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Table of Contents   
ORIGINAL ARTICLE  
Year : 2011  |  Volume : 14  |  Issue : 2  |  Page : 113-116
An ex vivo study to evaluate the remineralizing and antimicrobial efficacy of silver diamine fluoride and glass ionomer cement type VII for their proposed use as indirect pulp capping materials - Part I


Department of Conservative Dentistry and Endodontics, Centre for Dental Education and Research, All India Institute of Medical Sciences, New Delhi, India

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Date of Submission14-Sep-2010
Date of Decision28-Dec-2010
Date of Acceptance29-Jan-2011
Date of Web Publication7-Jul-2011
 

   Abstract 

Aim : Indirect pulp capping (IPC) preserves the pulp vitality by disinfecting and remineralizing remaining carious dentin. In the present study, glass ionomer (GC, FUJI VII) and silver diamine fluoride (SDF) were tested and compared to calcium hydroxide for their antimicrobial efficacy and remineralizing potential.
Materials and Methods : Dentin disks prepared from 45 freshly extracted first premolars were divided into three groups (n = 15). Each disk was cut into two equal parts, in which one half formed the control. Thirty dentin samples were used for ion estimation and the other 15 for microhardness testing. Atomic absorption spectrophotometry, colorimetric and potentiometric titration analyses were performed for calcium, phosphate and fluoride ion detection, respectively. The antimicrobial efficacy was analyzed using pure culture of Streptococcus mutans and mixed flora.
Results : Increase in the levels of calcium and phosphate ions was the highest in calcium hydroxide group. Both SDF and GC VII groups showed significant increase in fluoride ion levels. Samples treated with GC VII showed maximum increase in micro hardness. The highest zone of bacterial inhibition was found with SDF group.
Conclusions : This in vitro study documented the remineralizing, re-hardening and antimicrobial efficacy of both SDF and GC VII and hence can act as effective IPC materials.

Keywords: Calcium hydroxide; glass ionomer; high-fluoride releasing dental materials; indirect pulp capping; remineralization; silver diamine fluoride

How to cite this article:
Gupta A, Sinha N, Logani A, Shah N. An ex vivo study to evaluate the remineralizing and antimicrobial efficacy of silver diamine fluoride and glass ionomer cement type VII for their proposed use as indirect pulp capping materials - Part I. J Conserv Dent 2011;14:113-6

How to cite this URL:
Gupta A, Sinha N, Logani A, Shah N. An ex vivo study to evaluate the remineralizing and antimicrobial efficacy of silver diamine fluoride and glass ionomer cement type VII for their proposed use as indirect pulp capping materials - Part I. J Conserv Dent [serial online] 2011 [cited 2019 Apr 22];14:113-6. Available from: http://www.jcd.org.in/text.asp?2011/14/2/113/82603

   Introduction Top


In deep dentinal caries, dentin discoloration occurs far in advance of the infection by microorganisms, and as much as 2 mm of the softened or discolored dentin is not infected. Carious dentin actually consists of two distinct layers having different ultramicroscopic and chemical structures. [1] The outer carious layer is irreversibly denatured, infected and incapable of being remineralized and should be removed. The inner carious layer is reversibly denatured, not infected, and capable of being remineralized and should be preserved. [2] To prevent pulp exposure, indirect pulp capping (IPC) is done, in which caries is excavated and the tooth is restored with suitable IPC material. Over the years, calcium hydroxide has emerged as gold standard for IPC. Glass ionomer cement (GIC) has seen its popularity as a restorative material, as it was identified that fluoride release has an anticariogenic property. [3] Type VII GIC, which is a high-fluoride releasing material, was introduced for pit and fissure sealing. In addition, it has an excellent sealing property.

Silver diamine fluoride (SDF) has also been used as a cariostatic agent. Various clinical studies [4] have reported its utility in the treatment and prevention of caries. SDF helps in the deposition of silver phosphate to restore mineral content, resulting in rehardening of tooth structure. It also releases fluoride.

