|Year : 2016 | Volume
| Issue : 2 | Page : 67-71
Comparative evaluation of remineralization efficacy of GC tooth mousse plus and enafix on artificially demineralized enamel surface: An in vitro study
Department of Conservative Dentistry and Endodontics, Swargiya Dadasaheb Kalmegh Smruti Dental College and Hospital, Nagpur, Maharashtra, India
|Date of Web Publication||19-Dec-2016|
Plot No. BA2, Shreenath Nagar, Near Onkar Nagar Square, Manewada Ring Road, Nagpur - 440 027, Maharashtra
Source of Support: None, Conflict of Interest: None
Background: The most common dental disease facing the human race is dental caries. For caries, management of early lesions, prevention of initiation, and interruption of progression are desirable. Remineralization is an effective way of preventive management of caries. Aim: To evaluate remineralization efficacy of two different remineralizing agents. Materials and Methods: Forty enamel samples were taken, and they were divided into four groups (n = 10). Demineralization was carried out with Groups A, B, and C. Remineralization was carried out with Groups A and B for 7 continuous days using casein phosphopeptide-amorphous calcium phosphate with fluoride (CPP-ACPF) and calcium sucrose phosphate (CaSP). Group D was kept as positive control with intact enamel without any surface treatment, whereas Group C was kept as negative control with surface demineralization of enamel. Microhardness testing was done using Vickers microhardness tester after 7-day remineralization cycle. Statistical Analysis: One-way analysis of variance and post hoc Tukey tests were performed. Results: Microhardness of CaSP and CPP-ACPF was comparable. Conclusion: Both remineralizing agents showed surface remineralization.
Keywords: Calcium sucrose phosphate, casein phosphopeptide-amorphous calcium phosphate with fluoride, Enafix, GC Tooth Mousse Plus, Remineralization
|How to cite this article:|
Gade V. Comparative evaluation of remineralization efficacy of GC tooth mousse plus and enafix on artificially demineralized enamel surface: An in vitro study. Indian J Oral Health Res 2016;2:67-71
|How to cite this URL:|
Gade V. Comparative evaluation of remineralization efficacy of GC tooth mousse plus and enafix on artificially demineralized enamel surface: An in vitro study. Indian J Oral Health Res [serial online] 2016 [cited 2018 May 25];2:67-71. Available from: http://www.ijohr.org/text.asp?2016/2/2/67/196097
| Introduction|| |
Dental caries is the most common oral ailment, affecting a large number of population in urban areas.  Enamel demineralization starts when pH drops below 5. This is the beginning of early enamel caries. Subsurface layer gets demineralized during primary stages of demineralization. However, neutralizing the oral pH can oppose demineralization. This process is achieved by remineralization. Remineralization increases amount of calcium and phosphate ions in the immediate environment of the lesion. In remineralization, building up of partly dissolved apatite crystals occurs and further caries process is prevented. 
Numerous remineralizing agents such as topical fluoride, stannous fluoride have been used for remineralization. It involves direct delivery of ions to the affected area. 
Restriction of initiation of dental caries can be achieved by milk and its products. Casein, calcium, and phosphate provide anticarcinogenic property to milk, giving its protective action.  When casein phosphopeptide-amorphous calcium fluoride is applied to oral environment, casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) complex molecule binds readily to enamel, biofilm, and soft tissues and delivers the calcium and phosphate ions strictly where it is needed. Studies done by Patil et al. and Jayarajan et al. reported that CPP-ACP with fluoride (CPP-ACPF) has more potential for remineralization as compared to CPP-ACP. ,
Recently, calcium sucrose phosphate (CaSP) was reintroduced as Enafix in the Indian market as a remineralizing agent. It decomposes to calcium, phosphate, and sucrose ions, thus resulting in increased rate of remineralization. 
This randomized control study focuses on evaluating the enamel remineralizing potential of CPP-ACPF (GC Tooth Mousse Plus, Leuven, Belgium) and CaSP (Enafix, Group Pharmaceuticals Limited, Bengaluru, Karnataka, India) using surface microhardness analysis (Vickers hardness test). Null hypothesis of this study was that both remineralizing agents, CPP-ACPF and Enafix, had similar remineralizing efficacy.
| Materials and methods|| |
In this study, forty freshly extracted sound maxillary premolars, extracted for orthodontic reasons were used. Teeth with any visible or detectable caries, or any white spot lesions, or any hypoplastic lesions were excluded. 
The teeth were cleaned of gross debris. Elimination of microbial growth was achieved by using an autoclave cycle for 40 min. 
The teeth were decoronated, and the crown portions were divided into four segments of two buccal and two palatal halves each, using a double-faced diamond disc mounted on a contra-angle handpiece. 
Enamel samples were embedded in self-cure acrylic with the enamel surface exposed. These samples were stored in deionized water until further use. A total of forty acrylic slabs embedded with enamel samples were produced and were then randomly divided into four groups of ten samples each [Figure 1] and [Table 1].
