Evaluation of Microhardness of Residual Dentin in Primary Molars Following Caries Removal with Conventional and Chemomechanical Techniques: An In vitro Study.

BACKGROUND: Many patients consider removal of caries to be a very unpleasant experience. Removal of caries with conventional drill is considered traumatic mainly due to fear and anxiety of children and their parents. Minimally invasive dentistry adopts a philosophy that integrates prevention, remineralization, and minimal intervention for the placement and replacement of restorations, thus reaching the treatment objective using the least invasive surgical approach, with the removal of the minimal amount of healthy tissues. Chemomechanical caries removal (CMCR) is a method for minimally invasive, gentle dentin caries removal based on biological principles which is an effective alternative to the traditional method. The present study was done to compare the microhardness of sound dentin before and after carious removal using a chemomechanical method and a conventional method.
MATERIALS AND METHODS: The present in vitro study was done on 28 proximal surfaces of fourteen extracted primary molars (with active caries on one proximal surface and sound side as control). The study was done to assess the Knoop microhardness of remaining dentinal surface after caries removal using a slow speed conventional bur and a chemomechanical method (Carie-Care™). Results and.
CONCLUSION: The rotary instrument group showed a consistent microhardness value with not much difference according to depth. The chemomechanical group showed a lesser microhardness value closer to the cavity floor than away from it. The microhardness values at all depths were significantly different for each treatment group with an increased value seen in the rotary group. The mean microhardness values of residual dentin in treated side were found to be insignificant when compared among each interval in each group. The microhardness of sound dentin had high significant difference from that of residual dentin in both the rotary group and the chemomechanical group.

INTRODUCTION

Dental caries is one of the most common chronic oral infections and is the second largest cause of tooth loss after periodontitis.[1] Whether dental caries progresses, stops, or reverses is dependent on a balance between demineralisation and remineralisation. This will lead to either cavitation within the tooth or repair and reversal of the lesion, or maintenance of the status quo.[2] Remineralization is frequent, especially when the biofilm pH is restored by saliva, which acts as a buffer. The remineralized areas have a higher concentration of fluoride and less microporous enamel structure than the original tooth structure because of the acquisition of calcium and phosphates from saliva.[3]

Many patients consider removal of caries to be a very unpleasant experience. Removal of caries with conventional drill is considered traumatic mainly due to fear and anxiety of children and their parents. However, mechanical preparation often induces pain, and local anesthesia is therefore needed.[4] Dentin caries removal with the drill may be traumatic to the pulp due to pressure, thermal damage, and vibration.[56] It is often difficult to establish how much tooth material should be removed, which often leads to overextended cavities.[5]

With the development of new dental restorative materials, advances in adhesive dentistry, the management of dental caries has drastically evolved from G.V. Black's “Extension for prevention” to “Construction with conservation.”[7] This concept includes the early detection of lesions, individual caries risk assessment, nonsurgical interventions, and modified surgical approach that includes smaller tooth preparations with modified cavity designs and adhesive dental materials and repair rather than replacement of failing restorations.

Minimally invasive dentistry adopts a philosophy that integrates prevention, remineralization, and minimal intervention for the placement and replacement of restorations, thus reaching the treatment objective using the least invasive surgical approach, with the removal of the minimal amount of healthy tissues. It includes the following different techniques such as air abrasion, atraumatic restorative technique,[89] sono-abrasion,[10] laser,[11] and chemomechanical caries removal (CMCR).[12]

CMCR is a method for minimally invasive, gentle dentin caries removal based on biological principles which is an effective alternative to the traditional method. It involves the application of a chemical solution to the carious dentin followed by gentle removal with hand instruments,[13] avoiding pulp irritation and patient discomfort. A variety of CMCR agents have been used since 1972 such as GK-101,[14] GK-101E,[15] Caridex,[16] Carisolv™ gel,[15] papacarie.[17]

Carie-Care™ gel is a chemomechanical caries removal agent developed in collaboration with Vittal Mallya Scientific Research Foundation in May 2011. It contains papaya extract (Carica papaya) and clove oil (Syzygium aromaticum).[18]

Studies regarding the fate of microhardness of the remaining dentin after removal of caries in primary teeth using Carie-Vare™ are minimal. Thus, the purpose of this study was to compare the microhardness of sound dentin before and after carious removal using a chemomechanical method and a conventional method.