The present study was planned to evaluate the remineralizing, rehardening and antimicrobial properties of these two high-fluoride releasing materials, i.e. GIC type VII and SDF, as potential IPC materials and compare them with calcium hydroxide.


   Materials and Methods Top


Forty-five mandibular first premolars extracted for orthodontic treatment (average age 16 years) were kept in deionized water at room temperature for a maximum period of 1 month before commencing the study. The teeth were sectioned perpendicular to the long axis at 2.5 mm from the buccal cusp tip to make dentin disks of 2 mm thickness. They were demineralized using 22 ml of 6% carboxymethyl cellulose acid gel by weight at pH 5.0 for 2 weeks without renewal. Thereafter, the specimens were thoroughly rinsed with deionized water and were divided into three groups of 15 each. Each dentin disk was further divided into two equal parts, where one half served as the control and the other as test. The three test groups were as follows:

Group I: Calcium hydroxide (Dycal® ,Dentsply Caulk, Dentsply International.Inc, DE, USA.)

Group II: SDF (Saforide® , J. Morita Corporation. Osaka, Japan)

Group III: GC type VII (GC Corporation. Tokyo, Japan)

All the three materials were mixed as per manufacturers' instructions. As the SDF was in a gel form, it was mixed with zinc oxide powder. The specimens were allowed to set for an hour and subsequently coated with nail polish except at the pulpal side of the dentin disk. They were placed in 5 ml of remineralizing buffer at 37°C for 6 weeks.

After 6 weeks, the test materials were carefully separated from the dentin disk samples and washed with deionized water. Thirty dentin samples were used for ion estimates and 15 for microhardness testing. Atomic absorption spectrophotometry (AAS), colorimetric and potentiometric titration analyses were performed for calcium, phosphate and fluoride ion detection, respectively.

Bacteriological assay

The antimicrobial efficacy of the test materials was tested on pure culture against Streptococcus mutans and mixed flora. Zone of inhibition was measured on the plates at 24 and 72 hours.

Statistical analysis

The raw data were used for the statistical analysis using SPSS version 11. One-way Analysis of Variance (ANOVA) was used to test for significance of differences between class means of all the groups. [5]


   Results Top


Calcium, phosphorous and fluoride ions in demineralized samples did not show any statistically significant difference in all three groups, confirming the uniformity of samples.

The percentage increase in calcium, phosphorous and fluoride ions after treatment with the three test materials is shown in [Table 1] and intergroup comparison is shown in [Table 2].
Table 1: Percentage increase in ion content of dentin after treatment with test materials


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Table 2: Inter-group comparison of mean differences in calcium, phosphate and fluoride levels with test materials


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It is found that increase in calcium level was the highest in calcium hydroxide group as compared to both SDF and GC VII groups, with GCVII group showing the least increase. There was no statistically significant difference between increase in phosphate levels after application of calcium hydroxide and SDF, but there was statistically significant difference between calcium hydroxide and GC VII groups with regard to increase in phosphate levels. The difference between percentage increases of phosphate level in SDF and GC VII groups was statistically significant. Overall, calcium hydroxide exhibited the highest phosphate deposition within dentin than either SDF or GC VII.

Both SDF and GC VII groups showed significant increase in fluoride levels as compared to calcium hydroxide group. The difference between fluoride ion increase in SDF and GC VII groups was statistically significant. Overall, SDF exhibited the highest F deposition within dentin than either calcium hydroxide or GC VII.

Microhardness

Increase in microhardness was found in all the three groups. GC VII was seen to have the maximum increase in knoop Hardness Number (KHN) followed by SDF and calcium hydroxide groups.

Antimicrobial efficacy

The antimicrobial efficacy (zone of inhibition) against S. mutans by calcium hydroxide and SDF groups was more than that of GC VII group, both after 1 and 3 days. For mixed flora, the zone of inhibition was the highest in SDF group, followed by calcium hydroxide and GC VII groups, both after 1 and 3 days for the mixed flora.


