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Year : 2022  |  Volume : 25  |  Issue : 1  |  Page : 42-46
Effect of nonthermal atmospheric plasma, grape seed extract, and bromelain on immediate bonding of composite to bleached and microabraded surfaces

Department of Conservative Dentistry and Endodontics, Sibar Institute of Dental Sciences, Guntur, Andhra Pradesh, India

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Date of Submission19-Oct-2021
Date of Decision01-Dec-2021
Date of Acceptance03-Jan-2022
Date of Web Publication02-May-2022


Background: Tooth whitening procedures such as bleaching and microabrasion alters the enamel surface and thus reduce the composites' bond strength. Hence, various surface treatments were introduced to overcome this problem.
Aim: To assess the effect of nonthermal atmospheric plasma (NTAP), grape seed extract (GSE) and bromelain on the shear bond strength (SBS) of the composite after bleaching and microabrasion.
Materials and Methods: Eighty extracted maxillary anterior teeth were distributed into two groups. Group 1: bleaching and Group 2: Microabrasion and further subdivided into four subgroups (n = 10) based on the form of surface treatment employed. Group A: no surface treatment, Group B: bromelain, Group C: GSE, Group D: NTAP. Composite resin was bonded to the labial surface and placed in artificial saliva for 24 h. SBS testing was done for all the samples.
Statistical Analysis: Analysis was performed using two-way analysis of variance and post hoc Tukey's test with P < 0.05 considered statistically significant.
Results: Group 1D revealed the highest bond strength (35.4 Mpa) and Group 1A showed the least bond strength values (15.7 Mpa). Among the bleaching groups, significant difference was observed between all the subgroups except Group 1A and 1B (P = 0.972). In microabrasion group, significant difference was observed between Group 2A and 2D (P = 0.0001), Group 2B and 2D (P = 0.0010), and Group 2C and 2D (P = 0.0001).
Conclusions: Following bleaching, NTAP application and GSE significantly improved the SBS of composite resin. Application of NTAP to microabraded surfaces increased the bond strength of composite resin.

Keywords: Bleaching; bromelain; grape seed extract; microabrasion; nonthermal atmospheric plasma

How to cite this article:
Banu MA, Bolla N, Tammineedi S, Vemuri S, Basam RC, Ganapati AK. Effect of nonthermal atmospheric plasma, grape seed extract, and bromelain on immediate bonding of composite to bleached and microabraded surfaces. J Conserv Dent 2022;25:42-6

How to cite this URL:
Banu MA, Bolla N, Tammineedi S, Vemuri S, Basam RC, Ganapati AK. Effect of nonthermal atmospheric plasma, grape seed extract, and bromelain on immediate bonding of composite to bleached and microabraded surfaces. J Conserv Dent [serial online] 2022 [cited 2023 Oct 1];25:42-6. Available from:

   Introduction Top

Intrinsic enamel defects, such as opacities and brown pigmentations can adversely affect the color, translucency of enamel. Depending on the severity of the defects, various treatment modalities such as bleaching, microabrasion, veneers, and crowns are performed.[1] However, bleaching remains the mainstay of treatment for these intrinsic stains. The bleaching agent slowly interacts with stains located within the superficial layers of the tooth and thus lightens the tooth surface.[2] Hydrogen peroxide (H2O2) has side effects such as tooth sensitivity, external cervical resorption, and reduction of composites' bond strength when bonded immediately to the bleached surface. The standard method to overcome this problem is to delay the bonding for up to 4 weeks following bleaching.[3] A variety of methods have been proposed to overcome this problem, which includes superficial tooth reduction, alcohol application before bonding, using acetone-based adhesives, and antioxidants such as alpha-tocopherol and sodium ascorbate.[4]

Grape seed extract (GSE) is a natural antioxidant that contains 98% of oligomeric proanthocyanidin complexes (OPCs). OPCs contain electron donor sites, i.e., hydroxyl sites, which binds to the free radicals. They neutralize the free radicals trapped in the enamel following the degradation of H2O2. There is a need to compare the effect of applying these antioxidants to bleached surfaces with other surface treatment procedures.[4]

Espinosa et al. stated that Enamel deproteinization before etching increases the retentive surface of the enamel to 94.47%.[5] Deproteinizing enzymes like bromelain remove the unsupported collagen, thereby increasing the dentin permeability and enhancing the diffusion of monomers. Bromelain was reported to decrease nanoleakage following the removal of collagen.[6] However, there are no studies evaluating the efficacy of bromelain following bleaching on the shear bond strength (SBS) of composite to the tooth surface.

