and enamel demineralization around orthodontic brackets after artificial aging significantly. The enhanced protective effect of the bioactive adhesive indicates its ability to be used as a preventative bonding agent, in addition to its use for providing bracket adhesion, which can also support enamel preservation. The results presented here indicate that bioactive adhesives have the potential to mitigate orthodontic complications, such as white spot lesion formation, although more clinical studies are needed to confirm these results.

1. Liu, F., et al., Comparison of fixed braces and clear braces for malocclusion treatment. BMC Oral Health, 2024. 24(1): p. 941.
2. De Ridder, L., et al., Prevalence of orthodontic malocclusions in healthy children and adolescents: a systematic review. International journal of environmental research and public health, 2022. 19(12): p. 7446.
3. Lone, I., et al., Global Map of Skeletal and Dental Malocclusion Prevalence: From Classes to Continents. J Dentistry Oral Disorders, 2024. 10: p. 1183.
4. Wambier, L.-M. and D.-S. Wambier, Orthodontic bracket bonding techniques and adhesion failures: A systematic review and meta-analysis. Journal of Clinical and Experimental Dentistry, 2022. 14(9): p. e746.
5. Koaban, A.M., et al., Recent advances in orthodontic brackets: from aesthetics to smart technologies. Cureus, 2025. 17(6).
6. Heboyan, A., et al., Dental luting cements: an updated comprehensive review. Molecules, 2023. 28(4): p. 1619.
7. Liang, X., et al., Three-dimensional printing resin-based dental provisional crowns and bridges: recent progress in properties, applications, and perspectives. Materials, 2025. 18(10): p. 2202.
8. Alnazeh, A.A., et al., Photodynamic therapy and argon laser treatment of white spot lesions: Effects on enamel remineralization and orthodontic bracket bond strength. Journal of Photochemistry and Photobiology B: Biology, 2025: p. 113316.
9. Ludovichetti, F.S., et al., Prevention of White Spot Lesions Induced by Fixed Orthodontic Therapy: A Literature Review. Dentistry Journal, 2025. 13(3): p. 103.
10. Inchingolo, A.D., et al., Oralbiotica/oralbiotics: the impact of oral microbiota on dental health and demineralization: a systematic review of the literature. Children, 2022. 9(7): p. 1014.
11. Adali, N., D.S. Gill, and F.B. Naini, Care of Fixed Appliances. Preadjusted Edgewise Fixed Orthodontic Appliances: Principles and Practice, 2023: p. 235-246.
12. He, L., et al., Applications of nanotechnology in orthodontics: a comprehensive review of tooth movement, antibacterial properties, friction reduction, and corrosion resistance. BioMedical Engineering OnLine, 2024. 23(1): p. 72.
13. Nizami, M.Z.I., et al., Advances in bioactive dental adhesives for caries prevention: a state-of-the-art review. Journal of Functional Biomaterials, 2025. 16(11): p. 418.
14. Masarykova, N., et al., Comparison of microleakage under orthodontic brackets bonded with five different adhesive systems: in vitro study. BMC Oral Health, 2023. 23(1): p. 637.
15. Ali, N.A.M., et al., Enamel demineralization around orthodontic brackets bonded with new bioactive composite (in-vitro study). Journal of Baghdad College of Dentistry, 2024. 36(2): p. 54-62.
16. Yang, S.-Y., et al., Acid neutralizing and remineralizing orthodontic adhesive containing hydrated calcium silicate. Journal of Dentistry, 2022. 123: p. 104204.
17. Kamber, R., et al., Efficacy of sealants and bonding materials during fixed orthodontic treatment to prevent enamel demineralization: a systematic review and meta-analysis. Scientific reports, 2021. 11(1): p. 16556.
18. Demircioglu, R.M., et al., Do Different Types of Adhesive Agents Effect Enamel Demineralization for Orthodontic Bonding? An In Vitro Study. Coatings, 2023. 13(2): p. 401.
19. Hennig, C.-L., et al., Influence of Different Bracket Adhesive Systems on Enamel Demineralization—An In Vitro Study. Journal of Clinical Medicine, 2023. 12(13): p. 4494.
20. Khalifa, O.M., M.F. Badawi, and T.A. Soliman, Bonding durability and remineralizing efficiency of orthodontic adhesive containing titanium tetrafluoride: an invitro study. BMC Oral Health, 2023. 23(1): p. 340.
21. Abbassy, M.A., A.S. Bakry, and R. Hill, The efficiency of fluoride bioactive glasses in protecting enamel surrounding orthodontic bracket. BioMed Research International, 2021. 2021(1): p. 5544196.
22. Domantaitė, M. and G. Trakinienė, Influence of the use of remineralizing agents on the tensile bond strength of orthodontic brackets. Scientific Reports, 2023. 13(1): p. 507.