Demineralizing solution was prepared in the Department of Biochemistry. A digital pH meter (Slope Labtronics, Model LT-11, Panchkula, Haryana, India) was used to check pH during and after preparation of the solution. The composition of demineralizing solution used was as follows:
- 2.2 mM calcium chloride, CaCl 2 .2H 2 O (Loba Chemie Pvt. Ltd., Mumbai, Maharashtra, India)
- 2.2 mM monosodium phosphate, NaH 2 PO 4 .7H 2 O (Loba Chemie Pvt. Ltd., Mumbai, Maharashtra, India)
- 0.05 M lactic acid, C 3 H 6 O 3 (Loba Chemie Pvt. Ltd., Mumbai, Maharashtra, India).
The final pH was adjusted to 4.5 with 50% sodium hydroxide, NaOH (Loba Chemie Pvt. Ltd., Mumbai, Maharashtra, India). 
All the samples of Groups A, B, and D were then immersed into a glass container containing 50 ml of prepared demineralizing solution for 48 h at 37°C inside a Universal Incubator (Coslab, ISO 9001:2000, Ambala Cantt, Haryana, India). This demineralizing procedure was contemplated to produce a consistent subsurface lesion. After 48 h of incubation in the demineralizing solution, the teeth were washed with deionized water, dried with the help of an air syringe, and placed in different clean glass containers until further evaluation. 
The samples in Groups A and B were treated with respective remineralizing agents at every 24 h for 7 days. Samples were rubbed with respective remineralizing agent with the help of polishing cup attached to a contra-angle handpiece for 4 min [Figure 2], washed with deionized water, and then placed in artificial saliva. All samples were placed in the Universal Incubator at 37°C between each remineralizing cycle. In the control groups, samples were only washed with deionized water and placed in artificial saliva. Artificial saliva was renewed every 24 h just before immersion of freshly treated samples. 
After 7 cycles of remineralization, the surfaces were assessed for surface microhardness using Vickers microhardness testing machine [Figure 3]. The surface microhardness of the specimens was determined using MITUTOYO microhardness tester (Kawasaki, Japan). A load of 100 g was exercised steadily to the surface of specimens for 10 s using Vickers elongated diamond pyramid indenter under a ×40 objective lens. Five indentations were placed on the surface and the average value was considered for each specimen. Accuracy of values of diagonal length of indentation was determined under high magnification of ×400. The depth of the indentation was measured through a built-in scaled microscope and the values were converted to Vickers microhardness values.
The surface microhardness of samples was compared across study groups. The mean and standard deviation of microhardness of samples was obtained for each group and comparison was performed using one-way analysis of variance. The pair-wise comparison of mean microhardness between groups was carried out using Tukey's post hoc test. The statistical significance was tested at 5%, and the analysis was performed using SPSS 18.0 software (SPSS Inc., IBM, New York, USA).
| Results|| |
The analysis resulting into P < 0.0001 indicates statistically significant difference in the mean levels of surface microhardness (SMH) of different materials.
The mean for positive control was the highest 282.35 ± 28.97 MPa while that of negative control was the lowest 198.74 ± 25.46 MPa. In case of materials, CaSP that had a mean value of 234.24 ± 19.05 while CPP-ACPF had the least mean value of 213.91 ± 31.81 MPa [Table 2].
Pair-wise comparison of mean SMH between different study groups reveals that the mean microhardness of positive control group is significantly higher than all other groups as P < 0.05. The mean microhardness of CaSP (Enafix) was slightly higher than CPP-ACPF. However, the difference was statistically insignificant as P < 0.05. Yet, its mean difference was statistically significant from that of negative control [Table 3].
|Table 3: Pair-wise comparison of mean surface microhardness using post hoc Tukey|
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In summary, the study reveals that the overall difference of mean microhardness across groups is mainly due to positive and negative control groups. Mean values for two experimental groups are insignificantly different from each other. Thus, the null hypothesis was accepted.
| Discussion|| |
A subsurface carious lesion of enamel is clinically seen as a white spot lesion. The most prominent feature of a white spot lesion is that there is a subsurface demineralized zone with top layer of intact enamel. However, the mineral content of surface is deficient. As compared to sound enamel, white spot lesion shows a lower microhardness. 
Demineralization process is directly affected by the acidic environment produced by the metabolism of bacteria. During demineralization, calcium, phosphate, fluoride, carbonate, sodium, and magnesium ions diffuse out from the enamel surface into the saliva. More the acidic environment more is the outflow of the calcium ion and phosphate ion. Mineral content of surface is higher than the body of the lesion. As calcium and phosphate ions diffuse to the exterior, there is more probability of remineralization at the surface. 
In the present study, the specimens were placed in the demineralizing solution for 48 h at 37°C in the Universal Incubator. This resulted in a subsurface demineralization. 