In recent times, plasma has gained considerable importance in the medical field. Plasma is a partially ionized gas and known as the fourth state of matter. Nonthermal atmospheric plasma (NTAP) is a low-temperature artificially generated gas, which does not use a vacuum system.[7] Various studies have reported the increase in composite resin's bond strength to the tooth surface using proteolytic enzymes, antioxidants, and NTAP. However, limited literature is available regarding the effect of bromelain, and NTAP on the bond strength of composites immediately following the tooth whitening systems.

Microabrasion is a simultaneous procedure of abrasion and erosion, reducing the superficial enamel resulting in a compact, prism-free layer. It comprises dense mineralized tissue in an organic matrix, which is replaced by a densely compacted prismless layer. Thus, by altering the enamel surface, it reduces the composites' bond strength.[8] Opinya et al. observed that removal of the compact mineralized layer can improve the composites' bond strength.[9] Hence, there is a need to study the effect of various surface treatments on microabraded surfaces. The present work aimed to assess the impact of bromelain, GSE, and NTAP on the SBS of composite resin following bleaching and microabrasion.

   Materials and Methods Top

After obtaining ethical clearance from the institutional ethics committee (Pr. 139/IEC/SIBAR/2018), eighty human maxillary anterior teeth (n = 80), extracted for various reasons, were selected for the study. The samples were thoroughly cleaned and were mounted in plastic molds with self-cure acrylic resin until the cementoenamel junction. The facial tooth surfaces were cleaned with pumice slurry. The samples were allocated into two groups based on the teeth whitening systems (n = 40 each).

Treating with tooth whitening systems

Group 1 (n = 40): the labial tooth surfaces were dried. Pola Office syringe (SDI limited, Australia) is taken, the plunger is pulled back to release pressure and extrude the contents into the pot supplied by the manufacturer. Immediately mixing is done using an applicator tip until a homogeneous mixture is obtained. Generous amount of this gel is applied on the labial surface for 8 min. The bleaching gel is then rinsed and blot dried.

In Group 2 (n = 40): a fine grit diamond bur with water coolant is used to slightly remove the labial surface superficially to initiate micro reduction. A plastic mac tip supplied by the manufacturer was attached to the Opalustre (Ultradent, USA) syringe. 1 mm thick layer is applied uniformly on the tooth's labial surface. A micromotor handpiece with Opal prophy cups was used at RPM of 400, along on the enamel surface for 120 s. Following this, the teeth were rinsed with distilled water.

The groups were subdivided into four subgroups: A, B, C, and D, respectively, based on the type of surface treatment performed.

Subgroup A (n = 10): No surface treatment is done (control group).

Subgroup B (n = 10): 5% bromelain solution for 10 min and then rinsed with distilled water.

Subgroup C (n = 10): 5% GSE solution for 10 min and then rinsed with distilled water.

Subgroup D (n = 10): NTAP exposed for 15 s.

Preparation of solutions

  1. Proteolytic enzyme– 5% bromelain solution-5 gm of bromelain (Meteoric Exim Pvt. Ltd., India) dissolved in 100 ml of distilled water
  2. Antioxidant– 5% GSE solution-5 g of GSE (All pure organics, India) dissolved in 100 ml of distilled water.

Nonthermal atmospheric plasma application

To generate the plasma jet, compressed He gas with a pulsing head of 2 GHz at 3.25 standard liters was utilized. The NTAP was applied at a pressure <2 Pascal's and a power of 2 Watt [Figure 1]. The application was carried out in an isolated closed chamber at a distance of 5–6 mm from the tooth surface for 15 s for each sample with an alternating cycle of activation and resting phase of 5 s each. The temperature on the tooth surface was also examined using a thermocouple which ranged from 27 to 29.4°C, that is endured by pulpal tissues.
Figure 1: Application of nonthermal atmospheric plasma

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Placement of composite restoration