MATERIALS AND METHODS

The present in vitro study was done in the Department of Pedodontics and Preventive Dentistry, Rajah Muthiah Dental College and Hospital, Annamalai University in association with Centre for Materials Joining Research, Annamalai University, to assess the remaining dentinal surface after carious tooth tissue removal with a slow speed conventional bur and a chemomechanical method (Carie-Care™) using the Knoop microhardness test.

Fourteen extracted primary molars, with active carious cavities on one proximal surface, were divided into two experimental groups as follows, in accordance with the carious tissue removal method: conventional mechanical treatment – slow speed rotary instrument – and chemomechanical method – Carie-Care™. Radiographs were taken to confirm the amount of dentin present in each tooth. Those teeth with <2 mm of dentin were excluded from the study.

Carious tissue removal using the conventional technique

Carious tissue removal using the conventional technique was performed with a spherical diamond bur with the largest diameter compatible with the cavity size, at slow speed, under water cooling, by a single operator. To gauge carious tissue removal, a dental explorer was used to check, until hard dentin was obtained.

Carious tissue removal using chemomechanical method

For the Carie-Care group, the product was applied and left in the cavity for 30 s, and carious dentin was afterward removed with a Maillefer curette that comes with the Carisolv™ system kit. The gel was reapplied until it presented a light coloring, indicative of nonexistence of softened carious tissue, and confirmed with the use of the dental explorer, to assess the remaining dentin hardness.

Preparing test specimens for microhardness test

After carious tissue removal, the teeth were longitudinally sectioned – under water cooling – in the mesiodistal direction of the crown, at the center of the cavity, until two sections were obtained. One of the sections was embedded in acrylic resin[19] so that the area to be analyzed remained exposed. Polishing was done at the Centre for Material Joining Research, Department of Manufacturing Engineering, Annamalai University, in a rotary polisher, with 600, 1000, 1200, 2000 and 4/0 grit abrasive papers, and final polishing was done with a felt cloth and diamond paste (0.2 μ). This preparation was considered ideal when, under optic microscopy, the specimens appeared to be shiny and without the presence of scratches.

The microhardness test was performed at Apollo Test House, Bangalore, on sound dentin and on treated dentin of the same specimen. For this analysis, a Leitz Miniload microhardness tester was used, with a Knoop indenter using a static load of 25 g applied for 30 s on the sound dentin and 15 g for 30 s on treated dentin. On the dentin submitted to carious tissue removal, 18 indentations were made – three at each distance – starting from 50, 250, and 500 μm from the base of the carious cavity [Figure 1], and on the sound dentin, 18 indentations were made – three at each distance – from 50, 250, and 500 μm from the dentinoenamel junction [Figure 2]. The indentations were made with a distance of 100 μm between them.[20]

Figure 1

Indentations at 50, 250 and 500 μm from the base of the carious cavity

Figure 2

Indentations at 50, 250 and 500 μm from the dentinoenamel junction

RESULTS

The factors analyzed in this study were “type of treatment” (rotary instrument and chemomechanical methods), “indentation intervals” (50, 250, 500 μm depths), and “type of tissue” (sound and treated).

The measurement of the longest diagonal of the indentation was measured. The value obtained was applied to the formula: HK (kgf/mm2) = (14230 × P) ÷ l2

Where HK = Knoop hardness in kgf/mm2

P = Load in grams

l = Length of the longer diagonal in microns.

The obtained data were analyzed using two group t-test for comparison after checking the variance using f-test. The statistical values were then tabulated to arrive at the result of the present study.

The mean Knoop microhardness (kgf/mm2) of dentin after carious tissue removal in both the groups at each depth is shown in Table 1. It was observed that the rotary instrument group showed a consistent microhardness value with not much difference according to depth. In the chemomechanical group, microhardness value at 50 μm was lesser than microhardness at 250 and 500 μm.

Table 1

Mean microhardness at each interval

The mean Knoop microhardness of dentin after caries removal in each depth for both the groups is compared in Graph 1. The statistical evaluation of the data showed that the mean microhardness of dentin at 50 μm for the bur group to be 47.54 ± 7.14 kgf/mm2 and that for Carie-Care™ group to be 25.84 ± 6.96 kgf/mm2. At 250 μm, the mean value for bur group was 47.90 ± 8.00 kgf/mm2 and that for Carie-Care™ group was 30.19 ± 9.09 kgf/mm2. At 500 μm, the mean microhardness value for bur group was 47.32 ± 7.36 kgf/mm2 and for Carie-Care™ group was 29.33 ± 8.17 kgf/mm2. According to two group t-test, all the comparisons showed high statistical significance (P < 0.05), showing that the microhardness value at all depths were significantly different for each treatment group.