   Discussion Top


The present study was designed to assess quantitatively any change in the chemical composition of artificially demineralized human dentin stored in remineralizing buffer, treated with high-fluoride releasing materials for their potential use as IPC materials.

While calcium hydroxide has been extensively utilized in the past for IPC, [6] the use of GC VII and SDF has been reserved for remineralization of superficial enamel lesions. [7] Partial removal of carious tissue treated with calcium hydroxide had shown good results based on clinical, radiographic, ultra structural and microbiological outcomes. [8] Remineralization of remaining carious dentine has been detected both biochemically and radiographically. [9] The success of calcium hydroxide as an IPC agent is ascribed to its ability to inhibit bacterial enzymes by means of hydroxyl ions that act on the cytoplasmic membrane of bacteria (generating the antibacterial effect) and that of activating tissue enzymes, such as alkaline phosphatase, which has an influence on mineralization. [10] However, this effect was significantly weaker than that found with fluoride. [11] If fluoride ions are present at the crystal surface in sufficient concentration, these ions can get adsorbed onto the surface of crystals and markedly inhibit further demineralization by acid. [12] Calcium and phosphate, primarily from saliva, and other topical sources, diffuse into the tooth and with the help of fluoride, build on existing crystal remnants. [13] The new crystal surface may be similar to fluorapatite depending on the concentration of fluoride present. However, remineralization with fluoride alone is more superficial than from calcium, phosphate and fluoride together. All the three ions together can effect remineralization to a greater depth and increase the resistance to future acid attack. [14]

Thus, high-fluoride releasing materials in the form of GC VII and SDF were selected to deliver fluoride in sufficient concentrations and test the hypothesis that if these two materials have equal or greater potential than calcium hydroxide in remineralization of dentin, then they can be promising IPC materials in the future.

In the present study, the samples were subjected to acid dissolution for inducing artificial demineralization by a lactic acid based demineralizing agent. The gel contained 0.1 M lactic acid titrated to pH 5 using a concentrated potassium hydroxide solution. To simulate the pulpal environment present in the oral cavity, the dentin disks were stored in a remineralizing solution containing 1.5 mmol/l CaCl 2 , 0.9 mmol/l KH 2 PO 4 , 130 mmol/l KCl, and 20 mmol/l 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES). The pH was then adjusted to 7 with NaOH. [15]

In the present study, AAS was used to evaluate the changes in calcium level in dentin. [16] Wet digestion method was used with concentrated HNO 3 . Colorimetric assays were used for phosphorous ion estimation. Potentiometric titration was used for fluoride determination. [17] To test whether any discrepancy existed between different sources of samples, specimens from each test group were tested before application of the test materials. These specimens acted as control groups. No statistically significant difference in the mean ion concentration existed between the control specimens from the three groups, which confirmed the consistency of the nature of specimen. In the present study, calcium hydroxide caused the maximum increase in calcium ions. The possible mechanism of remineralization with calcium hydroxide in vitro is by the dissociation of calcium hydroxide into calcium and hydroxyl ions when it comes in contact with water present in dentin and deposition of these calcium ions on the superficial layer of demineralized dentin. [10] This was further evidenced by crystal units of calcium and phosphorous precipitated on the surface of calcium hydroxide capped dentin samples, [18] suggesting that calcium ions were released from calcium hydroxide containing materials that modify the dentin surface.

Specifically, SDF group showed statistically significant increase in the level of calcium. While literature has shown SDF to be associated with definite increase in fluoride level, [19] it is interesting to conjecture what may have caused the increased value for calcium. It may be attributed to the influx of calcium ions from remineralizing buffer, which simulated dental pulp environment. The underlying mechanism may be attributed to the formation of SDF reaction products like silver phosphate and fluoride. These fluoride ions may have attracted calcium from the environment (in remineralizing buffer solution) as its reaction with tooth calcium is competed by silver phosphate.