For all the groups, the composite placement was carried out immediately after surface treatment. Self-etch adhesive (Solare Universal bond, GC, India) was applied to the labial surfaces of the tooth. A cylindrical plastic matrix of 3 mm diameter and 3 mm height was prepared. The matrix was placed on the labial surface and the composite resin (Spectrum, Dentsply, USA) was condensed incrementally. It was then photopolymerized in all directions using light-emitting diode light-curing unit (Orikam Eighteenth Curing Pen, India) for 60 s. The matrix was removed carefully following polymerization. The samples were then placed in artificial saliva for 24 h at 37°C. SBS testing was performed for all the samples using Universal Testing Machine (UTM) (Instron, Model 8801). Each sample was secured in UTM in such a way that the leading edge of the plunger is targeted at the interface between composite and labial surface at 1 mm/min speed. The force needed to dislodge the composite material was noted. The SBS was calculated using N/mm2 (peak load value/composite base area).

Statistical analysis

The data obtained were analyzed using version 20.0. SPSS software (Statistical Package for the Social science, IBM Corp. Ltd., Armonk, NY, USA). All the groups were analyzed for overall significance values using two-way analysis of variance test, whereas, comparison within the group and intergroup comparisons were made utilizing post hoc Tukey's tests with P < 0.05 was considered as statistically significant.

   Results Top

The results showed that Group 1D had higher bond strength (35.4 Mpa) out of all the groups, and Group 1A with least bond strength values (15.7 Mpa). In the bleaching groups, significant differences were observed between all the subgroups except Group 1A and 1B (P = 0.972) [Table 1]. In microabrasion group, significant differences were observed between Group 2A and 2D (P = 0.0001), Group 2B and 2D (P = 0.0010), and Group 2C and 2D (P = 0.0001) [Table 2].
Table 1: Comparison of interactions between the four subgroups (A, B, C, and D) with mean shear bond strength using Tukey's multiple post hoc test in Group 1

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Table 2: Comparison of interactions between the four subgroups (A, B, C, and D) with mean shear bond strength using Tukey's multiple post hoc test in Group 2

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   Discussion Top

The bleaching procedure might exhibit definite antagonistic effects on the enamel. These include a decrease in the surface microhardness, morphological as well as compositional changes.[10] These changes may affect the bonding of restorations to enamel when applied immediately postbleaching. According to the literature, tooth bleaching may reduce composites' bond strength applied to bleached enamel. Smith et al. stated significant surface structural changes in bleached enamel. According to them, fundamental changes may be the cause of the reduction in bonding strength.[11] Several reports have been reported regarding the association between bleaching agents and changes in the chemical or morphological structure of enamel.[12] These modifications include loss of calcium and regular prism margins, change in the ratio of the organic and inorganic constituents of the tooth.

In the present study, bleaching followed by immediate bonding of composite resin with no surface treatments showed the least bond strength. This finding is similar to the previous investigation by Nimet et al. who reported a drop in the mean bond strength value of composite by half due to the application of 35% H2O2.[13] Physical alterations in the tooth structure after bleaching were the probable reason for the decrease in enamel bond strength. Similar decrease in bond strength was reported by Cinthia et al., Khamverdi et al.[14],[15] this decrease in bond strength may be due to scanty, small, and poorly distinct resin tags bonded to bleached teeth. The eminence of these resin tags, their frequency, and their depth of penetration effect the bond strength of composite to the enamel, as mentioned in the research done by Titley et al.[11] Similar short, sparse, structurally incomplete, or complete absent resin tags in bleached enamel was reported by Dishman et al.[16]

Torneck et al. showed a substantial decrease in composite resin's adhesive bond strength following exposure of enamel to H2O2.[17] The residual peroxides on the enamel surface are responsible for the bond failure that happened at the resin-enamel interface. Rinsing leads to the residual H2O2 washout only on the superficial enamel (0.5–5 nm).[18] However, the residual free radicals trapped deep inside the enamel could have attributed to the compromised bond strength. Reversal of the reduced bond strength can be achieved in two ways. The first one is the time-dependent delay of composite placement, while the latter is by surface treatments performed on the bleached enamel using alcohol, acetone-based adhesives, and antioxidants.