Graph 1

Comparison of Microhardness of Bur group and Carie-CareTM group at each interval

The comparison mean Knoop microhardness of dentin at each depth interval for bur group is shown in Graph 2. The values for all the three depths were compared among each other which showed P > 0.05 suggesting statistical insignificance. The same was done with the chemomechanical group [Graph 3]. This group also showed statistical insignificance (P > 0.05).

Graph 2

Mean Microhardness of dentin at each interval for Bur group

Graph 3

Mean Microhardness of dentin at each interval for Carie-Care™ group

The comparison between the mean Knoop microhardness of dentin in sound side and treated side of bur group is compared in Graph 4 and for Carie-Care™ in Graph 5. The statistical evaluation of the mean microhardness at 50, 250, and 500 μm showed high statistical significance (P < 0.05). This showed that there was high significant difference between the microhardness of sound dentin and residual dentin in the treated side of the bur group and Carie-Care™ group.

Graph 4

Microhardness of dentin in sound side and treated side (Bur group)

Graph 5

Microhardness of dentin in sound side and treated side (Carie-Care™ group)

DISCUSSION

Dentin caries can be divided into two distinct layers. The outer layer is contaminated by bacteria causing a nonremineralizable necrotic collagen matrix. In the inner layer, bacteria are much less frequently observed and the collagen has been reversibly denaturated but retains the crossbanded ultrastructure. If the acid challenge is removed, the inner layer has the potential to remineralize.[21]

Contemporary concepts of caries management deal with preservation of the inner layer. Chemomechanical technique is the only method that is selective in caries dentin removal.[22] Various studies have advocated the advantages of chemomechanical carious tissue removal.[2324252627]

The present in vitro study was conducted to assess and compare the microhardness of remaining dentinal surface after carious tooth tissue removal with a slow speed conventional bur and a chemomechanical method (Carie-Care™) in primary molars.

Conventional caries removal techniques include the use of high- and low-speed burs. This technique allows faster treatment but may promote unnecessary structure removal. In the present study, slow-speed diamond bur was used for the conventional technique of carious removal. This was adopted to standardize the speed at which the caries was removed. A round carbide bur generally works well in its removal of caries-infected dentin and is used in a far more conservative low-speed handpiece system.[28]

The CMCR agent used in the present study, Carie-Care™, was launched in May 2011, and studies regarding this material were minimal. Carie-Care™ is a gel-based formulation containing a purified proteolytic cysteine enzyme, papain, from the plant C. papaya (Papaya) along with clove oil. Each milliliter of Carie-Care™ contains 100 mg of papaya extract and 2 mg of clove oil. It also contains small amounts of chloramines. Sodium methylparaben and sodium propylparaben are used as preservatives.[18]

In the present study, mechanical carious tissue removal was performed until sound dentin was obtained, gauged by the test performed with the dental explorer as done in previous studies by Flukiger et al.[29] and Haak et al.[10] In CMCR, the manufacturer's guidelines were followed. Although the Carie-Care™ manufacturer recommends the use of blunt curettes, it was opted to use the Maillefer curette supplied with the Carisolv™ Kit in accordance with the study done by Corrêa et al.[26]

Microhardness analysis has been used as a method to assess loss and reincorporation of minerals to the dental tissue because the reduction in the numerical hardness value presents a linear relation to mineral loss. Among the various microhardness tests, Knoop microhardness analysis has significant correlation with the amount of mineral loss from the tooth structure.[30] The microhardness value from various areas of dentin was significantly different. This may be due to the difference in inorganic compositions in different locations. The hardness of dentin also depends on the state of mineralization of tissue.[31]

Two types of dentinal tissues were assessed in this study – sound and treated, so different loads were used, to obtain indentations of similar quality. This technique was followed in the study done by Corrêa et al.[26] The Knoop indenter loads used were 15 g for 30 s on carious dentin and 25 g for 30 s on sound dentin.[20] Such loads were considered to be sufficient to permanently deform the assessed structures, without the occurrence of elastic deformations that could lead to any alteration.[32]