Calcium hydroxide group also exhibited greater deposition of phosphate than either SDF or GC VII groups. However, when calcium hydroxide and SDF were considered, the difference was statistically insignificant. Significant difference was found, however, with GC VII which showed least amount of phosphorous deposition as compared to calcium hydroxide. This finding is consistent with that of Conrado [20] who showed an increase in phosphorus concentration from both vital and pulp-extirpated teeth, treated with chemically pure calcium hydroxide. He could not, however, explain the mechanism for this increased level in pulpless teeth. We propose that the observed increase in phosphate level (10.01%) may be attributed to the calcium depositing effect of calcium hydroxide that creates a zone of increased saturation, which may encourage diffusion of anions like phosphorous from pulpal end (remineralizing solution). With SDF, although direct release from the material is not known, the mechanism most likely follows the pattern as seen for calcium ions. Also, SDF may be more useful in remineralizing dentin caries which presented a greater amount of protein substrate, carbonates and phosphates for reaction with silver ion released by SDF. [19] Increase in phosphorous ion was also seen with GC VII; however, the increase was minimal (6.186%). This is well supported by Kitasako et al.[21] who reported increased phosphate and magnesium content underneath GIC restorations.

All samples following SDF application had significant rise in fluoride level. It was also seen that this rise was greater than with GC VII, while specimens restored with calcium hydroxide showed minimal increase in fluoride level. SDF (38% solution) contains fluoride levels up to 100,000 ppm. [23] Thus, the high percentage increase (802.417%) may be due to the high content of fluoride in SDF. A high value of fluoride ions was also seen with GC VII (404.78%). This finding is in accordance with previous studies utilizing conventional GIC. [15] Calcium hydroxide showed insignificant increase (0.86%) owing to the absence of any fluoride source in its composition.

In the present study, all three groups showed increase in microhardness value, the value being highest in GC and lowest in calcium hydroxide groups. GC VII and SDF create an environment of fluoride hypersaturation that results in the formation of fluorapatite crystals. On the other hand, with calcium hydroxide, no such phenomenon is seen and it results in the formation of only hydroxyapatite crystals. Fluorapatite crystals are larger in size and thus result in closely packed structure with fewer voids. This accounts for greater microhardness value when compared with hydroxyapatite crystals.

Surprisingly, SDF had comparatively lower microhardness value than GC VII in spite of higher fluoride release. This finding can be attributed to the fact that SDF deposits silver phosphate which is the main mechanism of action of SDF responsible for increased hardness. [23] The silver might actively compete with the binding site of fluoride to calcium of tooth sample. The fluorapatite layer formed with SDF may thus only be superficial, leading to lower microhardness value when compared to GCVII.

The present study highlights that both GC type VII and SDF have greater remineralization potential than that of calcium hydroxide in terms of increased mineral content, particularly fluoride, and microhardness value, when applied to artificially demineralized human dentin and hence can be used as effective IPC materials.

 
   References Top

1.Fusayama T. Two layers of carious dentin: Diagnosis and treatment. Oper Dent 1979;4:63-70.  Back to cited text no. 1
    
2.Miyauchi H, IW aku M, Fusayama T. Physiological recalcification of carious dentin. Bnl Tokyo Med Dent Univ 1978;25:169-79.  Back to cited text no. 2
    
3.Wilson AD, Kent BE. A New transluscent cement for dentistry. The glass ionomer cement. Br Dent J 1972:132-3.  Back to cited text no. 3
    
4.Yamaga R, Yoshita S, Yokomizo I. Diamine silver fluoride and its clinical application. J Osaka Univ Dent Sch 1972;12:1-20.  Back to cited text no. 4
    
5.Lindman HR. Analysis of variance in complex experimental designs. San Francisco: W. H. Freeman and Co; 1974. p. 297.  Back to cited text no. 5
    
6.Oliveira EF, Carminatti G, Fontanella V, Maltz M. The monitoring of deep caries lesions after incomplete dentine caries removal: Results after 14-18 months. Clinical Oral Inv. 2006;2:134-9.  Back to cited text no. 6
    
7.Domenick TZ. Dentifrices, mouthwashes, and remineralization/caries arrestment strategies. BMC Oral Health 2006;6:1-9.   Back to cited text no. 7
    