In the current study, bromelain was used as an experimental surface treatment agent. Bromelain is commonly used in medical supplements. The pH of the bromelain ranges from 3 to 6.5. At this pH, the bromelain components are stable, and it disrupts adhered proteins causing stains. According to Aarti et al. there is a significant increase in microleakage after bleaching with 35% H2O2 and effectively reduced when the bleached teeth were treated with bromelain.[19] However, their methodology involved class V cavity preparation on the cervical third of the teeth. Better results for bromelain group in their study could be due to dentin involvement while cavity preparation, thereby exposing the collagen substrate for the bromelain to act. In the present study, bromelain showed no statistical significance when compared to the control group. This could be because the bromelain acts on the protein content, which is less in enamel.

To overcome the failure of immediate bonding to bleached enamel, Vidya et al. studied the effectiveness of antioxidants in the reversal of decreased adhesion to the bleached surface.[4] Nair et al. and Rose et al. studied that antioxidant agents have the potential to eliminate free radicals, thereby minimizing the interference with the polymerization of adhesives, which in turn can improve the bond strength of composites to bleached surface.[20]

GSE is a natural antioxidant that contains 98% of OPCs.[20] When compared to control, GSE significantly improved the bond strength. OPCs contain electron donor sites, i.e., hydroxyl sites, which binds to the free radicals. They neutralize the free radicals trapped in the enamel following the degradation of H2O2. This potential of OPCs is said to be 50 times superior than other antioxidants.[4],[6]

In the microabrasion group, control samples showed the least bond strength. Microabrasion increases the roughness of the enamel surface and is also related to the reduction in the enamel microhardness. The abrasion of enamel rods causes the formation of by-products which get compacted on the micro abraded surface. This results in a surface which is impervious to etching.[8] Group 2B showed no improvement in mean SBS values, which could be because the bromelain acts on the protein content, which is less in enamel. As there is no free radical production, even the application of antioxidants did not show improved bond strength in Group 2C.

Plasma is the fourth state of matter, an extremely reactive material comprises charged particles, and a strong electric field. Plasma treatment increased hydrophilicity of the dental materials on the dentin surface, thereby reducing the interface defects/voids and improve the interfacial quality.[21] In the current study, NTAP application to both bleached and microabraded surfaces, i.e., Groups 1D, 2D significantly improved the bond strength. This is in accordance with several studies which have proven NTAP to increase bond strength at the interface of tooth and composite by 60%.[22] This bonding at the interface helps in improving durability, longevity, and performance of the composite resin.[23]

Plasma application increases the wettability, hydrophilicity of the substrate and modifies its chemical structure by provisionally exposing and partially dispersing the functional groups. This results in improved infiltration of the bonding agent into the enamel and enhances the adhesive bond strength producing bonds that depend on surface chemistry rather than surface porosity, which is in accordance with Cho et al.[24] Even though the resin tags do not significantly contribute to the bond strength in the case of self-etch adhesives, the enhanced resin tag formation by NTAP in the present study could have contributed to the rise in initial bond strength.[7],[21]

NTAP causes molecular alterations such as reducing the molecules containing nitrogen and carbon and exposing the hydroxyapatite crystals, which is considered a significant improvement in the wettability of enamel. Short exposure periods of enamel surfaces to NTAP for only 30 s lead to the formation of super hydrophilic surfaces.[25] There appears to be a need for further clinical studies to compare and gather more details of plasma effects on the organic components of enamel, analyzing the different periods of plasma application and its implications on adhesive properties to the bleached and micro abraded enamel to support the present hypothesis.

Limitations of the study

  1. In the present study, even though plasma has shown better results of increased bond strength, the use of the same in the clinical scenario is practically difficult
  2. The results have to be confirmed with large-scale and in vivo studies.

   Conclusions Top

  • Composite resin application immediately following bleaching and microabrasion showed decreased mean SBS values
  • Following bleaching, NTAP, and GSE application significantly improved the SBS of composite resin
  • Following microabrasion, NTAP significantly increased the SBS of composite resin.


We would like to thank the Department of chemistry, Indian institute of space science and technology (IIST), Kerala, for the provision of the NTAP jet equipment and the Department of Conservative Dentistry and Endodontics, and the management of the SIBAR institute of dental sciences for their kind support and guidance throughout the study.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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Correspondence Address:
Dr. Sravanthi Tammineedi
Department of Conservative Dentistry and Endodontics, Sibar Institute of Dental Sciences, Takkellapadu, Guntur - 522 509, Andhra Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jcd.jcd_530_21

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