Materials that are prone to cracking and possess microcrystalline structure are appropriate for microhardness measurement. As there are intact surface and subsurface demineralization, SMH measurement was a suitable technique for this experimental design. Therefore, in the present study, the microhardness values for each specimen were measured. 
In the present study, mean for positive control was the highest 282.35 ± 28.97 MPa while that of negative control was the lowest 198 ± 25.46 MPa, which showed that mean microhardness of the sound enamel was highest. Between the both experimental groups, CaSP, i.e., Enafix showed more microhardness (234.24 = 19.05) as compared to CPP-ACPF (213.91 = 31.81) but statistically both are insignificant.
CPP-ACPF is supersaturated solution of amorphous and crystalline calcium phosphate phases. It has added fluoride content. It is a stabilized composition so that spontaneous precipitation of calcium phosphate is stopped. The remineralizing capacity is directly proportional to the levels of free calcium and phosphate ions that are stabilized by CPP. 
When CPP-ACPF is applied on the tooth surface, its sticky CPP part readily mixes with enamel and biofilm releasing the calcium and phosphate ions. The free calcium and phosphate ions enter the enamel rods and form the apatite crystals again. 
Mehta et al. reported that CPP-ACP molecules need an acidic exposure to get activated and this would separate ACP from the casein. In the present study, the samples underwent demineralization only once. Less value of CPP-ACP in this study might be because the samples were not acid activated when CPP-ACPF was applied on the tooth surface. 
This is due to a difference in time between the release of ACP from CPP during the acid challenge and the time required to deposit calcium and phosphate into the lesion during remineralization. Another reason can be due to the short duration of application of the material. Therefore, it is necessary to have a longer period of application to be able to detect deposition of calcium and phosphate in the demineralized lesion.
CaSP is a combination of calcium salts of sucrose phosphate esters, mixed with inorganic calcium. It readily breaks down and releases calcium ions, phosphate ions, and sucrose phosphate ions into saliva. It is composed of 10%-12% calcium (wt%) and 8%-10% phosphorous (wt%). Calcium and phosphate ions in aqueous media form insoluble precipitates. CaSP forms aqueous solutions consisting of high concentration of calcium and phosphate without occurrence of precipitation. It acts as an ideal carrier for calcium and phosphate in water. Enafix acts by adsorption of sucrose phosphate ion rapidly on the enamel surface, thereby reducing the rate of acid dissolution of hydroxyapatite and quick remineralization by calcium and phosphate ion by common ion effect. 
This study was aimed to evaluate the effectiveness of newly introduced Enafix, which is an inexpensive, substitute for costly remineralizing agents and is suitable for the Indian economical background.
Drawbacks observed in the study include failure to remineralize artificial caries completely in 7-days time. Hence, the period of application for complete remineralization cannot be described for the remineralizing agents used. Although surface remineralization was confirmed, enamel subsurface remineralization was not evaluated in the study. Within the limitations of this in vitro study, one can conclude that remineralization takes place with the use of CaSP and CPP-ACPF. However, complete remineralization did not occur within 7 days.
| Conclusion|| |
Enafix being a cost effective material as compared to GC Tooth mousse plus, it can be used as an alternative for better remineralization in Indian scenario.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Patil N, Choudhari S, Kulkarni S, Joshi SR. Comparative evaluation of remineralizing potential of three agents on artificially demineralized human enamel: An in vitro
study. J Conserv Dent 2013;16:116-20.
Lata S, Varghese NO, Varughese JM. Remineralization potential of fluoride and amorphous calcium phosphate-casein phosphopeptide on enamel lesions: An in vitro
comparative evaluation. J Conserv Dent 2010;1:42-6.
Jayarajan J, Janardhanam P, Jayakumar P, Deepika. Efficacy of CPP-ACP and CPP-ACPF on enamel remineralization - An in vitro
study using scanning electron microscope and DIAGNOdent. Indian J Dent Res 2011;22:77-82.
Sargod SS, Bhat SS, Hegde S, Karunakaran R. Remineralization potential using calcium sucrose phosphate (Enafix) on artificial carious lesion: A Polaroid microscopic study. Indian J Appl Res 2015;5:421-3.
Shetty S, Hegde MN, Bopanna TP. Enamel remineralization assessment after treatment with three different remineralizing agents using surface microhardness: An in vitro
study. J Conserv Dent 2014;17:49-52.
Harris R, Schamschula RG, Beveridge J, Gregory G. The cariostatic effect of calcium sucrose phosphate in a group of children aged 5-17 years. IV. Aust Dent J 1969;14:42-9.
Mehta R, Nandlal B, Prashanth S. Comparative evaluation of remineralization potential of casein phosphopeptide-amorphous calcium phosphate and casein phosphopeptide-amorphous calcium phosphate fluoride on artificial enamel white spot lesion: An in vitro
light fluorescence study. Indian J Dent Res 2013;24:681-9.
[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2], [Table 3]