8.Marchi JJ, Araujo FB, Fröner AM, Straffon LH, Nör JE. Indirect pulp capping in the primary dentition: A 4-year follow-up study. J Clin Pediatr Dent. 2006;31:68-71.  Back to cited text no. 8
    
9.Eidelman E, Finn S, Koulourides T. Remineralization of carious dentin treated with calcium hydroxide. J Dent Child 1965;32:218-25.  Back to cited text no. 9
    
10.Estrela C, Sydney GB, Bammann LL, Junior OF. Mechanism of action of Calcium and Hydroxyl ions of Calcium hydroxide on tissues and bacteria. Braz. Dent J 1995;6:85-90.  Back to cited text no. 10
    
11.SaitoT, Toyooka H, Ito S, Miles A. In vitro Study of Remineralization of Dentin: Effects of Ions on Mineral Induction by Decalcified Dentin Matrix. Caries Res 2003;37:445-9.  Back to cited text no. 11
    
12.Featherstone JD, Glena R, Shariati M, Shields CP. Dependence of in vitro demineralization and remineralization of dental enamel on fluoride concentration. J Dent Res 1990;69:620-5.  Back to cited text no. 12
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13.Featherstone JD. The science and practice of caries prevention. J Am Dent Assoc 2000;131:887-99.  Back to cited text no. 13
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14.Reynolds EC, Walsh LJ. Additional aids to the remineralisation of tooth structure. In: Mount GJ, Hume WR editors. Preservation and Restoration of Tooth Structure. 2 nd ed. Brighton, Australia: Knowledge Books and Software; 2005: Chapter 8.  Back to cited text no. 14
    
15.Exterkate RA, Damen JJ , Ten Cate JM. Effect of Fluoride- Releasing Filling Materials on Underlying Dentinal Lesions in vitro. Caries Res 2005;39:509-13.  Back to cited text no. 15
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16.M Adel, Khairy P, Aliaa E. In vitro Evaluation of the Remineralizing Capacity of fluoride Releasing Restorative Materials at the Internal Cavity Wall and External Margin. Cairo Dent J 2007;23:239-47.  Back to cited text no. 16
    
17.Yin F, Yao Y, Liu CC, Wena ML. Developments in the analysis of fluoride.1997-1999 . Fluoride 2001;34:114-25.   Back to cited text no. 17
    
18.Tziafas D, Economides N. Formation of crystals on the surface of calcium hydroxide-containing materials in vitro. J Endod 1999;25;539-42.  Back to cited text no. 18
    
19.Delbem AC, Bergamaschi M, Takebayashi KS, Frederico R. Effect of fluoridated varnish and silver diamine fluoride solution on enamel demineralization: pH-cycling study. J Appl Oral Sci 2006;14:88-92.  Back to cited text no. 19
    
20.Conrado CA. Remineralization of carious dentin: In vivo micro radiographic and chemical studies in human permanent teeth capped with Calcium hydroxide. Braz. Dent J 2004;3:186-9.  Back to cited text no. 20
    
21.Kitasako Y, Nakajima M, Foxton R.M, Aoki K, Pereira PN, Tagami J. Physiological remineralization of artificially demineralized dentin beneath glass ionomer cements with and without bacterial contamination. Oper Dent 2003;28:274-80.  Back to cited text no. 21
    
22.Gotjamanos T, Orton V. Fluoride ion concentration in 40 per cent silver fluoride solutions determined by ion selective electrode and ion chromatography techniques. Aust Dent J 1998;43:55-6.  Back to cited text no. 22
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23.Kato S, Fusayama T. Recalcification of Artificially Decalcified Dentin In vivo. J Dent Res 1970;49:1060-7.  Back to cited text no. 23
[PUBMED]  [FULLTEXT]  

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Correspondence Address:
N Shah
Department of Conservative Dentistry and Endodontics, Centre for Dental Education and Research, All India Institute of Medical Sciences, New Delhi - 110 029
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0972-0707.82603

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