The mean microhardness of dentin for bur group at 50, 250, and 500 μm in the treated side was 47.55 ± 7.14, 47.90 ± 8.00, and 47.32 ± 7.36 kgf/mm2, respectively. The mean microhardness values of dentin that were obtained for Carie-Care™ group at 50, 250, and 500 μm in the treated side were 25.84 ± 6.96, 30.19 ± 9.09, and 29.33 ± 8.17 kgf/mm2, respectively. The results of the present study showed that the mean microhardness of bur group was more than the Carie-Care™ group in the treated side. There was significant difference between two treatment groups (P < 0.05). The mean microhardness values of residual dentin in the treated side for the bur group showed a consistent microhardness value with not much difference with respect to depth. The treated side of Carie-Care™ group showed a lesser microhardness value closer to the cavity floor than away from it. This may be because the bur method of excavation tended to overprepare cavities due to lack of sensitivity of the tactile feedback. This resulted in gross, rapid removal of tissue with reduced control over the whole process. The operator could not identify the true clinical end point, so the excavation procedure continued into healthier dentin leading to eventual overpreparation.[6]

Splieth et al.[33] said that caries removal with Carisolv™ leaves up to 50 μm more of remineralizable carious dentin in the inner caries-inactive layer than round burs.

The selectivity lies in the mechanism of action of Carie-Care™ which contains papain. The chemomechanical agents rely on the action of proteolytic agents such as papain and sodium hypochloride to further degrade the partially demineralized and altered dentin matrix that has been previously exposed to bacterial action (infected dentin), thus facilitating its removal and preventing damage to the underlying demineralizable tissues (affected dentin).[6] The papain enzyme is a plant-derived cysteine protease of broad proteolytic activity,[18] used as a chemomechanical material since its introduction by Bussadori's group. The selective interaction of the enzyme with the affected components of carious dentin has been suggested to be due to the lack of an antiprotease α1-antitrypsin, which inhibits protein digestion in sound collagen-based tissues. α1-antitrypsin is a molecule produced by hepatocyte and mononuclear phagocytes and is extensively found in the lower respiratory tract during the degradation of the lung parenchyma in respiratory diseases, such as pulmonary emphysema (Hogg and Timens, 2009). It is likely that the inflammatory process that would release from hepatocytes and mononuclear phagocytes should be mainly confined to the pulp chamber, where the tooth inflammatory response takes place. If α1-antitrypsin in sound dentin prevented enzymatic degradation, similar effects might be expected against the proteolytic action of bacteria, which does not appear to be the case. This would depend upon the type of bacteria and the byproducts they were releasing.[17]

The effectiveness of papain in facilitating removal of carious dentin was described in studies by Bussadori et al., (2005), Corrêa et al.,[26] and Piva et al.[34]

The selective action of the papain gel in carious dentin is due to its inability to digest collagen fibrils that are protected by mineral in sound dentin, as opposed to the putative “protective” action of the α1-antitrypsin in intact collagen fibrils. Former studies have identified that papain promoted superficial degradation of Type I collagen fibrils and this was independent of previous partial degeneration from bacterial action.[17]

The mean Knoop hardness number value for the sound side in the present study was 68.06 ± 5.03 kgf/mm2. The mean microhardness of dentin in bur-treated side was 47.59 ± 7.39 kgf/mm2 and that for Carie-Care™-treated side was 28.45 ± 8.21 kgf/mm2. In the present study, when the mean microhardness values of both the types of treated dentin were compared with the estimated value on the sound side, it showed statistically significant difference (P < 0.05). Higher microhardness values for sound side than treated side were shown in former studies by Banerjee et al.,[35] Hossain et al.,[36] and Corrêa et al.[26] The present result also showed a reduction in mean microhardness for the caries-treated side.

Dental instrumentation results in a smear layer which cover the normal structural components of dentin and penetrates several micrometers into the tubules to form smear plugs.[37] This layer is composed of a mixture of partly denatured collagen and mineral.[3839] Studies done by Eric et al.[40] showed that bur cavity surface showed flat appearance and exhibited a debris-like smear layer that may interfere with adhesion, wetting, penetration, and hardness of the prepared cavity.[39] The studies by Wennerberg et al.,[41] Banerjee et al.,[35] and Hossain et al.[36] observed that dentinal surface after chemomechanical treatment (Carisolv™) was irregular and there remained a minimal debris-like smear layer and most of the dentinal tubules were opened.

CONCLUSION

In accordance with the results obtained in this study, it may be concluded that:

The rotary instrument group showed a consistent microhardness value with not much difference according to depth. The chemomechanical group showed a lesser microhardness value closer to the cavity floor than away from it

The microhardness values at all depths were significantly different for each treatment group with an increased value seen in the rotary group

The mean microhardness values of residual dentin in treated side were found to be insignificant when compared among each interval in each group

The microhardness of sound dentin had high significant difference from that of residual dentin in both the rotary group and the chemomechanical group.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.